Next Article in Journal
Contributions to Knowledge of the Dictyocaulus Infection of the Red Deer
Previous Article in Journal
Post-Slaughter Age Classification and Sex Determination in Deboned Beef Using Lipofuscin Autofluorescence and Amelogenin Gene Analysis
 
 
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Review

piRNAs as Potential Regulators of Mammary Gland Development and Pathology in Livestock

College of Veterinary Medicine, Basic Veterinary Medicine, Panhe Campus, Shandong Agricultural University, Taian 271018, China
*
Authors to whom correspondence should be addressed.
These authors contributed equally to this work.
Vet. Sci. 2025, 12(6), 594; https://doi.org/10.3390/vetsci12060594
Submission received: 29 April 2025 / Revised: 1 June 2025 / Accepted: 16 June 2025 / Published: 17 June 2025

Simple Summary

PIWI-interacting RNAs (piRNAs) have been demonstrated to maintain transposon silencing and regulate reproductive development and disease progression by binding to PIWI proteins. However, research on piRNAs in livestock animals is still in its infancy, with only preliminary studies available regarding their roles in the reproduction of swine, bovine, and ovine species. Given the regulatory mechanisms of piRNAs in mammary cancer and inflammation, as well as the established roles of other non-coding RNAs (ncRNAs) in mammary development, this paper focuses on the dairy cow mammary system to explore the potential functions of piRNAs in mammary development and mastitis. This review offers a novel perspective on mammary gland biology in livestock and provides a theoretical foundation for understanding the role of piRNAs in mammary gland development and associated diseases, representing significant value for both basic research and practical applications.

Abstract

PiRNAs are a subclass of non-coding RNAs, 26–31 nucleotides (nt) in length, that form regulatory complexes through their interaction with PIWI proteins. Studies in model organisms have demonstrated that piRNAs play crucial roles in tissue development and in predicting disease outcomes, positioning them as promising targets for developmental regulation and therapeutic intervention. In contrast, research on piRNAs in animal husbandry is still in its early stages and has not received sufficient attention. Despite this, the few studies available in livestock research have revealed that piRNAs serve as key regulators of reproductive development, underscoring their significant regulatory potential in farm animals and justifying further investigation. Accordingly, this review uses the bovine mammary gland as an exemplary case to summarize the progress in piRNA research related to mammary development and disease. The role of piRNAs in regulating breast cancer stem cell proliferation and modulating inflammatory progression is a highly active area of research. We hypothesize that piRNAs may play a potential role in regulating both mammary gland development and mastitis, making them promising targets for enhancing mammary development and overall health in dairy cattle and providing a theoretical foundation for further piRNA applications in animal husbandry.

Graphical Abstract

1. Introduction

The livestock farming industry confronts a dual challenge of global population growth and an increasing demand for food. This dual challenge entails enhancing production efficiency and adopting green, circular development practices. Consequently, this imposes increased demands on fundamental aspects such as genetic enhancement, disease prevention and control, and reproductive efficiency. In this context, epigenetic regulatory mechanisms have emerged as a promising research field with the potential to alleviate production bottlenecks in livestock, owing to their crucial roles in gene expression regulation and transgenerational inheritance [1].
The impact of epigenetic regulatory networks on key phenotypic traits—including growth performance, stress tolerance, and reproductive efficiency—in livestock is well documented. This regulation is mediated through DNA methylation, histone modifications, and ncRNAs, whose significance in livestock is increasingly recognized [2,3]. Among these, DNA methylation has been identified as a prominent feature of heat stress in dairy cows. It has been demonstrated that the exposure of bovines to elevated ambient temperatures during the period of gestation results in the occurrence of differential DNA methylation in utero, as well as intrauterine growth restriction of the offspring, which is partially responsible for long-term phenotypic alterations [4]. It was demonstrated that heat stress resulted in a significant increase in the methylation level of the Dual Leucine Zipper Kinase-like (DNLZ) promoter in dairy cows. This finding implies that the changes could be used as an epigenetic marker for heat tolerance traits and provide direction for breeding heat-resistant dairy cows [5]. In mastitis regulation, the presence of differentially expressed miRNAs in the milk exosome from healthy cows and cows with mastitis suggests that such molecules may serve as a significant research tool for studying the molecular mechanisms of mastitis in dairy cows [6]. Furthermore, differentially expressed lncRNAs were identified in the mammary tissues of cows in the early and non-lactating stages of lactation. Through pathway analyses, the authors showed that these differentially expressed lncRNAs may be involved in important signaling cascades and regulatory processes associated with immune responses, cell growth, and intracellular signaling [7]. Despite the present focus on DNA methylation and histone modification in livestock epigenetic studies, the importance of ncRNAs is becoming increasingly recognized as research in this area progresses. Consequently, a comprehensive understanding of ncRNAs is of great value for the development of animal husbandry.
NcRNAs include a variety of forms, such as microRNAs (miRNAs), small interfering RNAs (siRNAs), piRNAs, and long non-coding RNAs (lncRNAs), among others. Of particular note is the discovery of piRNAs in 2006 [8,9,10,11,12], which have been shown to maintain genome stability and regulate protein synthesis by binding to PIWI proteins [13,14]. In addition to this, piRNAs have been shown to play a crucial role in the maintenance of normal gonadal development [15]. Recently, the significance of piRNAs in inflammatory responses and cancer research, particularly in breast cancer, has gained increasing recognition. This nc RNA is emerging as potential prognostic markers and therapeutic targets for breast cancer [16,17].
At the present time, research into the mechanism of piRNAs has been primarily focused on model organisms such as nematodes, Drosophila, and mice. To date, studies pertaining to animal husbandry have exclusively encompassed a proportion of the expression profiles, and no in-depth mechanistic studies have been undertaken to date. The present focus of research in the field of piRNAs is on cancer and inflammation, where the role of these molecules is to regulate the proliferation of breast cancer cells [18], as well as to inhibit inflammatory signaling pathways and thus regulate inflammatory progression [19]. The studies of piRNAs in the specific inflammatory response of mastitis or in mammary development have not been found. In this paper, we present a systematic review of the production, characteristics, and functions of piRNAs, as well as their role in inflammation and breast cancer, and propose a hypothesis regarding their potential roles in mammary gland development and mastitis with the generalization of the inflammatory response and the characteristics of mammary gland development. Additionally, we provided a concise overview of the prospects for the utilization of piRNAs in domestic animals. To explore the potential roles of piRNAs in mammary development and disease through a review of their functions and mechanisms, providing new directions for research in livestock biology.

2. The Process of Generating piRNAs

As a class of ncRNAs, piRNAs exhibit a unique biogenesis pathway that encompasses both primary processing and secondary amplification (also known as the ping-pong cycle, a self-reinforcing amplification loop of piRNA production). The process of piRNA biogenesis is found to be identical in the majority of animal germ cells [20]. Notably, piRNA biogenesis occurs independently of the Dicer enzyme. Primary piRNAs are characterized by a single-stranded linear structure featuring a 5′-monophosphate and a 2′-O-methyl modification at the 3′ end [21]. In contrast, secondary piRNAs display a 10 nt complementarity at their 5′ ends with primary piRNAs [22]. This characteristic underpins the efficient repression of transposons by the piRNA pathway.

2.1. Primary Processing

Primary processing represents a crucial stage in piRNA biogenesis. The sources of piRNAs include transposon-derived piRNAs, lncRNA-derived piRNAs, and mRNA-derived piRNAs [22]. The primary processing of piRNAs is a complex process that involves multiple proteins, whose identities vary across species (Table 1). In Drosophila, the Rhino−Deadlock−Cutoff (RDC) complex, located in the nucleus, plays a pivotal role in recognizing heterochromatin regions. Subsequently, the RDC complex recruits the Moonshiner protein, activates RNA polymerase II (Pol II), and initiates the transcription of piRNA clusters from both genomic strands. These processes culminate in the production of either sense or antisense piRNA precursors (pre-piRNA) [23]. The pre-piRNA is subsequently cleaved by the 5′-terminal enzyme—Zucchini (Zuc) protein, typically at a uracil-enriched region, resulting in piRNAs with a strong 5′ uracil bias (1U bias) [24]. The cleaved precursor RNAs are trimmed by the 3′-terminal enzyme—Trimmer protein to yield single-stranded, linear piRNAs with a final length of 26–31 nt, thereby forming primary piRNAs [25,26,27,28].

2.2. Secondary Amplification

Secondary amplification, also known as the “ping-pong cycle,” represents a crucial stage in piRNA biogenesis, resulting in an increased production of piRNAs through synergistic interactions with diverse PIWI proteins [32,37]. In Drosophila, for instance, primary antisense piRNAs bind to the Aub protein to form the piRNA-Aub complex, which exhibits endonuclease activity. This complex subsequently targets the sense pre-piRNA through a process of base complementary pairing, then trimmed by exonucleases and modified by hua enhancer 1 (HEN1) methyltransferase, resulting in the formation of the secondary piRNA. The 5′ end of the secondary piRNA begins at the 10th nt from the cleavage site of the target RNA, which typically features an adenine residue (10A bias) [37,38,39]. The secondary piRNA then associates with the Ago3 protein, forming a complex that targets antisense pre-piRNA through base-pairing. The Ago3 complex cleaves the target RNA, thereby generating a new antisense pre-piRNA [40]. The 5′ end of this new antisense pre-piRNA begins at the 10th nt from the cleavage site—typically featuring a uracil residue—and, after exonuclease trimming and modification by HEN1 methyltransferase, matures into a new primary antisense piRNA [41]. The newly formed primary antisense piRNA then re-associates with the Aub protein, initiating another round of the ping-pong cycle; this cycle is represented schematically in Figure 1.

3. Characteristics and Functions of piRNAs

The principal function of piRNAs is believed to involve their binding to PIWI proteins, thereby forming functional piRNA–PIWI complexes. Their main functions include silencing transposons to ensure genomic stability, preserving fertility, and regulating disease development. In early studies, research focused on the mechanism of piRNA silencing of transposable factors at the transcriptional and post-transcriptional levels in animal germ cells. This is closely related to the regulatory function of reproductive development [22,42]. Subsequent evidence has shown that piRNAs can also regulate reproductive development by regulating the protein-coding genes involved in embryogenesis [43,44]. More recent studies have focused on the regulatory roles of piRNAs in inflammation and breast cancer, with their expression being modulated by environmental, nutritional, stress-related, pathological, and genetic factors, as well as by maternal transgenerational effects [45,46].

3.1. The Characteristics of piRNAs

Within the non-coding RNA families, piRNAs, miRNAs, and siRNAs exhibit significant disparities in their generation mechanisms, action proteins, and functional levels, despite their common classification as small molecule RNAs.
In contrast to the biogenesis of miRNAs and siRNAs, which is Dicer-dependent, piRNA generation occurs independently of Dicer. It is initiated from a dedicated piRNA cluster and non-cluster templates, resulting in the production of mature piRNAs (typically 26–31 nt in length) through primary processing and secondary amplification [22]. In contrast, miRNA biogenesis initiates with the transcription of primary miRNA transcripts (pri-miRNAs) by Pol II. These pri-miRNAs fold into characteristic hairpin structures within the nucleus, where they are recognized and cleaved by Drosha. This processing generates precursor miRNAs (pre-miRNAs), which are subsequently processed by the cytoplasmic RNase III enzyme Dicer into miRNA duplexes with a length of 21–24 nt. One strand of the duplex is selectively loaded onto AGO subfamily proteins to form the functional RNA-induced silencing complex (RISC) [47]. Similar to miRNAs, siRNAs originate from exogenous viral genomes or endogenous repetitive sequences, which are transcribed into long double-stranded RNAs (dsRNAs). These dsRNAs undergo precise cleavage by the ribonuclease Dicer, generating short RNA duplexes, which are 20–24 nt long. Following processing, one strand of the duplex (the guide strand) is selectively incorporated into AGO subfamily proteins within the RISC [48,49]. Secondly, it is important to note that the binding proteins of piRNAs are distinct from those of miRNAs and siRNAs. The silencing of small molecule RNAs has been observed to interact with Argonaute family proteins, which constitute the core part of RNA-induced silencing [50]. Argonaute proteins have been identified as multidomain proteins, primarily comprising the PAZ and PIWI domains, and encompassing both AGO subfamily and PIWI subfamily proteins [51,52]. It has been demonstrated that, in contrast to the interaction of miRNAs and siRNAs with AGO subfamily proteins, piRNAs interact with the PIWI subfamily proteins [53].
In conclusion, the core functions of piRNAs are distinct from those of miRNAs and siRNAs. The primary function of piRNAs is to maintain the stability of the germ cell genome, primarily through the process of transposon silencing. This process involves the modification of transposon genes at the transcriptional level, resulting in the suppression of their expression (e.g., DNA methylation and histone modification). Furthermore, at the post-transcriptional level, the process directly cleaves transposon-originated RNA molecules in order to regulate gene expression [54,55,56]. Meanwhile, miRNAs play a pivotal role in regulating gene expression at the post-transcriptional level. By binding to the 3′ untranslated region (3′UTR) of target mRNAs, miRNAs either inhibit translation via partial complementary pairing or promote mRNA deadenylation and subsequent degradation [57]. The siRNA is fully complementary to the target mRNA and functions by mediating RISC cleavage of the mRNA, resulting in degradation and subsequent gene silencing [58]. A comparative analysis reveals significant disparities among these classes (Figure 2). Notably, the investigation of piRNAs in livestock is a relatively recent development compared to research on other ncRNAs.

3.2. The Role of piRNAs in Silencing Transposons and Stabilizing Genomes

The piRNA pathway is critical for maintaining transposon silencing and sustaining gene stability, and is centrally dependent on the integrity of PIWI proteins. It has been demonstrated in model animals that piRNAs interact with PIWI proteins to generate recognition complexes which silence distinct transposons Tc3, GypsyDR1, long interspersed nuclear element-1 (L1), etc. [30,54]. This process is dependent on the RNA-dependent RNA polymerase (RdRPase) for the synthesis of 22-nucleotide guanine-starting RNA (22G-RNA), which in turn maintains long-term silencing through the worm-specific Argonaute (WAGO) pathway [59]. The absence of PIWI proteins (e.g., PRG-1, ZIWI, or PIWI) has been demonstrated to result in transposon activation, reduced germ cell numbers, and gonadal hypoplasia [31,38,60]. It has been established that certain proteins—defective P granules and sterile (DEPS-1), four ankyrin repeats, a sterile alpha motif, and leucine zipper 1 protein (Asz1), Maelstrom (Mael), etc.—are indispensable for the maintenance of complex structure or function, and their absence directly disrupts transposon repression [15,61,62].
The repression of transposons by piRNAs can be achieved through epigenetic mechanisms. It has been established that MILI and MIWI2, in conjunction with DNA methyltransferase 3-like (DNMT3L), are indispensable for the initiation of methylation processes. Furthermore, it has been determined that L1 and intracisternal A particle (IAP) play a pivotal role in this process. In the absence of MILI/MIWI2, there is a concomitant reduction in the level of transposon CpG methylation, resulting in the activation of the transposons [63,64]. 22G-RNA, in conjunction with the WAGO family protein heritable RNA interference (RNAi) defective-1 (HRDE-1) and the RNAi pathway protein nuclear RNAi defective-2 (NRDE-2), has been shown to collaborate in the recruitment of histone methyltransferase SET-25/32, thereby facilitating the catalysis of histone H3 lysine 9 trimethylation (H3K9me3) modification and, consequently, resulting in transcriptional repression through the establishment of heterochromatin structures by the heterochromatin protein like-2 (HPL-2). Concurrently, 22G-RNA has been demonstrated to directly mediate the degradation of target mRNAs and enhance the silencing effect [65]. Abnormal PIWI function has been demonstrated to reduce the level of H3K9me modification and heterochromatin protein 1 (HP1) enrichment, whilst concomitantly increasing the level of histone H3 lysine 4 methylation (H3K4me2/3) modification. This has been shown to inhibit chromatin formation in the transposons and its surrounding regions, thereby ultimately activating the transposons [66,67,68].
In addition to epigenetic regulation, it has been established that piRNAs are capable of obstructing the process of translation of transposon mRNAs through their capacity to bind to the 3′UTR region of the transposon mRNAs. Furthermore, it has been demonstrated that chromatin assembly factor 1 (CAF1) interacts with the MIWI/piRNA complex, thereby inducing the decay of the target mRNA [44], which further enhances the silencing effects on transposons. A significant number of studies have been conducted on model animals such as nematodes, Drosophila, and mice. However, there is a paucity of mechanistic studies of piRNAs in livestock animals.

3.3. Physiological Functions of piRNAs

The piRNA pathway is paramount for reproductive development, playing a critical role in safeguarding gametogenesis and maintaining fertility. At the level of chromosome dynamics, piRNAs play a pivotal role in assembling telomere protection complexes, thereby ensuring genomic stability [69]. Furthermore, the piRNA pathway directly influences chromosome condensation and segregation. In Drosophila germ cells, mitotic bodies composed of piRNA pathway proteins have been observed to bind to pericentromeric, piRNA-producing loci, thereby regulating condensing loading. Mutations in the piRNA pathway lead to aberrant condensing loading, causing delays in chromosome condensation and segregation defects [70]. The role of PIWI proteins in maintaining germline stem cells (GSCs) is underscored by observations showing that the deletion of PIWI results in reduced GSC numbers, whereas its overexpression enhances stem cell division [71]. Notably, mice deficient in the piRNA pathway protein Miwi2 exhibit a significant reduction in germ cell numbers with age [64]. The importance of fertility maintenance is further evidenced by studies in mice showing that mitochondrial phospholipase D (MitoPLD) gene dysfunction disrupts the piRNA biogenesis pathway, resulting in spermatogonial meiotic arrest [72]. In golden hamsters, piRNAs are essential for the development of spermatogonia and the formation of fertile oocytes [73]. Thus, the aberrant expression of genes associated with the piRNA pathway leads to the unscheduled activation of transposons in germ cells, meiotic arrest, impaired spermatogenesis and oogenesis, disrupted early embryonic development, and ultimately infertility. The above findings suggest that the piRNA pathway plays a pivotal role in preserving developmental potential and reproductive health.

3.4. Factors Regulating piRNAs

The expression of piRNAs is regulated by a multitude of factors. Recent studies have demonstrated in model animals that elevated temperatures convert Drosophila piRNA clusters from an inactive to an active state, leading to the generation of novel piRNAs. These novel piRNAs are characterized by enhanced stability and can be maternally inherited by subsequent generations [74]. Conversely, elevated temperatures have also been shown to decrease piRNA levels and reduce offspring fitness in C. elegans. However, following bacterial infection, the restoration of piRNA levels correlates with improved offspring fitness, suggesting that the piRNA pathway can dynamically respond to environmental signals and exert a lasting influence on progeny [75]. Altered nutritional levels modulate piRNA expression; for instance, differential piRNA profiles have been observed in the sperm of obese versus lean males [76]. Similarly, a Western-style diet has been found to alter piRNA expression in the testes of male mice [77], while a high-fat diet influences sperm piRNA profiles in both males and their offspring, subsequently impacting offspring metabolic function [78]. Furthermore, the present study demonstrates that short-term endurance training induces reversible changes in sperm piRNA expression [79]. Early traumatic stress has been shown to significantly downregulate piRNA cluster 110 in male mouse sperm [80]. Furthermore, microcystin–leucine–arginine (MC-LR) exposure has been reported to alter piRNA expression in the testes and prostate of offspring, leading to decreased testicular indices and prostate hyperplasia in male mice [45,81].
Consequently, fluctuations in environmental factors, stress levels, and nutritional intake can result in alterations in piRNA expression, which may be transmitted to subsequent generations. In the context of livestock production, environmental stressors—including nutritional imbalances and heat stress—often have detrimental effects on animal health and performance. Given the role of piRNAs in responding to environmental and nutritional cues, it is hypothesized that they may hold considerable potential for enhancing livestock productivity and resilience. We propose the following pivotal research questions: Can piRNAs be used as biomarkers of heat stress resilience in dairy cows? Can piRNA expression levels influence mammary gland development?

4. The Role of ncRNAs in Mammary Gland Development

The function of piRNAs as a class of ncRNAs in the development of the mammary gland remains to be elucidated; however, the role of other ncRNAs in this process has been the subject of recent research. In dairy farming, a cow’s economic value depends on its milk yield, which is driven by mammary gland development and lactation physiology. The mammary gland, a distinctive mammalian organ, displays diverse morphologies across different developmental stages—from embryonic morphogenesis and pubertal ductal expansion to gestational alveolar differentiation. Within livestock production systems, the extent of mammary gland development directly impacts the economic viability of the industry, as evidenced by strong positive correlations with key productivity metrics: lactation performance in mature dairy cows [82,83] and sow nursing capacity (reflected in piglet weaning weight) [84,85]. Notably, declining lactation performance and a high incidence of mastitis causing significant economic losses are challenges for the livestock industry. Therefore, research aimed at promoting mammary epithelial cell proliferation and differentiation, maintaining lactation homeostasis, and reducing mastitis through molecular regulation offers a promising avenue for addressing current challenges in the industry [86,87]. Within the ncRNA regulatory network, miRNAs have been shown to play a pivotal regulatory role in various aspects of animal development and cell differentiation [57]. In contrast, piRNAs have been identified as being essential for the formation of germ cells [73]. Although the precise functional mechanisms of piRNAs in mammary gland development remain to be fully elucidated, ample evidence demonstrates that miRNAs and lncRNAs in the ncRNA family play significant roles in mammary gland development and milk synthesis [88,89]. This underscores the multifaceted regulatory functions of the ncRNA family in mammary gland development.
As a prominent member of the short ncRNA family, miRNAs play a pivotal role in embryonic development [90], and their dysfunction has been associated with a range of diseases [91]. MiRNAs regulate gene expression through the sequence-specific recognition of the 3′ untranslated region (3′UTR) or other regulatory elements of target mRNAs. This regulatory mechanism exhibits unique network characteristics: a single miRNA can modulate the stability or translational efficiency of multiple target mRNAs, while a single mRNA may be concurrently regulated by several miRNAs, thereby orchestrating protein synthesis and signaling pathways [92]. Recent studies indicate that miRNAs serve as key regulators in the dynamic process of mammary gland development. The expression of miR-30b was found to be significantly increased in mice at the pubertal and maturity virgin stages. The overexpression of miR-30b resulted in reduced alveolar filling, smaller lumens, fewer lipid droplets, and structurally altered lipid droplets [93]. In addition, it was found that the process of mammary gland degradation was delayed. The expression of miR-31 increased progressively from the pubertal to the mature stages in mice. The inhibition of miR-31 expression resulted in a loss of mammary stem cells (MaSCS), fewer lipid droplets, and less alveolar tissue [94]. In goats, the expression of miR-103 was significantly higher at mid-lactation than at dry period. The overexpression of miR-103 promoted the accumulation of milk fat droplets and triglycerides in Goat Mammary Epithelial Cells (GMECs) [95]. Consequently, the expression patterns of miRNAs undergo characteristic changes throughout mammary gland development—including the formation of the embryonic mammary gland primordium, pubertal ductal morphogenesis, gestational alveolar proliferation, and functional maturation during lactation (Table 2). Collectively, these dynamic changes play a crucial regulatory role in every stage of mammary gland development. The livestock farming industry confronts a dual challenge of global population growth and an increasing demand for food. This dual challenge entails enhancing production efficiency and adopting green, circular development practices.
Although not as extensively studied as miRNAs, lncRNAs—another significant class of non-coding RNAs—have increasingly garnered research attention in recent years due to their essential roles in regulating mammary gland development. It has been demonstrated that the lncRNA SOX2OT harbors the transcription factor SOX2—one of the Yamanaka factors—which plays a pivotal role in embryonic development and is essential for maintaining the pluripotency of various stem cells. SOX2 has also been shown to be a key determinant [124]. Studies have demonstrated that steroid receptor RNA activator (SRA) significantly promotes the proliferation and differentiation of mammary epithelial cells through the synergistic activation of estrogen (ER) and progesterone (PR) receptors, while also triggering apoptosis. Furthermore, SRA enhances the progression of lobule–alveolar structures during pregnancy relative to control mice [89]. Zfas1 is expressed in developing a mammary gland after puberty, particularly in the epithelial cells of the ducts and alveoli during pregnancy. The knockdown of Zfas1 in HC11 cells has been shown to increase cellular proliferation, induce β-casein expression, and promote epithelial dome formation, suggesting that high Zfas1 expression in late gestational mammary glands may regulate proliferation and inhibit the terminal differentiation of alveolar cells [125].The H19 locus is regulated by estradiol and corticosterone, with high expression levels observed in alveolar cells during pregnancy and degeneration. H19 is developmentally regulated, exhibiting elevated transcript levels during both puberty and pregnancy [126]. Pregnancy-induced noncoding RNA (PINC) is a developmentally regulated lncRNA that is highly expressed in alveolar cells during pregnancy and in degenerating terminal ductal lobule-like structures during transplacental labor. PINC may inhibit the terminal differentiation of alveolar cells during pregnancy, thereby preventing the premature secretion of large quantities of milk [127,128].
Overall, ncRNAs play a pivotal role in the regulation of mammary gland development, with both miRNAs and lncRNAs having been demonstrated to exert a direct influence on this process. However, there remains a significant research gap regarding the specific mechanism of piRNAs in the development of the mammary gland. piRNAs have been demonstrated to perform distinctive functions in the context of reproductive development, and they also act as a member of ncRNAs. Consequently, it can be hypothesized that piRNAs may also be a pivotal factor influencing mammary gland development. Nevertheless, current research on piRNAs has been predominantly focused on model animals, and research on piRNAs in livestock is still limited.

5. Prospects for piRNAs in Livestock Animals

As members of ncRNAs, piRNAs play critical roles in numerous biological processes, including maintaining genomic stability, silencing transposons, and facilitating germ cell development. Recent research has revealed the potential significance of piRNAs in livestock species, with ongoing studies focusing on pigs, cattle, and sheep. Numerous investigations have characterized the expression profiles of piRNAs in various gonadal tissues across different developmental stages in livestock, offering novel insights into their roles in germ cell, embryonic, and overall gonadal development. In pigs, a “ping-pong” amplification loop of piRNAs has been proposed, suggesting that piRNAs repress transposons expression and regulate the post-transcriptional expression of multiple protein-coding genes critical for normal spermatogenesis, thereby enhancing our understanding of porcine spermatozoa development [129,130]. In cattle, hybrid male sterility (HMS) has been linked to promoter hypermethylation-induced silencing of PIWI/piRNA pathway genes. DNA methylation influences this pathway by affecting gene expression and the production of robust piRNAs during spermatogenesis, underscoring its central role in bovine HMS [131]. Studies have shown that piRNA expression in bovine frozen semen differs significantly between high-motility (HM) and low-motility (LM) sperm, suggesting that piRNAs may be involved in sperm development and overall fertility [132]. Additionally, the expression profiles of piRNAs in sheep ovaries during the luteal (LP) and follicular (FP) phases have been examined to provide a reference for understanding the role of ovarian piRNAs throughout the estrous cycle [133]. The results of the present study suggest that piRNAs play an integral role in domestic animals (Table 3). Moreover, analysis of milk exosomal ncRNAs has revealed the presence of 88 piRNAs of unknown function within milk, indicating a potential association between piRNAs and immune function [134]. Additionally, the antiviral defense function of piRNAs has been demonstrated in mosquito cells, and knocking down piRNA pathway proteins leads to the enhanced replication of the Semliki Forest virus, thereby underscoring the antiviral properties of the piRNA pathway. Consequently, the potential of piRNAs to enhance reproductive efficiency, optimize production performance, and augment disease resistance has been recognized. Compared with research on model organisms or humans, studies on piRNAs in animal husbandry are still in their infancy and warrant further investigation.

6. The Role of piRNAs in Breast Cancer

Although the precise mechanisms of piRNA action in mammary gland development remain ambiguous, recent analyses of piRNAs in human and mice have revealed defined endogenous piRNA expression patterns and mechanistic functions in breast cancer, with implications for regulatory control in mammary tissues [153]. Using small RNA sequencing facilitates the identification of differentially expressed piRNAs in both tumor and non-tumor tissues of breast cancer. For instance, Huang et al. reported that piR-4987, piR-20365, piR-20485, and piR-20582 are up-regulated in tumors, with elevated piR-4987 expression correlating with positive lymph node status [154]. In addition, Hashim et al. found that in breast cancer cells, piR-34377, piR-35407, and piR-36743 are up-regulated, whereas piR-36026, piR-36249, piR-36318, and piR-36712 are down-regulated—suggesting potential regulatory roles in the cell cycle, apoptosis, cell–cell interactions, and DNA replication and repair [155]. Other studies have identified differentially expressed piRNAs in various breast cancer types and following different treatments [156,157].
Further studies have revealed that piRNAs act through multiple pathways and clarified their molecular mechanisms. piR-651 is highly expressed in breast tumor tissues and cells, and binds to PIWIL2 to form a complex that promotes the DNMT1-mediated methylation of the phosphatase and tensin homolog (PTEN) promoter, which promotes cell proliferation and invasion and elevates MDM2, CKD4, and CyclinD1 protein levels, as well as inhibits apoptosis; the disruption of piR-651 results in the opposite effect [158]. Similarly, piR-823 has been shown to be upregulated in breast cancer cells and enhance the expression of stem cell regulators (OCT4, SOX2, KLF4, NANOG, h-TERT) and methyltransferases (DNMT1, DNMT3A, DNMT3B), thereby promoting the hypermethylation of the adenomatous polyposis coli (APC) promoter, activating Wnt signaling, and driving tumor growth—whereas piR-823 knockdown, which also increases ERα and decreases h-TERT expression via the inhibition of the PI3K/Akt/mTOR pathway, suppresses cancer cell proliferation [159,160]. In addition, piR-2158 is downregulated in human and rodent breast cancer tumors and inhibits interleukin-11 (IL-11) expression by competing with FOS-related antigen 1 (FOSL1), inactivates the JAK/STAT pathway, and suppresses cell proliferation, migration, epithelial–mesenchymal transition (EMT), stemness, and angiogenesis [18]. Collectively, these findings underscore the regulatory role of piRNAs in breast cancer (Table 4).
Given the above outlined mechanisms, piRNAs have shown a great deal of promise for therapeutic use. The use of piR-2158 nanoparticles as a therapeutic target for the treatment of breast cancer has been validated by their demonstrated significant inhibitory effects on the growth of mouse breast cancer tumors [18]. Additionally, piRNAs can be utilized as a potential molecular staging and prognosis tool and has been shown to be closely associated with breast cancer metastasis, staging, and response to treatment. Consequently, piRNAs may serve as both prognostic markers and therapeutic targets for breast cancer. However, more study is still required to accurately diagnose piRNAs and develop targeted therapies because our present understanding of the precise molecular pathways behind their significance in breast cancer is still restricted.
Beyond oncogenesis, numerous studies have highlighted critical roles for piRNAs in maintaining genome stability, regulating stem cell function, and modulating molecular signaling pathways. Many studies have demonstrated the pivotal role of the piRNA pathway in the maintenance, division, and differentiation of GSCs [172,173]. These findings offer important clues to their physiological roles in normal mammary gland development. Mammary gland development is a complex, multi-stage process encompassing the embryonic formation of the mammary anlage, pubertal ductal branching, and gestational differentiation of glandular structures [174]. The central roles of stem cells, hormonal stimulation, and signaling pathways underpin both normal development and breast cancer pathogenesis. Indeed, mammary stem cell MaSCs maintain tissue homeostasis and regeneration through self-renewal, whereas breast cancer stem cells (BCSCs) drive tumor growth and recurrence [174,175,176]. Moreover, both mammary development and breast cancer have been associated with the activation of key pathways—including Wnt, Notch, and Hedgehog—and with the pivotal influence of the estrogen receptor ERα in mammary morphogenesis and tumorigenesis [177,178,179]. These correspondences suggest that the physiological development of breast tissue shares common molecular characteristics with the pathological progression of breast cancer; accordingly, it is plausible that piRNAs contribute to the regulation of cell proliferation, differentiation, and homeostasis in normal mammary gland development.

7. The Role of piRNAs in Inflammation

The inflammatory response is characterized by the activation of immune cells, such as dendritic cells, neutrophils, and mast cells [180], and the release of pro-inflammatory factors, including tumor necrosis factor-alpha (TNF-α), IL-1, and IL-6 [181]. The role of non-coding RNAs (ncRNAs) in the regulation of inflammation has become a subject of considerable interest in the research community [182]. Among them, piRNA, as a significant member of the ncRNA family, is progressively being revealed to possess an inflammatory regulatory function in addition to its role in breast cancer. It has been shown that piRNAs are involved in the regulation of inflammatory responses through a multilevel molecular mechanism. First, piRNAs can play a regulatory role by directly acting on key inflammatory signaling pathways. For example, pir-has-216911 binds to Toll-like Receptors 4(TLR4) mRNA and suppresses the TLR4/NF-κB/NLRP3 inflammatory signaling pathway, thereby inhibiting caspase-1–induced activation of GSDMD and reducing the pro-inflammatory effects of pyroptosis [183]. Similarly, piR-112710 directly binds to the 3′UTR of the thioredoxin-interacting protein (Txnip), suppressing its expression and inactivating the Txnip/NLRP3 pathway; this leads to reduced levels of pro-inflammatory factors (IL-18 and IL-1β) and the lower expression of proteins related to inflammasome activation (NLRP3, caspase-1, and GSDMD-N) [19,184]. Furthermore, hsa_piR_019949 is significantly down-regulated in response to IL-1β, and it may modulate the inflammatory response by repressing the expression of the lncRNA NEAT1—which in turn lowers NLRP3 levels and regulates the NOD-like receptor signaling pathway [185]. Secondly, it has been demonstrated that certain piRNAs regulate inflammation-related genes through epigenetic modification pathways. It was found that piRNA-6426 has been shown to inhibit inflammation by increasing methylation at the SOAT1 promoter via the recruitment of DNMT3B, which reduces the secretion of IL-1β and TNF-α and ameliorates the inflammatory microenvironment in heart failure [186]. This epigenetic regulatory mechanism provides a new direction for the treatment of inflammatory diseases. Thirdly, in disparate cell types, piRNAs manifested particular regulatory functions. In endothelial cells, rno-piR-017330 is up-regulated in response to TNF-α stimulation, suggesting that piRNAs contribute to the regulation of inflammation in these cells [187]. In contrast, in chondrocytes, IL-1β promotes the expression of piRNA mmu_piR_037459, suggesting that inflammatory factors may drive osteoarthritis (OA) pathology by regulating piRNA levels [188]. Finally, clinical studies have revealed correlations between piRNA expression profiles and disease states. Saha et al. identified 19 differentially expressed piRNAs in the plasma of patients with chronic pancreatitis (CP) compared to healthy individuals [189]. This discrepancy in expression indicates that piRNA may be of pathophysiological significance in inflammatory diseases. It can thus be concluded that piRNAs also have an important role in the inflammatory response (Table 5).
Although the mechanisms by which piRNAs regulate inflammatory responses are gradually being elucidated, their specific role in mastitis—a common inflammatory condition of the mammary gland—remains unreported. Mastitis exhibits pathological similarities to other systemic or organ-specific inflammatory diseases (e.g., arthritis, pneumonia), providing a useful theoretical foundation for investigating shared mechanisms. The following three lines of evidence were used to formulate a hypothesis regarding the potential involvement of piRNAs in the regulation of mastitis in dairy cows through key pathways such as TLR4/NF-κB: First, mastitis and other inflammatory diseases share a similar immune microenvironment, characterized by the infiltration of inflammatory cells such as monocytes, dendritic cells, and macrophages that release reactive oxygen species (ROS) and proteases, thereby exacerbating tissue damage [192]. Second, key pro-inflammatory cytokines—namely, TNF-α, IL-1β, and IL-6—are elevated in mastitis, driving the inflammatory response and contributing to tissue destruction [71,193]. Third, mastitis shares essential inflammatory pathways such as NF-κB, the NLRP3 inflammasome, and MAPK with other inflammatory conditions [194,195,196]. Therefore, although the role of piRNAs in mastitis remains ambiguous, their established functions in regulating cytokine secretion, NLRP3 inflammasome expression, and NF-κB signaling in other inflammatory disorders suggest that piRNAs may similarly influence mastitis progression.
In light of the functions of piRNAs in breast cancer progression and inflammatory responses, a comprehensive analysis was conducted regarding the potential roles of piRNAs in mammary gland development and mastitis (Figure 3). This investigation offers new insights into the advancement of animal husbandry.

8. Conclusions

‘Livestock are economically vital’ is punchier, providing a stable, high-quality supply of meat, eggs, and milk, while its scientific breeding mode is directly related to environmental safety, disease prevention, and biosecurity [197,198]. Despite their importance, research into the roles of piRNAs in livestock remains in its infancy, and systematic investigations are scarce. In this paper, we demonstrate that piRNAs are involved in numerous biological processes—including gamete formation, embryonic development, and disease regulation. Moreover, we propose that these small ncRNAs may influence livestock growth performance, disease resistance, and environmental adaptation through epigenetic mechanisms. At present, however, we are still unable to answer the questions “Can piRNAs regulate mammary gland development and can piRNAs be used as markers for mastitis in dairy cows?” Consequently, a comprehensive analysis of the molecular mechanisms underlying piRNA function in livestock is imperative to optimize breeding strategies, mitigate disease risks, and cultivate high-quality breeds—ultimately contributing to the industrial upgrading and high-quality development of animal husbandry. Notably, this study innovatively suggests that piRNAs might regulate the mammary developmental cycle and the pathological process of mastitis, offering new perspectives on enhancing lactation performance and developing innovative disease prevention and control strategies, which advance the development of dairy and animal husbandry in response to the dual challenges of a growing global population and increasing food demand.

Author Contributions

W.Y. and Z.Z.: writing—original draft. W.Y., Z.Z., and X.D.: writing—review and editing and literature search. X.D. and Q.H.: proofreading and supervision: X.D., Q.H., and Z.W.: resources and funding. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Cattle Agro-industry Technology Research System of Shandong Province (SDAIT-09-20), China Postdoctoral Science Foundation (2024M761856), and Shandong Postdoctoral Science Foundation (SDBX202302020).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding authors.

Conflicts of Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this review, and there is no professional or other personal interest of any nature or kind in any product, service, and/or company that could be construed as influencing the content of this review.

References

  1. Thompson, R.P.; Nilsson, E.; Skinner, M.K. Environmental epigenetics and epigenetic inheritance in domestic farm animals. Anim. Reprod. Sci. 2020, 220, 106316. [Google Scholar] [CrossRef] [PubMed]
  2. Ibeagha-Awemu, E.M.; Khatib, H. Epigenetics of livestock health, production, and breeding. In Handbook of Epigenetics; Elsevier: Amsterdam, The Netherlands, 2023; pp. 569–610. [Google Scholar]
  3. Do, D.N.; Suravajhala, P. Editorial: Role of Non-Coding RNAs in Animals. Animals 2023, 13, 805. [Google Scholar] [CrossRef] [PubMed]
  4. Ouellet, V.; Boucher, A.; Dahl, G.E.; Laporta, J. Consequences of maternal heat stress at different stages of embryonic and fetal development on dairy cows’ progeny. Anim. Front. 2021, 11, 48–56. [Google Scholar] [CrossRef]
  5. Yang, Y.; Chen, Y.; Hu, L.; Zhang, C.; Chen, G.; Hou, L.; Xu, Q.; Wang, Y.; Li, M. Molecular regulation of whole genome DNA methylation in heat stress response of dairy cows. BMC Genom. 2025, 26, 464. [Google Scholar] [CrossRef]
  6. Stefanon, B.; Cintio, M.; Sgorlon, S.; Scarsella, E.; Licastro, D.; Zecconi, A.; Colitti, M. Regulatory Role of microRNA of Milk Exosomes in Mastitis of Dairy Cows. Animals 2023, 13, 821. [Google Scholar] [CrossRef]
  7. Ghulam Mohyuddin, S.; Liang, Y.; Xia, Y.; Wang, M.; Zhang, H.; Li, M.; Yang, Z.; Niel, A.K.; Mao, Y. Identification and Classification of Long Non-Coding RNAs in the Mammary Gland of the Holstein Cow. Int. J. Mol. Sci. 2023, 24, 13585. [Google Scholar] [CrossRef]
  8. Vagin, V.V.; Sigova, A.; Li, C.; Seitz, H.; Gvozdev, V.; Zamore, P.D. A distinct small RNA pathway silences selfish genetic elements in the germline. Science 2006, 313, 320–324. [Google Scholar] [CrossRef] [PubMed]
  9. Aravin, A.; Gaidatzis, D.; Pfeffer, S.; Lagos-Quintana, M.; Landgraf, P.; Iovino, N.; Morris, P.; Brownstein, M.J.; Kuramochi-Miyagawa, S.; Nakano, T.; et al. A novel class of small RNAs bind to MILI protein in mouse testes. Nature 2006, 442, 203–207. [Google Scholar] [CrossRef]
  10. Girard, A.; Sachidanandam, R.; Hannon, G.J.; Carmell, M.A. A germline-specific class of small RNAs binds mammalian Piwi proteins. Nature 2006, 442, 199–202. [Google Scholar] [CrossRef]
  11. Grivna, S.T.; Beyret, E.; Wang, Z.; Lin, H. A novel class of small RNAs in mouse spermatogenic cells. Genes Dev. 2006, 20, 1709–1714. [Google Scholar] [CrossRef]
  12. Lau, N.C.; Seto, A.G.; Kim, J.; Kuramochi-Miyagawa, S.; Nakano, T.; Bartel, D.P.; Kingston, R.E. Characterization of the piRNA complex from rat testes. Science 2006, 313, 363–367. [Google Scholar] [CrossRef] [PubMed]
  13. Zhang, H.; Ren, Y.; Xu, H.; Pang, D.; Duan, C.; Liu, C. The expression of stem cell protein Piwil2 and piR-932 in breast cancer. Surg. Oncol. 2013, 22, 217–223. [Google Scholar] [CrossRef] [PubMed]
  14. Chuma, S.; Nakano, T. piRNA and spermatogenesis in mice. Philos. Trans. R. Soc. Lond. B Biol. Sci. 2013, 368, 20110338. [Google Scholar] [CrossRef]
  15. Ahmad, A.; Bogoch, Y.; Shvaizer, G.; Guler, N.; Levy, K.; Elkouby, Y.M. The piRNA protein Asz1 is essential for germ cell and gonad development in zebrafish and exhibits differential necessities in distinct types of germ granules. PLoS Genet. 2025, 21, e1010868. [Google Scholar] [CrossRef]
  16. Chalbatani, G.M.; Dana, H.; Memari, F.; Gharagozlou, E.; Ashjaei, S.; Kheirandish, P.; Marmari, V.; Mahmoudzadeh, H.; Mozayani, F.; Maleki, A.R.; et al. Biological function and molecular mechanism of piRNA in cancer. Pract. Lab. Med. 2019, 13, e00113. [Google Scholar] [CrossRef]
  17. Trzybulska, D.; Vergadi, E.; Tsatsanis, C. miRNA and Other Non-Coding RNAs as Promising Diagnostic Markers. Ejifcc 2018, 29, 221–226. [Google Scholar] [PubMed]
  18. Zhao, Q.; Qian, L.; Guo, Y.; Lü, J.; Li, D.; Xie, H.; Wang, Q.; Ma, W.; Liu, P.; Liu, Y.; et al. IL11 signaling mediates piR-2158 suppression of cell stemness and angiogenesis in breast cancer. Theranostics 2023, 13, 2337–2349. [Google Scholar] [CrossRef]
  19. Jiao, A.; Liu, H.; Wang, H.; Yu, J.; Gong, L.; Zhang, H.; Fu, L. piR112710 attenuates diabetic cardiomyopathy through inhibiting Txnip/NLRP3-mediated pyroptosis in db/db mice. Cell. Signal. 2024, 122, 111333. [Google Scholar] [CrossRef]
  20. Gainetdinov, I.; Colpan, C.; Arif, A.; Cecchini, K.; Zamore, P.D. A Single Mechanism of Biogenesis, Initiated and Directed by PIWI Proteins, Explains piRNA Production in Most Animals. Mol. Cell 2018, 71, 775–790. [Google Scholar] [CrossRef]
  21. Zhang, Y.; Liu, W.; Li, R.; Gu, J.; Wu, P.; Peng, C.; Ma, J.; Wu, L.; Yu, Y.; Huang, Y. Structural insights into the sequence-specific recognition of Piwi by Drosophila Papi. Proc. Natl. Acad. Sci. USA 2018, 115, 3374–3379. [Google Scholar] [CrossRef]
  22. Iwasaki, Y.W.; Siomi, M.C.; Siomi, H. PIWI-Interacting RNA: Its Biogenesis and Functions. Annu. Rev. Biochem. 2015, 84, 405–433. [Google Scholar] [CrossRef] [PubMed]
  23. Liu, Y.; Zhang, J.; Li, A.; Liu, Z.; He, Z.; Yuan, X.; Tuo, S. Prediction of cancer-associated piRNA–mRNA and piRNA–lncRNA interactions by integrated analysis of expression and sequence data. Tsinghua Sci. Technol. 2018, 23, 115–125. [Google Scholar] [CrossRef]
  24. Goriaux, C.; Desset, S.; Renaud, Y.; Vaury, C.; Brasset, E. Transcriptional properties and splicing of the flamenco piRNA cluster. EMBO Rep. 2014, 15, 411–418. [Google Scholar] [CrossRef] [PubMed]
  25. Ipsaro, J.J.; Haase, A.D.; Knott, S.R.; Joshua-Tor, L.; Hannon, G.J. The structural biochemistry of Zucchini implicates it as a nuclease in piRNA biogenesis. Nature 2012, 491, 279–283. [Google Scholar] [CrossRef]
  26. Zhang, R.; Tu, Y.X.; Ye, D.; Gu, Z.; Chen, Z.X.; Sun, Y. A Germline-Specific Regulator of Mitochondrial Fusion is Required for Maintenance and Differentiation of Germline Stem and Progenitor Cells. Adv. Sci. 2022, 9, 2203631. [Google Scholar] [CrossRef]
  27. Saito, K.; Ishizu, H.; Komai, M.; Kotani, H.; Kawamura, Y.; Nishida, K.M.; Siomi, H.; Siomi, M.C. Roles for the Yb body components Armitage and Yb in primary piRNA biogenesis in Drosophila. Genes Dev. 2010, 24, 2493–2498. [Google Scholar] [CrossRef]
  28. Andersen, P.R.; Tirian, L.; Vunjak, M.; Brennecke, J. A heterochromatin-dependent transcription machinery drives piRNA expression. Nature 2017, 549, 54–59. [Google Scholar] [CrossRef]
  29. Pastore, B.; Hertz, H.L.; Tang, W. Comparative analysis of piRNA sequences, targets and functions in nematodes. RNA Biol. 2022, 19, 1276–1292. [Google Scholar] [CrossRef]
  30. Houwing, S.; Kamminga, L.M.; Berezikov, E.; Cronembold, D.; Girard, A.; van den Elst, H.; Filippov, D.V.; Blaser, H.; Raz, E.; Moens, C.B.; et al. A role for Piwi and piRNAs in germ cell maintenance and transposon silencing in Zebrafish. Cell 2007, 129, 69–82. [Google Scholar] [CrossRef]
  31. Houwing, S.; Berezikov, E.; Ketting, R.F. Zili is required for germ cell differentiation and meiosis in zebrafish. EMBO J. 2008, 27, 2702–2711. [Google Scholar] [CrossRef]
  32. Brennecke, J.; Aravin, A.A.; Stark, A.; Dus, M.; Kellis, M.; Sachidanandam, R.; Hannon, G.J. Discrete small RNA-generating loci as master regulators of transposon activity in Drosophila. Cell 2007, 128, 1089–1103. [Google Scholar] [CrossRef] [PubMed]
  33. Hirakata, S.; Siomi, M.C. piRNA biogenesis in the germline: From transcription of piRNA genomic sources to piRNA maturation. Biochim. Biophys. Acta 2016, 1859, 82–92. [Google Scholar] [CrossRef] [PubMed]
  34. Grivna, S.T.; Pyhtila, B.; Lin, H. MIWI associates with translational machinery and PIWI-interacting RNAs (piRNAs) in regulating spermatogenesis. Proc. Natl. Acad. Sci. USA 2006, 103, 13415–13420. [Google Scholar] [CrossRef]
  35. Watanabe, T.; Takeda, A.; Tsukiyama, T.; Mise, K.; Okuno, T.; Sasaki, H.; Minami, N.; Imai, H. Identification and characterization of two novel classes of small RNAs in the mouse germline: Retrotransposon-derived siRNAs in oocytes and germline small RNAs in testes. Genes Dev. 2006, 20, 1732–1743. [Google Scholar] [CrossRef] [PubMed]
  36. Ding, D.; Liu, J.; Dong, K.; Midic, U.; Hess, R.A.; Xie, H.; Demireva, E.Y.; Chen, C. PNLDC1 is essential for piRNA 3′ end trimming and transposon silencing during spermatogenesis in mice. Nat. Commun. 2017, 8, 819. [Google Scholar] [CrossRef]
  37. Saito, K.; Sakaguchi, Y.; Suzuki, T.; Suzuki, T.; Siomi, H.; Siomi, M.C. Pimet, the Drosophila homolog of HEN1, mediates 2′-O-methylation of Piwi- interacting RNAs at their 3’ ends. Genes Dev. 2007, 21, 1603–1608. [Google Scholar] [CrossRef]
  38. Gunawardane, L.S.; Saito, K.; Nishida, K.M.; Miyoshi, K.; Kawamura, Y.; Nagami, T.; Siomi, H.; Siomi, M.C. A slicer-mediated mechanism for repeat-associated siRNA 5′ end formation in Drosophila. Science 2007, 315, 1587–1590. [Google Scholar] [CrossRef]
  39. Olivieri, D.; Senti, K.A.; Subramanian, S.; Sachidanandam, R.; Brennecke, J. The cochaperone shutdown defines a group of biogenesis factors essential for all piRNA populations in Drosophila. Mol. Cell 2012, 47, 954–969. [Google Scholar] [CrossRef]
  40. Nagao, A.; Mituyama, T.; Huang, H.; Chen, D.; Siomi, M.C.; Siomi, H. Biogenesis pathways of piRNAs loaded onto AGO3 in the Drosophila testis. RNA 2010, 16, 2503–2515. [Google Scholar] [CrossRef]
  41. Horwich, M.D.; Li, C.; Matranga, C.; Vagin, V.; Farley, G.; Wang, P.; Zamore, P.D. The Drosophila RNA methyltransferase, DmHen1, modifies germline piRNAs and single-stranded siRNAs in RISC. Curr. Biol. 2007, 17, 1265–1272. [Google Scholar] [CrossRef]
  42. Ozata, D.M.; Gainetdinov, I.; Zoch, A.; O’Carroll, D.; Zamore, P.D. PIWI-interacting RNAs: Small RNAs with big functions. Nat. Rev. Genet. 2019, 20, 89–108. [Google Scholar] [CrossRef] [PubMed]
  43. Rouget, C.; Papin, C.; Boureux, A.; Meunier, A.C.; Franco, B.; Robine, N.; Lai, E.C.; Pelisson, A.; Simonelig, M. Maternal mRNA deadenylation and decay by the piRNA pathway in the early Drosophila embryo. Nature 2010, 467, 1128–1132. [Google Scholar] [CrossRef] [PubMed]
  44. Gou, L.T.; Dai, P.; Yang, J.H.; Xue, Y.; Hu, Y.P.; Zhou, Y.; Kang, J.Y.; Wang, X.; Li, H.; Hua, M.M.; et al. Pachytene piRNAs instruct massive mRNA elimination during late spermiogenesis. Cell Res. 2014, 24, 680–700. [Google Scholar] [CrossRef]
  45. Zhang, L.; Zhang, H.; Zhang, H.; Benson, M.; Han, X.; Li, D. Roles of piRNAs in microcystin-leucine-arginine (MC-LR) induced reproductive toxicity in testis on male offspring. Food Chem. Toxicol. 2017, 105, 177–185. [Google Scholar] [CrossRef]
  46. Donkin, I.; Barrès, R. Sperm epigenetics and influence of environmental factors. Mol. Metab. 2018, 14, 1–11. [Google Scholar] [CrossRef] [PubMed]
  47. Bartel, D.P. Metazoan MicroRNAs. Cell 2018, 173, 20–51. [Google Scholar] [CrossRef]
  48. Santos, D.; Mingels, L.; Vogel, E.; Wang, L.; Christiaens, O.; Cappelle, K.; Wynant, N.; Gansemans, Y.; Van Nieuwerburgh, F.; Smagghe, G.; et al. Generation of Virus- and dsRNA-Derived siRNAs with Species-Dependent Length in Insects. Viruses 2019, 11, 738. [Google Scholar] [CrossRef]
  49. Liu, Q.; Ding, C.; Chu, Y.; Zhang, W.; Guo, G.; Chen, J.; Su, X. Pln24NT: A web resource for plant 24-nt siRNA producing loci. Bioinformatics 2017, 33, 2065–2067. [Google Scholar] [CrossRef]
  50. Swarts, D.C.; Makarova, K.; Wang, Y.; Nakanishi, K.; Ketting, R.F.; Koonin, E.V.; Patel, D.J.; van der Oost, J. The evolutionary journey of Argonaute proteins. Nat. Struct. Mol. Biol. 2014, 21, 743–753. [Google Scholar] [CrossRef]
  51. Tolia, N.H.; Joshua-Tor, L. Slicer and the argonautes. Nat. Chem. Biol. 2007, 3, 36–43. [Google Scholar] [CrossRef]
  52. Peters, L.; Meister, G. Argonaute proteins: Mediators of RNA silencing. Mol. Cell 2007, 26, 611–623. [Google Scholar] [CrossRef] [PubMed]
  53. Xiao, Y.; Ke, A. PIWI Takes a Giant Step. Cell 2016, 167, 310–312. [Google Scholar] [CrossRef] [PubMed]
  54. Aravin, A.A.; Sachidanandam, R.; Girard, A.; Fejes-Toth, K.; Hannon, G.J. Developmentally regulated piRNA clusters implicate MILI in transposon control. Science 2007, 316, 744–747. [Google Scholar] [CrossRef]
  55. Nishida, K.M.; Saito, K.; Mori, T.; Kawamura, Y.; Nagami-Okada, T.; Inagaki, S.; Siomi, H.; Siomi, M.C. Gene silencing mechanisms mediated by Aubergine piRNA complexes in Drosophila male gonad. RNA 2007, 13, 1911–1922. [Google Scholar] [CrossRef]
  56. Han, H.; Fan, G.; Song, S.; Jiang, Y.; Qian, C.; Zhang, W.; Su, Q.; Xue, X.; Zhuang, W.; Li, B. piRNA-30473 contributes to tumorigenesis and poor prognosis by regulating m6A RNA methylation in DLBCL. Blood 2021, 137, 1603–1614. [Google Scholar] [CrossRef]
  57. Gebert, L.F.R.; MacRae, I.J. Regulation of microRNA function in animals. Nat. Rev. Mol. Cell Biol. 2019, 20, 21–37. [Google Scholar] [CrossRef] [PubMed]
  58. Dana, H.; Chalbatani, G.M.; Mahmoodzadeh, H.; Karimloo, R.; Rezaiean, O.; Moradzadeh, A.; Mehmandoost, N.; Moazzen, F.; Mazraeh, A.; Marmari, V.; et al. Molecular Mechanisms and Biological Functions of siRNA. Int. J. Biomed. Sci. 2017, 13, 48–57. [Google Scholar] [CrossRef]
  59. Lee, H.C.; Gu, W.; Shirayama, M.; Youngman, E.; Conte, D., Jr.; Mello, C.C. C. elegans piRNAs mediate the genome-wide surveillance of germline transcripts. Cell 2012, 150, 78–87. [Google Scholar] [CrossRef]
  60. Batista, P.J.; Ruby, J.G.; Claycomb, J.M.; Chiang, R.; Fahlgren, N.; Kasschau, K.D.; Chaves, D.A.; Gu, W.; Vasale, J.J.; Duan, S.; et al. PRG-1 and 21U-RNAs interact to form the piRNA complex required for fertility in C. elegans. Mol. Cell 2008, 31, 67–78. [Google Scholar] [CrossRef]
  61. Suen, K.M.; Braukmann, F.; Butler, R.; Bensaddek, D.; Akay, A.; Lin, C.C.; Milonaitytė, D.; Doshi, N.; Sapetschnig, A.; Lamond, A.; et al. DEPS-1 is required for piRNA-dependent silencing and PIWI condensate organisation in Caenorhabditis elegans. Nat. Commun. 2020, 11, 4242. [Google Scholar] [CrossRef]
  62. Sienski, G.; Dönertas, D.; Brennecke, J. Transcriptional silencing of transposons by Piwi and maelstrom and its impact on chromatin state and gene expression. Cell 2012, 151, 964–980. [Google Scholar] [CrossRef] [PubMed]
  63. Kuramochi-Miyagawa, S.; Watanabe, T.; Gotoh, K.; Totoki, Y.; Toyoda, A.; Ikawa, M.; Asada, N.; Kojima, K.; Yamaguchi, Y.; Ijiri, T.W.; et al. DNA methylation of retrotransposon genes is regulated by Piwi family members MILI and MIWI2 in murine fetal testes. Genes Dev. 2008, 22, 908–917. [Google Scholar] [CrossRef]
  64. Carmell, M.A.; Girard, A.; van de Kant, H.J.; Bourc’his, D.; Bestor, T.H.; de Rooij, D.G.; Hannon, G.J. MIWI2 is essential for spermatogenesis and repression of transposons in the mouse male germline. Dev. Cell 2007, 12, 503–514. [Google Scholar] [CrossRef]
  65. Ashe, A.; Sapetschnig, A.; Weick, E.M.; Mitchell, J.; Bagijn, M.P.; Cording, A.C.; Doebley, A.L.; Goldstein, L.D.; Lehrbach, N.J.; Le Pen, J.; et al. piRNAs can trigger a multigenerational epigenetic memory in the germline of C. elegans. Cell 2012, 150, 88–99. [Google Scholar] [CrossRef] [PubMed]
  66. Klenov, M.S.; Sokolova, O.A.; Yakushev, E.Y.; Stolyarenko, A.D.; Mikhaleva, E.A.; Lavrov, S.A.; Gvozdev, V.A. Separation of stem cell maintenance and transposon silencing functions of Piwi protein. Proc. Natl. Acad. Sci. USA 2011, 108, 18760–18765. [Google Scholar] [CrossRef] [PubMed]
  67. Le Thomas, A.; Rogers, A.K.; Webster, A.; Marinov, G.K.; Liao, S.E.; Perkins, E.M.; Hur, J.K.; Aravin, A.A.; Tóth, K.F. Piwi induces piRNA-guided transcriptional silencing and establishment of a repressive chromatin state. Genes Dev. 2013, 27, 390–399. [Google Scholar] [CrossRef]
  68. Pezic, D.; Manakov, S.A.; Sachidanandam, R.; Aravin, A.A. piRNA pathway targets active LINE1 elements to establish the repressive H3K9me3 mark in germ cells. Genes Dev. 2014, 28, 1410–1428. [Google Scholar] [CrossRef]
  69. Khurana, J.S.; Xu, J.; Weng, Z.; Theurkauf, W.E. Distinct functions for the Drosophila piRNA pathway in genome maintenance and telomere protection. PLoS Genet. 2010, 6, e1001246. [Google Scholar] [CrossRef]
  70. Pek, J.W.; Kai, T. DEAD-box RNA helicase Belle/DDX3 and the RNA interference pathway promote mitotic chromosome segregation. Proc. Natl. Acad. Sci. USA 2011, 108, 12007–12012. [Google Scholar] [CrossRef]
  71. Cox, D.N.; Chao, A.; Lin, H. piwi encodes a nucleoplasmic factor whose activity modulates the number and division rate of germline stem cells. Development 2000, 127, 503–514. [Google Scholar] [CrossRef]
  72. Huang, H.; Gao, Q.; Peng, X.; Choi, S.Y.; Sarma, K.; Ren, H.; Morris, A.J.; Frohman, M.A. piRNA-associated germline nuage formation and spermatogenesis require MitoPLD profusogenic mitochondrial-surface lipid signaling. Dev. Cell 2011, 20, 376–387. [Google Scholar] [CrossRef] [PubMed]
  73. Loubalova, Z.; Fulka, H.; Horvat, F.; Pasulka, J.; Malik, R.; Hirose, M.; Ogura, A.; Svoboda, P. Formation of spermatogonia and fertile oocytes in golden hamsters requires piRNAs. Nat. Cell Biol. 2021, 23, 992–1001. [Google Scholar] [CrossRef] [PubMed]
  74. Casier, K.; Delmarre, V.; Gueguen, N.; Hermant, C.; Viodé, E.; Vaury, C.; Ronsseray, S.; Brasset, E.; Teysset, L.; Boivin, A. Environmentally-induced epigenetic conversion of a piRNA cluster. Elife 2019, 8, e39842. [Google Scholar] [CrossRef]
  75. Belicard, T.; Jareosettasin, P.; Sarkies, P. The piRNA pathway responds to environmental signals to establish intergenerational adaptation to stress. BMC Biol. 2018, 16, 103. [Google Scholar] [CrossRef]
  76. Donkin, I.; Versteyhe, S.; Ingerslev, L.R.; Qian, K.; Mechta, M.; Nordkap, L.; Mortensen, B.; Appel, E.V.; Jørgensen, N.; Kristiansen, V.B.; et al. Obesity and Bariatric Surgery Drive Epigenetic Variation of Spermatozoa in Humans. Cell Metab. 2016, 23, 369–378. [Google Scholar] [CrossRef]
  77. Grandjean, V.; Fourré, S.; De Abreu, D.A.; Derieppe, M.A.; Remy, J.J.; Rassoulzadegan, M. RNA-mediated paternal heredity of diet-induced obesity and metabolic disorders. Sci. Rep. 2015, 5, 18193. [Google Scholar] [CrossRef]
  78. de Castro Barbosa, T.; Ingerslev, L.R.; Alm, P.S.; Versteyhe, S.; Massart, J.; Rasmussen, M.; Donkin, I.; Sjögren, R.; Mudry, J.M.; Vetterli, L.; et al. High-fat diet reprograms the epigenome of rat spermatozoa and transgenerationally affects metabolism of the offspring. Mol. Metab. 2016, 5, 184–197. [Google Scholar] [CrossRef] [PubMed]
  79. Ingerslev, L.R.; Donkin, I.; Fabre, O.; Versteyhe, S.; Mechta, M.; Pattamaprapanont, P.; Mortensen, B.; Krarup, N.T.; Barrès, R. Endurance training remodels sperm-borne small RNA expression and methylation at neurological gene hotspots. Clin. Epigenetics 2018, 10, 12. [Google Scholar] [CrossRef]
  80. Gapp, K.; Jawaid, A.; Sarkies, P.; Bohacek, J.; Pelczar, P.; Prados, J.; Farinelli, L.; Miska, E.; Mansuy, I.M. Implication of sperm RNAs in transgenerational inheritance of the effects of early trauma in mice. Nat. Neurosci. 2014, 17, 667–669. [Google Scholar] [CrossRef]
  81. Han, R.; Zhang, L.; Gan, W.; Fu, K.; Jiang, K.; Ding, J.; Wu, J.; Han, X.; Li, D. piRNA-DQ722010 contributes to prostate hyperplasia of the male offspring mice after the maternal exposed to microcystin-leucine arginine. Prostate 2019, 79, 798–812. [Google Scholar] [CrossRef]
  82. Davis, S.R. TRIENNIAL LACTATION SYMPOSIUM/BOLFA: Mammary growth during pregnancy and lactation and its relationship with milk yield. J. Anim. Sci. 2017, 95, 5675–5688. [Google Scholar] [CrossRef] [PubMed]
  83. Hue-Beauvais, C.; Faulconnier, Y.; Charlier, M.; Leroux, C. Nutritional Regulation of Mammary Gland Development and Milk Synthesis in Animal Models and Dairy Species. Genes 2021, 12, 523. [Google Scholar] [CrossRef]
  84. Farmer, C.; Quesnel, H. Nutritional, hormonal, and environmental effects on colostrum in sows. J. Anim. Sci. 2009, 87, 56–64. [Google Scholar] [CrossRef] [PubMed]
  85. Hurley, W.L. Review: Mammary gland development in swine: Embryo to early lactation. Animal 2019, 13, s11–s19. [Google Scholar] [CrossRef]
  86. Akers, R.M.; Nickerson, S.C. Mastitis and its impact on structure and function in the ruminant mammary gland. J. Mammary Gland. Biol. Neoplasia 2011, 16, 275–289. [Google Scholar] [CrossRef]
  87. Guo, H.; Li, J.; Wang, Y.; Cao, X.; Lv, X.; Yang, Z.; Chen, Z. Progress in Research on Key Factors Regulating Lactation Initiation in the Mammary Glands of Dairy Cows. Genes 2023, 14, 1163. [Google Scholar] [CrossRef]
  88. Zhang, M.; Cao, M.; Kong, L.; Liu, J.; Wang, Y.; Song, C.; Chen, X.; Lai, M.; Fang, X.; Chen, H.; et al. MiR-204-5p promotes lipid synthesis in mammary epithelial cells by targeting SIRT1. Biochem. Biophys. Res. Commun. 2020, 533, 1490–1496. [Google Scholar] [CrossRef] [PubMed]
  89. Lanz, R.B.; Chua, S.S.; Barron, N.; Söder, B.M.; DeMayo, F.; O’Malley, B.W. Steroid receptor RNA activator stimulates proliferation as well as apoptosis in vivo. Mol. Cell Biol. 2003, 23, 7163–7176. [Google Scholar] [CrossRef]
  90. Pauli, A.; Rinn, J.L.; Schier, A.F. Non-coding RNAs as regulators of embryogenesis. Nat. Rev. Genet. 2011, 12, 136–149. [Google Scholar] [CrossRef]
  91. Esteller, M. Non-coding RNAs in human disease. Nat. Rev. Genet. 2011, 12, 861–874. [Google Scholar] [CrossRef]
  92. O’Brien, J.; Hayder, H.; Zayed, Y.; Peng, C. Overview of MicroRNA Biogenesis, Mechanisms of Actions, and Circulation. Front. Endocrinol. 2018, 9, 402. [Google Scholar] [CrossRef] [PubMed]
  93. Le Guillou, S.; Sdassi, N.; Laubier, J.; Passet, B.; Vilotte, M.; Castille, J.; Laloë, D.; Polyte, J.; Bouet, S.; Jaffrézic, F.; et al. Overexpression of miR-30b in the developing mouse mammary gland causes a lactation defect and delays involution. PLoS ONE 2012, 7, e45727. [Google Scholar] [CrossRef] [PubMed]
  94. Lv, C.; Li, F.; Li, X.; Tian, Y.; Zhang, Y.; Sheng, X.; Song, Y.; Meng, Q.; Yuan, S.; Luan, L.; et al. Author Correction: MiR-31 promotes mammary stem cell expansion and breast tumorigenesis by suppressing Wnt signaling antagonists. Nat. Commun. 2020, 11, 5308. [Google Scholar] [CrossRef] [PubMed]
  95. Lin, X.; Luo, J.; Zhang, L.; Wang, W.; Gou, D. MiR-103 controls milk fat accumulation in goat (Capra hircus) mammary gland during lactation. PLoS ONE 2013, 8, e79258. [Google Scholar] [CrossRef]
  96. Ibarra, I.; Erlich, Y.; Muthuswamy, S.K.; Sachidanandam, R.; Hannon, G.J. A role for microRNAs in maintenance of mouse mammary epithelial progenitor cells. Genes Dev. 2007, 21, 3238–3243. [Google Scholar] [CrossRef]
  97. Feuermann, Y.; Kang, K.; Shamay, A.; Robinson, G.W.; Hennighausen, L. MiR-21 is under control of STAT5 but is dispensable for mammary development and lactation. PLoS ONE 2014, 9, e85123. [Google Scholar] [CrossRef]
  98. Bonetti, P.; Climent, M.; Panebianco, F.; Tordonato, C.; Santoro, A.; Marzi, M.J.; Pelicci, P.G.; Ventura, A.; Nicassio, F. Correction: Dual role for miR-34a in the control of early progenitor proliferation and commitment in the mammary gland and in breast cancer. Oncogene 2020, 39, 2228. [Google Scholar] [CrossRef]
  99. Tanaka, T.; Haneda, S.; Imakawa, K.; Sakai, S.; Nagaoka, K. A microRNA, miR-101a, controls mammary gland development by regulating cyclooxygenase-2 expression. Differentiation 2009, 77, 181–187. [Google Scholar] [CrossRef]
  100. Ucar, A.; Vafaizadeh, V.; Jarry, H.; Fiedler, J.; Klemmt, P.A.; Thum, T.; Groner, B.; Chowdhury, K. miR-212 and miR-132 are required for epithelial stromal interactions necessary for mouse mammary gland development. Nat. Genet. 2010, 42, 1101–1108. [Google Scholar] [CrossRef]
  101. Lee, J.M.; Cho, K.W.; Kim, E.J.; Tang, Q.; Kim, K.S.; Tickle, C.; Jung, H.S. A contrasting function for miR-137 in embryonic mammogenesis and adult breast carcinogenesis. Oncotarget 2015, 6, 22048–22059. [Google Scholar] [CrossRef]
  102. Wang, C.; Li, Q.; Li, Y. miR-138 function and its targets on mouse mammary epithelial cells. Progress Biochem. Biophys. 2006. Available online: https://pesquisa.bvsalud.org/portal/resource/pt/wpr-592163 (accessed on 1 May 2025).
  103. Cui, Y.; Sun, X.; Jin, L.; Yu, G.; Li, Q.; Gao, X.; Ao, J.; Wang, C. MiR-139 suppresses β-casein synthesis and proliferation in bovine mammary epithelial cells by targeting the GHR and IGF1R signaling pathways. BMC Vet. Res. 2017, 13, 350. [Google Scholar] [CrossRef] [PubMed]
  104. Jiang, N.; Wu, C.; Li, Y.; Liu, J.; Yuan, Y.; Shi, H. Identification and profiling of microRNAs involved in the regenerative involution of mammary gland. Genomics 2022, 114, 110442. [Google Scholar] [CrossRef] [PubMed]
  105. Yoo, K.H.; Kang, K.; Feuermann, Y.; Jang, S.J.; Robinson, G.W.; Hennighausen, L. The STAT5-regulated miR-193b locus restrains mammary stem and progenitor cell activity and alveolar differentiation. Dev. Biol. 2014, 395, 245–254. [Google Scholar] [CrossRef]
  106. Shimono, Y.; Zabala, M.; Cho, R.W.; Lobo, N.; Dalerba, P.; Qian, D.; Diehn, M.; Liu, H.; Panula, S.P.; Chiao, E.; et al. Downregulation of miRNA-200c links breast cancer stem cells with normal stem cells. Cell 2009, 138, 592–603. [Google Scholar] [CrossRef] [PubMed]
  107. Lu, Y.; Cao, J.; Napoli, M.; Xia, Z.; Zhao, N.; Creighton, C.J.; Li, W.; Chen, X.; Flores, E.R.; McManus, M.T.; et al. miR-205 Regulates Basal Cell Identity and Stem Cell Regenerative Potential During Mammary Reconstitution. Stem Cells 2018, 36, 1875–1889. [Google Scholar] [CrossRef]
  108. Patel, Y.; Soni, M.; Awgulewitsch, A.; Kern, M.J.; Liu, S.; Shah, N.; Singh, U.P.; Chen, H. Correction: Overexpression of miR-489 derails mammary hierarchy structure and inhibits HER2/neu-induced tumorigenesis. Oncogene 2019, 38, 454. [Google Scholar] [CrossRef]
  109. Chu, M.; Zhao, Y.; Yu, S.; Hao, Y.; Zhang, P.; Feng, Y.; Zhang, H.; Ma, D.; Liu, J.; Cheng, M.; et al. miR-15b negatively correlates with lipid metabolism in mammary epithelial cells. Am. J. Physiol. Cell Physiol. 2018, 314, C43–C52. [Google Scholar] [CrossRef]
  110. Chen, Z.; Luo, J.; Sun, S.; Cao, D.; Shi, H.; Loor, J.J. miR-148a and miR-17-5p synergistically regulate milk TAG synthesis via PPARGC1A and PPARA in goat mammary epithelial cells. RNA Biol. 2017, 14, 326–338. [Google Scholar] [CrossRef]
  111. Wang, H.; Luo, J.; Chen, Z.; Cao, W.T.; Xu, H.F.; Gou, D.M.; Zhu, J.J. MicroRNA-24 can control triacylglycerol synthesis in goat mammary epithelial cells by targeting the fatty acid synthase gene. J. Dairy Sci. 2015, 98, 9001–9014. [Google Scholar] [CrossRef]
  112. Ma, L.; Qiu, H.; Chen, Z.; Li, L.; Zeng, Y.; Luo, J.; Gou, D. miR-25 modulates triacylglycerol and lipid accumulation in goat mammary epithelial cells by repressing PGC-1beta. J. Anim. Sci. Biotechnol. 2018, 9, 48. [Google Scholar] [CrossRef]
  113. Lin, X.Z.; Luo, J.; Zhang, L.P.; Wang, W.; Shi, H.B.; Zhu, J.J. MiR-27a suppresses triglyceride accumulation and affects gene mRNA expression associated with fat metabolism in dairy goat mammary gland epithelial cells. Gene 2013, 521, 15–23. [Google Scholar] [CrossRef] [PubMed]
  114. Bian, Y.; Lei, Y.; Wang, C.; Wang, J.; Wang, L.; Liu, L.; Liu, L.; Gao, X.; Li, Q. Epigenetic Regulation of miR-29s Affects the Lactation Activity of Dairy Cow Mammary Epithelial Cells. J. Cell Physiol. 2015, 230, 2152–2163. [Google Scholar] [CrossRef] [PubMed]
  115. Cui, W.; Li, Q.; Feng, L.; Ding, W. MiR-126-3p regulates progesterone receptors and involves development and lactation of mouse mammary gland. Mol. Cell Biochem. 2011, 355, 17–25. [Google Scholar] [CrossRef]
  116. Chu, M.; Zhao, Y.; Feng, Y.; Zhang, H.; Liu, J.; Cheng, M.; Li, L.; Shen, W.; Cao, H.; Li, Q.; et al. MicroRNA-126 participates in lipid metabolism in mammary epithelial cells. Mol. Cell Endocrinol. 2017, 454, 77–86. [Google Scholar] [CrossRef]
  117. Tian, L.; Zhang, L.; Cui, Y.; Li, H.; Xie, X.; Li, Y.; Wang, C. miR-142-3p Regulates Milk Synthesis and Structure of Murine Mammary Glands via PRLR-Mediated Multiple Signaling Pathways. J. Agric. Food Chem. 2019, 67, 9532–9542. [Google Scholar] [CrossRef] [PubMed]
  118. Wang, H.; Shi, H.; Luo, J.; Yi, Y.; Yao, D.; Zhang, X.; Ma, G.; Loor, J.J. MiR-145 Regulates Lipogenesis in Goat Mammary Cells Via Targeting INSIG1 and Epigenetic Regulation of Lipid-Related Genes. J. Cell Physiol. 2017, 232, 1030–1040. [Google Scholar] [CrossRef]
  119. Heinz, R.E.; Rudolph, M.C.; Ramanathan, P.; Spoelstra, N.S.; Butterfield, K.T.; Webb, P.G.; Babbs, B.L.; Gao, H.; Chen, S.; Gordon, M.A.; et al. Constitutive expression of microRNA-150 in mammary epithelium suppresses secretory activation and impairs de novo lipogenesis. Development 2016, 143, 4236–4248. [Google Scholar] [CrossRef]
  120. Chen, Z.; Shi, H.; Sun, S.; Xu, H.; Cao, D.; Luo, J. MicroRNA-181b suppresses TAG via target IRS2 and regulating multiple genes in the Hippo pathway. Exp. Cell Res. 2016, 348, 66–74. [Google Scholar] [CrossRef] [PubMed]
  121. Wang, J.; Aydoğdu, E.; Mukhopadhyay, S.; Helguero, L.A.; Williams, C. A miR-206 regulated gene landscape enhances mammary epithelial differentiation. J. Cell Physiol. 2019, 234, 22220–22233. [Google Scholar] [CrossRef]
  122. Chu, M.; Zhao, Y.; Yu, S.; Hao, Y.; Zhang, P.; Feng, Y.; Zhang, H.; Ma, D.; Liu, J.; Cheng, M.; et al. MicroRNA-221 may be involved in lipid metabolism in mammary epithelial cells. Int. J. Biochem. Cell Biol. 2018, 97, 118–127. [Google Scholar] [CrossRef]
  123. Li, D.; Xie, X.; Wang, J.; Bian, Y.; Li, Q.; Gao, X.; Wang, C. MiR-486 regulates lactation and targets the PTEN gene in cow mammary glands. PLoS ONE 2015, 10, e0118284. [Google Scholar] [CrossRef] [PubMed]
  124. Wang, Y.; Dong, J.; Li, D.; Lai, L.; Siwko, S.; Li, Y.; Liu, M. Lgr4 regulates mammary gland development and stem cell activity through the pluripotency transcription factor Sox2. Stem Cells 2013, 31, 1921–1931. [Google Scholar] [CrossRef] [PubMed]
  125. Askarian-Amiri, M.E.; Crawford, J.; French, J.D.; Smart, C.E.; Smith, M.A.; Clark, M.B.; Ru, K.; Mercer, T.R.; Thompson, E.R.; Lakhani, S.R.; et al. SNORD-host RNA Zfas1 is a regulator of mammary development and a potential marker for breast cancer. RNA 2011, 17, 878–891. [Google Scholar] [CrossRef] [PubMed]
  126. Adriaenssens, E.; Lottin, S.; Dugimont, T.; Fauquette, W.; Coll, J.; Dupouy, J.P.; Boilly, B.; Curgy, J.J. Steroid hormones modulate H19 gene expression in both mammary gland and uterus. Oncogene 1999, 18, 4460–4473. [Google Scholar] [CrossRef]
  127. Ginger, M.R.; Shore, A.N.; Contreras, A.; Rijnkels, M.; Miller, J.; Gonzalez-Rimbau, M.F.; Rosen, J.M. A noncoding RNA is a potential marker of cell fate during mammary gland development. Proc. Natl. Acad. Sci. USA 2006, 103, 5781–5786. [Google Scholar] [CrossRef]
  128. Shore, A.N.; Kabotyanski, E.B.; Roarty, K.; Smith, M.A.; Zhang, Y.; Creighton, C.J.; Dinger, M.E.; Rosen, J.M. Pregnancy-induced noncoding RNA (PINC) associates with polycomb repressive complex 2 and regulates mammary epithelial differentiation. PLoS Genet. 2012, 8, e1002840. [Google Scholar] [CrossRef]
  129. Ma, X.; Niu, X.; Huang, S.; Li, S.; Ran, X.; Wang, J.; Dai, X. The piRNAs present in the developing testes of Chinese indigenous Xiang pigs. Theriogenology 2022, 189, 92–106. [Google Scholar] [CrossRef]
  130. Gebert, D.; Ketting, R.F.; Zischler, H.; Rosenkranz, D. piRNAs from Pig Testis Provide Evidence for a Conserved Role of the Piwi Pathway in Post-Transcriptional Gene Regulation in Mammals. PLoS ONE 2015, 10, e0124860. [Google Scholar] [CrossRef]
  131. Zhang, G.W.; Wang, L.; Chen, H.; Guan, J.; Wu, Y.; Zhao, J.; Luo, Z.; Huang, W.; Zuo, F. Promoter hypermethylation of PIWI/piRNA pathway genes associated with diminished pachytene piRNA production in bovine hybrid male sterility. Epigenetics 2020, 15, 914–931. [Google Scholar] [CrossRef]
  132. Capra, E.; Turri, F.; Lazzari, B.; Cremonesi, P.; Gliozzi, T.M.; Fojadelli, I.; Stella, A.; Pizzi, F. Small RNA sequencing of cryopreserved semen from single bull revealed altered miRNAs and piRNAs expression between High- and Low-motile sperm populations. BMC Genom. 2017, 18, 14. [Google Scholar] [CrossRef]
  133. Li, C.; Zhang, R.; Zhang, Z.; Ren, C.; Wang, X.; He, X.; Mwacharo, J.M.; Zhang, X.; Zhang, J.; Di, R.; et al. Expression characteristics of piRNAs in ovine luteal phase and follicular phase ovaries. Front. Vet. Sci. 2022, 9, 921868. [Google Scholar] [CrossRef] [PubMed]
  134. Testroet, E.D.; Shome, S.; Testroet, A.; Reecy, J.; Jernigan, R.L.; Zhu, M.; Du, M.; Clark, S.; Beitz, D. Profiling of the Exosomal Cargo of Bovine Milk Reveals the Presence of Immune-and Growth-modulatory ncRNAs. FASEB J. 2018, 32, 747.25. [Google Scholar] [CrossRef]
  135. Ablondi, M.; Gòdia, M.; Rodriguez-Gil, J.E.; Sánchez, A.; Clop, A. Characterisation of sperm piRNAs and their correlation with semen quality traits in swine. Anim. Genet. 2021, 52, 114–120. [Google Scholar] [CrossRef]
  136. Kowalczykiewicz, D.; Pawlak, P.; Lechniak, D.; Wrzesinski, J. Altered expression of porcine Piwi genes and piRNA during development. PLoS ONE 2012, 7, e43816. [Google Scholar] [CrossRef]
  137. Wang, C.; Chen, Y.; Yang, X.; Du, Y.; Xu, Z.; Zhou, Y.; Yang, X.; Wang, X.; Zhang, C.; Li, S.; et al. The porcine piRNA transcriptome response to Senecavirus a infection. Front. Vet. Sci. 2023, 10, 1126277. [Google Scholar] [CrossRef] [PubMed]
  138. Weng, B.; Ran, M.; Chen, B.; Wu, M.; Peng, F.; Dong, L.; He, C.; Zhang, S.; Li, Z. Systematic identification and characterization of miRNAs and piRNAs from porcine testes. Genes Genom. 2017, 39, 1047–1057. [Google Scholar] [CrossRef]
  139. Yang, C.X.; Du, Z.Q.; Wright, E.C.; Rothschild, M.F.; Prather, R.S.; Ross, J.W. Small RNA profile of the cumulus-oocyte complex and early embryos in the pig. Biol. Reprod. 2012, 87, 117. [Google Scholar] [CrossRef]
  140. Xu, Z.; Xie, Y.; Zhou, C.; Hu, Q.; Gu, T.; Yang, J.; Zheng, E.; Huang, S.; Xu, Z.; Cai, G.; et al. Expression Pattern of Seminal Plasma Extracellular Vesicle Small RNAs in Boar Semen. Front. Vet. Sci. 2020, 7, 585276. [Google Scholar] [CrossRef]
  141. Russell, S.; Patel, M.; Gilchrist, G.; Stalker, L.; Gillis, D.; Rosenkranz, D.; LaMarre, J. Bovine piRNA-like RNAs are associated with both transposable elements and mRNAs. Reproduction 2017, 153, 305–318. [Google Scholar] [CrossRef]
  142. Spornraft, M.; Kirchner, B.; Pfaffl, M.W.; Riedmaier, I. Comparison of the miRNome and piRNome of bovine blood and plasma by small RNA sequencing. Biotechnol. Lett. 2015, 37, 1165–1176. [Google Scholar] [CrossRef]
  143. Wang, H.; Zhong, J.; Chai, Z.; Zhu, J.; Xin, J. Comparative expression profile of microRNAs and piRNAs in three ruminant species testes using next-generation sequencing. Reprod. Domest. Anim. 2018, 53, 963–970. [Google Scholar] [CrossRef] [PubMed]
  144. Sellem, E.; Marthey, S.; Rau, A.; Jouneau, L.; Bonnet, A.; Perrier, J.P.; Fritz, S.; Le Danvic, C.; Boussaha, M.; Kiefer, H.; et al. A comprehensive overview of bull sperm-borne small non-coding RNAs and their diversity across breeds. Epigenetics Chromatin 2020, 13, 19. [Google Scholar] [CrossRef]
  145. Roovers, E.F.; Rosenkranz, D.; Mahdipour, M.; Han, C.T.; He, N.; Chuva de Sousa Lopes, S.M.; van der Westerlaken, L.A.; Zischler, H.; Butter, F.; Roelen, B.A.; et al. Piwi proteins and piRNAs in mammalian oocytes and early embryos. Cell Rep. 2015, 10, 2069–2082. [Google Scholar] [CrossRef] [PubMed]
  146. Sellem, E.; Marthey, S.; Rau, A.; Jouneau, L.; Bonnet, A.; Le Danvic, C.; Guyonnet, B.; Kiefer, H.; Jammes, H.; Schibler, L. Dynamics of cattle sperm sncRNAs during maturation, from testis to ejaculated sperm. Epigenetics Chromatin 2021, 14, 24. [Google Scholar] [CrossRef]
  147. Shome, S.; Jernigan, R.L.; Beitz, D.C.; Clark, S.; Testroet, E.D. Non-coding RNA in raw and commercially processed milk and putative targets related to growth and immune-response. BMC Genom. 2021, 22, 749. [Google Scholar] [CrossRef] [PubMed]
  148. Chhabra, P.; Goel, B.M.; Arora, R. Analysis of differentially expressed diverse non-coding rnas in different stages of lactation of murrah buffalo. BIOINFOLET Q. J. Life Sci. 2023, 20, 524–530. [Google Scholar]
  149. Li, B.; He, X.; Zhao, Y.; Bai, D.; Bou, G.; Zhang, X.; Su, S.; Dao, L.; Liu, R.; Wang, Y.; et al. Identification of piRNAs and piRNA clusters in the testes of the Mongolian horse. Sci. Rep. 2019, 9, 5022. [Google Scholar] [CrossRef]
  150. Di, R.; Zhang, R.; Mwacharo, J.M.; Wang, X.; He, X.; Liu, Y.; Zhang, J.; Gong, Y.; Zhang, X.; Chu, M. Characteristics of piRNAs and their comparative profiling in testes of sheep with different fertility. Front. Genet. 2022, 13, 1078049. [Google Scholar] [CrossRef]
  151. He, X.; Li, B.; Fu, S.; Wang, B.; Qi, Y.; Da, L.; Te, R.; Sun, S.; Liu, Y.; Zhang, W. Identification of piRNAs in the testes of Sunite and Small-tailed Han sheep. Anim. Biotechnol. 2021, 32, 13–20. [Google Scholar] [CrossRef]
  152. Li, T.; Wang, H.; Ma, K.; Wu, Y.; Qi, X.; Liu, Z.; Li, Q.; Zhang, Y.; Ma, Y. Identification and functional characterization of developmental-stage-dependent piRNAs in Tibetan sheep testes. J. Anim. Sci. 2023, 101, skad189. [Google Scholar] [CrossRef]
  153. Cheng, J.; Guo, J.M.; Xiao, B.X.; Miao, Y.; Jiang, Z.; Zhou, H.; Li, Q.N. piRNA, the new non-coding RNA, is aberrantly expressed in human cancer cells. Clin. Chim. Acta 2011, 412, 1621–1625. [Google Scholar] [CrossRef] [PubMed]
  154. Huang, G.; Hu, H.; Xue, X.; Shen, S.; Gao, E.; Guo, G.; Shen, X.; Zhang, X. Altered expression of piRNAs and their relation with clinicopathologic features of breast cancer. Clin. Transl. Oncol. 2013, 15, 563–568. [Google Scholar] [CrossRef] [PubMed]
  155. Hashim, A.; Rizzo, F.; Marchese, G.; Ravo, M.; Tarallo, R.; Nassa, G.; Giurato, G.; Santamaria, G.; Cordella, A.; Cantarella, C.; et al. RNA sequencing identifies specific PIWI-interacting small non-coding RNA expression patterns in breast cancer. Oncotarget 2014, 5, 9901–9910. [Google Scholar] [CrossRef]
  156. Koduru, S.V.; Tiwari, A.K.; Leberfinger, A.; Hazard, S.W.; Kawasawa, Y.I.; Mahajan, M.; Ravnic, D.J. A Comprehensive NGS Data Analysis of Differentially Regulated miRNAs, piRNAs, lncRNAs and sn/snoRNAs in Triple Negative Breast Cancer. J. Cancer 2017, 8, 578–596. [Google Scholar] [CrossRef]
  157. Kärkkäinen, E.; Heikkinen, S.; Tengström, M.; Kosma, V.M.; Mannermaa, A.; Hartikainen, J.M. The debatable presence of PIWI-interacting RNAs in invasive breast cancer. Cancer Med. 2021, 10, 3593–3603. [Google Scholar] [CrossRef]
  158. Liu, T.; Wang, J.; Sun, L.; Li, M.; He, X.; Jiang, J.; Zhou, Q. Piwi-interacting RNA-651 promotes cell proliferation and migration and inhibits apoptosis in breast cancer by facilitating DNMT1-mediated PTEN promoter methylation. Cell Cycle 2021, 20, 1603–1616. [Google Scholar] [CrossRef]
  159. Ding, X.; Li, Y.; Lü, J.; Zhao, Q.; Guo, Y.; Lu, Z.; Ma, W.; Liu, P.; Pestell, R.G.; Liang, C.; et al. piRNA-823 Is Involved in Cancer Stem Cell Regulation Through Altering DNA Methylation in Association with Luminal Breast Cancer. Front. Cell Dev. Biol. 2021, 9, 641052. [Google Scholar] [CrossRef]
  160. Oner, C.; Colak, E. PIWI interacting RNA-823: Epigenetic regulator of the triple negative breast cancer cells proliferation. Eurasian J. Med. Oncol. 2022, 6, 339–344. [Google Scholar] [CrossRef]
  161. Ou, B.; Liu, Y.; Gao, Z.; Xu, J.; Yan, Y.; Li, Y.; Zhang, J. Senescent neutrophils-derived exosomal piRNA-17560 promotes chemoresistance and EMT of breast cancer via FTO-mediated m6A demethylation. Cell Death Dis. 2022, 13, 905. [Google Scholar] [CrossRef]
  162. Huang, S.; Chen, B.; Qiu, P.; Yan, Z.; Liang, Z.; Luo, K.; Huang, B.; Jiang, H. In vitro study of piwi interaction RNA-31106 promoting breast carcinogenesis by regulating METTL3-mediated m6A RNA methylation. Transl. Cancer Res. 2023, 12, 1588–1601. [Google Scholar] [CrossRef]
  163. Du, S.; Liu, J.; Ning, Y.; Yin, M.; Xu, M.; Liu, Z.; Liu, K. The piR-31115-PIWIL4 complex promotes the migration of the triple-negative breast cancer cell lineMDA-MB-231 by suppressing HSP90AA1 degradation. Gene 2025, 942, 149255. [Google Scholar] [CrossRef]
  164. Alexandrova, E.; Lamberti, J.; Saggese, P.; Pecoraro, G.; Memoli, D.; Cappa, V.M.; Ravo, M.; Iorio, R.; Tarallo, R.; Rizzo, F.; et al. Small Non-Coding RNA Profiling Identifies miR-181a-5p as a Mediator of Estrogen Receptor Beta-Induced Inhibition of Cholesterol Biosynthesis in Triple-Negative Breast Cancer. Cells 2020, 9, 874. [Google Scholar] [CrossRef]
  165. Tan, L.; Mai, D.; Zhang, B.; Jiang, X.; Zhang, J.; Bai, R.; Ye, Y.; Li, M.; Pan, L.; Su, J.; et al. PIWI-interacting RNA-36712 restrains breast cancer progression and chemoresistance by interaction with SEPW1 pseudogene SEPW1P RNA. Mol. Cancer 2019, 18, 9. [Google Scholar] [CrossRef] [PubMed]
  166. Lü, J.; Zhao, Q.; Ding, X.; Guo, Y.; Li, Y.; Xu, Z.; Li, S.; Wang, Z.; Shen, L.; Chen, H.W.; et al. Cyclin D1 promotes secretion of pro-oncogenic immuno-miRNAs and piRNAs. Clin. Sci. 2020, 134, 791–805. [Google Scholar] [CrossRef] [PubMed]
  167. Fu, A.; Jacobs, D.I.; Hoffman, A.E.; Zheng, T.; Zhu, Y. PIWI-interacting RNA 021285 is involved in breast tumorigenesis possibly by remodeling the cancer epigenome. Carcinogenesis 2015, 36, 1094–1102. [Google Scholar] [CrossRef] [PubMed]
  168. He, X.; Chen, X.; Zhang, X.; Duan, X.; Pan, T.; Hu, Q.; Zhang, Y.; Zhong, F.; Liu, J.; Zhang, H.; et al. An Lnc RNA (GAS5)/SnoRNA-derived piRNA induces activation of TRAIL gene by site-specifically recruiting MLL/COMPASS-like complexes. Nucleic Acids Res. 2015, 43, 3712–3725. [Google Scholar] [CrossRef]
  169. Jin, L.; Zhang, Z.; Wang, Z.; Tan, X.; Wang, Z.; Shen, L.; Long, C.; Wei, G.; He, D. Novel piRNA MW557525 regulates the growth of Piwil2-iCSCs and maintains their stem cell pluripotency. Mol. Biol. Rep. 2022, 49, 6957–6969. [Google Scholar] [CrossRef]
  170. Balaratnam, S.; West, N.; Basu, S. A piRNA utilizes HILI and HIWI2 mediated pathway to down-regulate ferritin heavy chain 1 mRNA in human somatic cells. Nucleic Acids Res. 2018, 46, 10635–10648. [Google Scholar] [CrossRef]
  171. Wu, L.; Huang, S.; Tian, W.; Liu, P.; Xie, Y.; Qiu, Y.; Li, X.; Tang, Y.; Zheng, S.; Sun, Y.; et al. PIWI-interacting RNA-YBX1 inhibits proliferation and metastasis by the MAPK signaling pathway via YBX1 in triple-negative breast cancer. Cell Death Discov. 2024, 10, 7. [Google Scholar] [CrossRef]
  172. Ma, X.; Wang, S.; Do, T.; Song, X.; Inaba, M.; Nishimoto, Y.; Liu, L.P.; Gao, Y.; Mao, Y.; Li, H.; et al. Piwi is required in multiple cell types to control germline stem cell lineage development in the Drosophila ovary. PLoS ONE 2014, 9, e90267. [Google Scholar] [CrossRef]
  173. Cox, D.N.; Chao, A.; Baker, J.; Chang, L.; Qiao, D.; Lin, H. A novel class of evolutionarily conserved genes defined by piwi are essential for stem cell self-renewal. Genes Dev. 1998, 12, 3715–3727. [Google Scholar] [CrossRef] [PubMed]
  174. Visvader, J.E. Keeping abreast of the mammary epithelial hierarchy and breast tumorigenesis. Genes Dev. 2009, 23, 2563–2577. [Google Scholar] [CrossRef] [PubMed]
  175. Visvader, J.E.; Clevers, H. Tissue-specific designs of stem cell hierarchies. Nat. Cell Biol. 2016, 18, 349–355. [Google Scholar] [CrossRef] [PubMed]
  176. Shackleton, M. Normal stem cells and cancer stem cells: Similar and different. Semin. Cancer Biol. 2010, 20, 85–92. [Google Scholar] [CrossRef]
  177. Yu, Q.C.; Verheyen, E.M.; Zeng, Y.A. Mammary Development and Breast Cancer: A Wnt Perspective. Cancers 2016, 8, 65. [Google Scholar] [CrossRef]
  178. Chen, W.; Wei, W.; Yu, L.; Ye, Z.; Huang, F.; Zhang, L.; Hu, S.; Cai, C. Mammary Development and Breast Cancer: A Notch Perspective. J. Mammary Gland. Biol. Neoplasia 2021, 26, 309–320. [Google Scholar] [CrossRef]
  179. Visbal, A.P.; Lewis, M.T. Hedgehog signaling in the normal and neoplastic mammary gland. Curr. Drug Targets 2010, 11, 1103–1111. [Google Scholar] [CrossRef]
  180. Singh, N.; Baby, D.; Rajguru, J.P.; Patil, P.B.; Thakkannavar, S.S.; Pujari, V.B. Inflammation and cancer. Ann. Afr. Med. 2019, 18, 121–126. [Google Scholar] [CrossRef]
  181. Borish, L.C.; Steinke, J.W. 2. Cytokines and chemokines. J. Allergy Clin. Immunol. 2003, 111, S460–S475. [Google Scholar] [CrossRef]
  182. Marques-Rocha, J.L.; Samblas, M.; Milagro, F.I.; Bressan, J.; Martínez, J.A.; Marti, A. Noncoding RNAs, cytokines, and inflammation-related diseases. FASEB J. 2015, 29, 3595–3611. [Google Scholar] [CrossRef]
  183. Liao, Z.; Yang, L.; Cheng, X.; Huang, X.; Zhang, Q.; Wen, D.; Song, Z.; Li, Y.; Wen, S.; Li, Y.; et al. pir-hsa-216911 inhibit pyroptosis in hepatocellular carcinoma by suppressing TLR4 initiated GSDMD activation. Cell Death Discov. 2025, 11, 11. [Google Scholar] [CrossRef] [PubMed]
  184. Radmehr, E.; Yazdanpanah, N.; Rezaei, N. Non-coding RNAs affecting NLRP3 inflammasome pathway in diabetic cardiomyopathy: A comprehensive review of potential therapeutic options. J. Transl. Med. 2025, 23, 249. [Google Scholar] [CrossRef] [PubMed]
  185. Zhang, X.; Wang, X.; Yu, F.; Wang, C.; Peng, J.; Wang, C.; Chen, X. PiRNA hsa_piR_019949 promotes chondrocyte anabolic metabolism by inhibiting the expression of lncRNA NEAT1. J. Orthop. Surg. Res. 2024, 19, 31. [Google Scholar] [CrossRef]
  186. Zhong, N.; Nong, X.; Diao, J.; Yang, G. piRNA-6426 increases DNMT3B-mediated SOAT1 methylation and improves heart failure. Aging 2022, 14, 2678–2694. [Google Scholar] [CrossRef]
  187. Liu, P.; Hu, L.; Shi, Y.; Liu, Y.; Yu, G.; Zhou, Y.; An, Q.; Zhu, W. Changes in the Small RNA Expression in Endothelial Cells in Response to Inflammatory Stimulation. Oxid. Med. Cell Longev. 2021, 2021, 8845520. [Google Scholar] [CrossRef]
  188. Zhang, Y.; Jiao, X.; Wang, T.; Yue, X.; Wang, Y.; Cai, B.; Wang, C.; Lu, S. piRNA mmu_piR_037459 suppression alleviated the degeneration of chondrocyte and cartilage. Int. Immunopharmacol. 2024, 128, 111473. [Google Scholar] [CrossRef] [PubMed]
  189. Saha, B.; Chakravarty, S.; Ray, S.; Saha, H.; Das, K.; Ghosh, I.; Mallick, B.; Biswas, N.K.; Goswami, S. Correlating tissue and plasma-specific piRNA changes to predict their possible role in pancreatic malignancy and chronic inflammation. Biomed. Rep. 2024, 21, 186. [Google Scholar] [CrossRef]
  190. Ren, R.; Tan, H.; Huang, Z.; Wang, Y.; Yang, B. Differential expression and correlation of immunoregulation related piRNA in rheumatoid arthritis. Front. Immunol. 2023, 14, 1175924. [Google Scholar] [CrossRef]
  191. Samir, M.; Vidal, R.O.; Abdallah, F.; Capece, V.; Seehusen, F.; Geffers, R.; Hussein, A.; Ali, A.A.H.; Bonn, S.; Pessler, F. Organ-specific small non-coding RNA responses in domestic (Sudani) ducks experimentally infected with highly pathogenic avian influenza virus (H5N1). RNA Biol. 2020, 17, 112–124. [Google Scholar] [CrossRef]
  192. Nathan, C. Nonresolving inflammation redux. Immunity 2022, 55, 592–605. [Google Scholar] [CrossRef]
  193. Akhtar, M.; Guo, S.; Guo, Y.F.; Zahoor, A.; Shaukat, A.; Chen, Y.; Umar, T.; Deng, P.G.; Guo, M. Upregulated-gene expression of pro-inflammatory cytokines (TNF-α, IL-1β and IL-6) via TLRs following NF-κB and MAPKs in bovine mastitis. Acta Trop. 2020, 207, 105458. [Google Scholar] [CrossRef] [PubMed]
  194. Tak, P.P.; Firestein, G.S. NF-kappaB: A key role in inflammatory diseases. J. Clin. Investig. 2001, 107, 7–11. [Google Scholar] [CrossRef]
  195. Kyriakis, J.M.; Avruch, J. Mammalian mitogen-activated protein kinase signal transduction pathways activated by stress and inflammation. Physiol. Rev. 2001, 81, 807–869. [Google Scholar] [CrossRef] [PubMed]
  196. Haneklaus, M.; O’Neill, L.A. NLRP3 at the interface of metabolism and inflammation. Immunol. Rev. 2015, 265, 53–62. [Google Scholar] [CrossRef] [PubMed]
  197. Godber, O.F.; Wall, R. Livestock and food security: Vulnerability to population growth and climate change. Glob. Chang. Biol. 2014, 20, 3092–3102. [Google Scholar] [CrossRef]
  198. Klous, G.; Huss, A.; Heederik, D.J.J.; Coutinho, R.A. Human-livestock contacts and their relationship to transmission of zoonotic pathogens, a systematic review of literature. One Health 2016, 2, 65–76. [Google Scholar] [CrossRef]
Figure 1. The primary and ping-pong mechanism for piRNA biogenesis.
Figure 1. The primary and ping-pong mechanism for piRNA biogenesis.
Vetsci 12 00594 g001
Figure 2. Differences between miRNA, siRNA, and piRNA.
Figure 2. Differences between miRNA, siRNA, and piRNA.
Vetsci 12 00594 g002
Figure 3. Possibility of piRNA in mammary development and mastitis.
Figure 3. Possibility of piRNA in mammary development and mastitis.
Vetsci 12 00594 g003
Table 1. Proteins involved in the generation of piRNAs in different species.
Table 1. Proteins involved in the generation of piRNAs in different species.
Species5′-Terminal Enzyme3′-Terminal EnzymeKey PIWI ProteinReference
NematodesUncertaintyUncertaintyPlasticity-related gene 1 (PRG-1), plasticity-related gene 2 (PRG-2)[29]
ZebrafishPhospholipase D family member 6 (PLD6)UncertaintyZIWI, ZILI[30,31]
DrosophilaZucTrimmerPIWI, Argonaute 3 (Ago3), Aubergin(Aub)[32,33]
MicePLD6Poly(A)-specific ribonuclease-like domain containing 1 (PNLDC1)MIWI, MIWI2, MILI[34,35,36]
Table 2. The role of miRNAs in mammary gland development.
Table 2. The role of miRNAs in mammary gland development.
Particular YearmiRNAResearch TargetModel TypeOutcomesReferences
Mammary gland development
2007let-7Comma-DβIn vitroInhibited self-renewal capacity of progenitor cells and promoted differentiation.[96]
2014miR-21HC11, miceIn vivo and in vitroRegulated mammary gland development and lactation.[97]
2012miR-30bMiceIn vivoInhibited normal mammary gland development and lipid droplet accumulation.[93]
2020miR-31HC11, miceIn vivo and in vitroPromoted MaSCS self-renewal, alveogenesis, and lipid droplet accumulation.[94]
2020miR-34aComma-Dβ, SUM159pt, miceIn vivo and in vitroInhibited MaSCs self-renewal, terminal end bud (TEB) development.[98]
2009miR-101aHC11, miceIn vivo and in vitroInhibited HC11 proliferation and β-casein expression, affected mammary gland development and degeneration.[99]
2010miR-132, miR-212MiceIn vivoPromoted ducts growth and modulated epithelial–stromal interactions.[100]
2015miR-137MDA-MB-231, 293T, miceIn vivo and in vitroPromoted thickening of the mammary substrate.[101]
2006miR-138Mouse mammary epithelial cells, miceIn vivo and in vitroRegulated mammary epithelial cell proliferation and mammary gland development, promoted β-casein expression.[102]
2017miR-139BMEC, Holstein cowsIn vivo and in vitroInhibited β-casein synthesis and BMEC proliferation.[103]
2022miR-142-5p, miR-148C, miR-152, miR-218, GoatsIn vivoRegulated mammary gland regenerative degeneration.[104]
2014miR-193b MEC, mice In vivo and in vitroInhibited mammary stem/progenitor cell activity and alveolar differentiation.[105]
2009miR-200c 293T, Tera-2, mice In vivo and in vitroInhibited mammary duct formation.[106]
2018miR-205MEC, miceIn vivo and in vitroImpacted mammary regenerative capacity and mammary homeostasis.[107]
2019miR-489Mouse mammary epithelial cells, miceIn vivo and in vitroInhibited duct growth and TEB formation.[108]
Milk component synthesis
2018miR-15bMCF-10A, mice, goatsIn vivo and in vitroInhibited lipid synthesis and metabolism.[109]
2017miR-17-5p
miR-148a
GMEC, goatsIn vivo and in vitroPromoted triacylglycerol (TAG) synthesis and milk fat droplet accumulation.[110]
2015miR-24GMEC, goatsIn vivo and in vitroIncreased unsaturated fatty acid concentrations, TAG levels, and milk fat droplet accumulation.[111]
2018miR-25GMEC, goatsIn vivo and in vitroInhibited TAG synthesis and lipid droplet accumulation.[112]
2013miR-27aGMEC, goatsIn vivo and in vitroInhibited TAG synthesis and reduced the ratio of unsaturated/saturated fatty acids.[113]
2015miR-29sDCMEC, 293T, Chinese Holstein cowsIn vivo and in vitroInhibited triglyceride, protein, and lactose secretion.[114]
2013miR-103GMEC, goatsIn vivo and in vitroPromoted lipid droplet accumulation and TAG accumulation.[95]
2011/2017miR-126-3pMCF-10A, miceIn vivo and in vitroInhibited β-casein secretion and lipid synthesis.[115,116]
2019miR-142-3pMMGEC, miceIn vivo and in vitroInhibited secretion of β-casein and TAG.[117]
2017miR-145GMEC, goatsIn vivo and in vitroPromoted lipid droplet enlargement and TAG accumulation, increased the relative content of unsaturated fatty acids.[118]
2016miR-150-5pMiceIn vivoInhibited the de novo synthesis of lipids and fatty acids.[119]
2016miR-181bGMEC, goatsIn vivo and in vitroIncreased TAG levels and cream droplet accumulation.[120]
2020miR-204 HC11, mice In vivo and in vitroPromoted β-casein and milk fat synthesis.[88]
2019miR-206 HC11, mice In vivo and in vitroPromoted lipid accumulation.[121]
2018miR-221 MEC, MCF-10A, mice In vivo and in vitroPromoted lipid synthesis.[122]
2015miR-486 BMEC, Holstein cows In vivo and in vitroPromoted beta-casein, lactose, and lipid secretion.[123]
Table 3. The role of piRNAs in domestic animals.
Table 3. The role of piRNAs in domestic animals.
Particular YearDetection MethodsSpeciesOutcomesReferences
2021Small RNA-seqPorcineCharacterization of the composition of piRNAs in spermatozoa suggests that piRNAs may be potential negative regulatory markers of sperm quality.[135]
2012Small RNA-seq, qRT-PCRPorcineIt was demonstrated that piRNAs were predominantly enriched in the mature gonads and were expressed more in the testis than in the ovary.[136]
2023Small RNA-seqPorcineExpression of piRNAs is regulated by Senecavirus A (SVA) and promotes apoptosis.[137]
2015Small RNA-seqPorcineCharacterization of the composition of piRNAs in testis suggests that mammalian piRNAs exist in the ping-pong cycle and have a role in the post-transcriptional regulation of protein-coding genes.[130]
2022Small RNA-seqXiang pigsIdentification of the composition of piRNAs in testicular tissues at different stages demonstrates that piRNAs regulate spermatogenesis.[129]
2017Small RNA-seq, qRT-PCRPorcineCharacterization of the expression profiles of testicular piRNAs at different stages of sexual maturation demonstrated that piRNAs regulate testicular development and spermatogenesis.[138]
2012Small RNA-seq, qRT-PCRPorcineEvidence for a potential role of piRNAs in female germ cell development.[139]
2020Small RNA-seq, qRT-PCRPorcineCharacterization of the expression profile of sperm plasma extracellular vesicles (SP-EVs) piRNAs suggests that piRNAs play a role in the physiological function of spermatozoa.[140]
2017Small RNA-seq, qRT-PCRBovidsThe piRNAs in the testis were identified as longer than the piIRNAs in oocytes and embryos.[141]
2020Small RNA-seqYattle, cattle, yaks, Promoter hypermethylation of PIWI/piRNA pathway genes leading to gene silencing and reduction in testis-thick piRNAs is a driver of bovine HMS.[131]
2015Small RNA-seqCalvesExpression of piRNAs in bovine blood and plasma was revealed, suggesting that they may originate from tissues other than blood cells and thus enter the circulation.[142]
2018Small RNA-seqCattle, yaks, dzoComparison of the expression characteristics of three ruminant piRNAs provides theoretical references for exploring their regulatory mechanisms in spermatogenesis and dzo reproductive therapy.[143]
2020Small RNA-seq, qRT-PCRBullsExpression of piRNAs in spermatozoa was detected, suggesting that they may play a role in embryonic development and may serve as biomarkers of semen fertility.[144]
2017Small RNA-seqBullsCharacterization of the composition of piRNAs in frozen spermatozoa suggests a role in sperm development and fertility.[132]
2015Small RNA-seqBovineDetection of the composition of mature testicular and ovarian piRNAs revealed that ovarian piRNAs were very similar to spermatogenesis thick-walled stage piRNAs.[145]
2018Small RNA-seqBovidsDetection of milk exosomal piRNAs expression suggests that they may be related to immune and developmental functions.[134]
2021Small RNA-seq CattleThe presence of piRNAs in ejaculated sperm was confirmed, suggesting that they may regulate sperm maturation, fertilization process, and embryonic genome activation.[146]
2021Small RNA-seqBovidsExpression of piRNAs was detected separately in both milks, suggesting a possible regulatory role in calf immunity and development.[147]
2023Small RNA-seqMurrah buffaloCharacterization of the composition of piRNAs at different stages of lactation implies that piRNAs can serve as potential targets for the regulation of lactation.[148]
2019Small RNA-seqMongolian horseCharacterization of piRNAs composition in sexually mature and immature testes suggests that piRNAs may regulate testicular development and spermatogenesis.[149]
2022Small RNA-seqSheepExpression profiles of piRNAs in LP and FP ovaries were characterized to facilitate understanding of the role of piRNAs in the estrous cycle.[133]
2022Small RNA-seqSheepCharacterization of the composition of testicular piRNAs demonstrates that piRNAs may mediate blood–testis barrier stability and spermatogonial stem cell differentiation.[150]
2021Small RNA-seqSunite (SN), Small-tailed Han (STH)Identification of differential expression of testicular piRNAs in different breeds suggests that piRNAs may be associated with male fecundity.[151]
2023Small RNA-seq, qRT-PCRTibetan sheepCharacterization of piRNAs expression profiles in different stages of testis suggests that piRNAs regulate male fertility and spermatogenesis.[152]
Table 4. The role of piRNAs in breast cancer.
Table 4. The role of piRNAs in breast cancer.
Particular YearpiRNAExpressionModel TypeSpeciesFindingReferences
2021piR-651UpregulationIn vivo and in vitroHumanBound to PIWIL2, promoted cell proliferation and migration through DNMT1-mediated methylation of the PTEN promoter.[158]
2021piR-823UpregulationIn vivo and in vitroHuman and miceIncreased the expression of DNMT1, DNMT3A, and DNMT3B genes to promote DNA methylation of APC genes to activate the Wnt signaling pathway.[159]
2022piR-823UpregulationIn vivo and in vitroHuman and miceInhibited piR-823 expression inhibited cell proliferation, PI3K/Akt/mTOR gene expression, and increased gene and protein expression of ERα.[160]
2013piR-932UpregulationIn vivo and in vitroHuman and miceBound to PIWIL2, promoted methylation of promoter CpG islands to repress Latexin expression.[13]
2023piR-2158DownregulationIn vivo and in vitroHuman and miceCompeted with FOSL1 to inhibit IL-11 expression and secretion, inactivating JAK/STAT signaling and thereby inhibiting breast cancer.[18]
2022piR-17560UpregulationIn vivo and in vitroHumanTargeted FTO-mediated m6A demethylation enhances ZEB1 expression, thereby promoting chemotherapy resistance and EMT.[161]
2013piR-4987, piR-20365, piR-20485, piR-20582UpregulationIn vivoHumanInfluenced cancer development and lymph node metastasis.[154]
2017piR-1282, piR-21131, piR-23672, piR-26526, piR-26527, piR-26528, piR-30293, piR-32745UpregulationIn vivo HumanCan be used as a biomarker for breast cancer and provided a therapeutic target.[156]
piR-23662Downregulation
2014piR-31106UpregulationIn vivo and in vitroHumanResponded to cell growth, cell cycle progression, and hormonal signaling.[155]
2021piR-31106, piR-34998, piR-40067UpregulationIn vivoHumanCan be used as a prognostic and therapeutic marker for breast cancer.[157]
2023piR-31106UpregulationIn vivo and in vitroHumanPromoted cell proliferation and migration as well as oncogene expression and METTL3-mediated m6A methylation.[162]
2025piR-31115UpregulationIn vivo and in vitroHumanBound to PIWIL4 and inhibits the degradation of HSP90AA1 protein, thereby promoting cell migration.[163]
2020piR-31143UpregulationIn vivo and in vitroHumanCan modulation of TNBC behavior through ERβ.[164]
2014piR-34377, piR-35407, piR-36743UpregulationIn vivo and in vitroHumanResponded to cell growth, cell cycle progression, and hormonal signals.[155]
piR-36026, piR-36249, piR-36318, piR-36712Downregulation
2019piR-36712DownregulationIn vivo and in vitroHuman and miceKnockdown of piR-36712 inhibits p53 activity through SEPW1, upregulates Slug/p21, and decreases E-calmodulin levels, ultimately inhibiting cell proliferation, migration, and invasion.[165]
2020piR-016658UpregulationIn vivo and in vitroHumanRegulated by cell Cyclin D1, affects stem cell function.[166]
piR-016975Downregulation
2015piR-021285UpregulationIn vivo and in vitroHumanIncreases the methylation level of the ARHGAP11A gene, which promotes cell invasion and inhibits cell apoptosis.[167]
2015piR-sno75UpregulationIn vivo and in vitroHuman and miceBinding to WDR5 recruits the MLL3/UTX complex to the TRAIL promoter region, thereby inducing H3K4 methylation and H3K27 demethylation.[168]
2022piR-MW557525UpregulationIn vivo and in vitroHumanPromotes the proliferation, migration, and invasion of Piwil2-iCSCs, promotes the expression of CD24, CD133, KLF4, and SOX2, and inhibits apoptosis.[169]
2018piR-FTH1DownregulationIn vivo and in vitroHumanBinds to HILI/HIWI2 and down-regulates FTH1, increasing sensitivity to chemotherapy.[170]
2024piR-YBX1DownregulationIn vivo and in vitroHuman and miceInhibition of YBX1 expression leads to inhibition of MEK and ERK1/2 MAPK signaling pathways, ultimately inhibiting cell proliferation and migration.[171]
Table 5. The role of piRNAs in inflammation.
Table 5. The role of piRNAs in inflammation.
Particular YearpiRNAExpressionResearch TargetFindingReferences
2024hsa-piR-3411, hsa-piR-24541, hsa-piR-27080, hsa-piR-28104, hsa-piR-32157 and 10 othersUpregulationHuman peripheral venous bloodIdentification of piRNAs in the plasma of CP patients demonstrated that piRNAs are associated with inflammation.[189]
hsa-piR-32835, hsa-piR-32836, hsa-piR-32986, hsa-piR-33168Downregulation
2022piRNA-6426DownregulationRat cardiomyocytes, ratsInhibits secretion of inflammatory factors IL-1β and TNF-α, cardiomyocyte apoptosis, oxidative stress, and improves the inflammatory microenvironment in heart failure.[186]
2023piR-has-27620, piR-has-27124UpregulationBlood samplesIdentification of peripheral leukocyte piRNA expression and their enrichment in Rap1, PI3K-Akt, and MAPK pathways as RA biomarkers.[190]
2021rno-piR-017330UpregulationEndothelial cells, ratsIdentification of piRNAs expression in endothelial cells under inflammatory conditions suggests that piRNAs may regulate inflammatory processes.[187]
2024hsa_piR_019949DownregulationC28/I2, SW1353Inhibition of NEAT1 and NLRP3 expression regulates the NOD-like receptor signaling pathway and modulates OA progression.[185]
2024mmu_piR_037459UpregulationMice cardiomyocytes, miceInhibition of collagenase II expression, promotion of chondrocyte apoptosis and inhibition of proliferation, inhibition of USP7 expression, and regulation of OA progression.[188]
2024piR-112710DownregulationMice cardiomyocytes, miceInhibits the Txnip/NLRP3 signaling pathway, reduces the levels of IL-18, IL-1β, and NLRP3, inhibits cardiomyocyte injury, and regulates inflammation progression.[19]
2025pir-has-216911UpregulationHL7702, Huh7, HepG2, Hep3B, nude miceInhibition of the TLR4/NFκB/NLRP3 inflammatory signaling pathway suppressed the inflammatory response.[183]
2020piRNAsDifferential expressionSudani duckThe composition of piRNAs in brain and lung was characterized, suggesting that they may be associated with lung inflammation.[191]
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Yu, W.; Zhang, Z.; Wang, Z.; Dong, X.; Hou, Q. piRNAs as Potential Regulators of Mammary Gland Development and Pathology in Livestock. Vet. Sci. 2025, 12, 594. https://doi.org/10.3390/vetsci12060594

AMA Style

Yu W, Zhang Z, Wang Z, Dong X, Hou Q. piRNAs as Potential Regulators of Mammary Gland Development and Pathology in Livestock. Veterinary Sciences. 2025; 12(6):594. https://doi.org/10.3390/vetsci12060594

Chicago/Turabian Style

Yu, Wenjing, Zixuan Zhang, Zhonghua Wang, Xusheng Dong, and Qiuling Hou. 2025. "piRNAs as Potential Regulators of Mammary Gland Development and Pathology in Livestock" Veterinary Sciences 12, no. 6: 594. https://doi.org/10.3390/vetsci12060594

APA Style

Yu, W., Zhang, Z., Wang, Z., Dong, X., & Hou, Q. (2025). piRNAs as Potential Regulators of Mammary Gland Development and Pathology in Livestock. Veterinary Sciences, 12(6), 594. https://doi.org/10.3390/vetsci12060594

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop