Search Results (26)

Ovarian cancer (OC) is the most lethal gynecological cancer globally. Its incidence and mortality consistently rise after menopause. While parity reduces the risk of OC, nulliparity during a woman’s fertile years increases it. Although the association between reproductive history and OC risk is well-established, the long-term impact of pregnancy on the postmenopausal human ovary has received little to no attention. Parity apparently delays the natural decline of the ovarian reserve, but this association also remains unexplored to date. Based on data from cellular, biochemical, and histological markers, as well as epidemiological studies, transcriptomic analyses, and gene knockout mouse models, we review compelling evidence suggesting a critical intraovarian interplay between the residual ovarian reserve and the ovarian surface epithelium (OSE) in the aged ovary. This interaction appears to be a key factor underlying the protective effect of parity on ovarian cancer (OC) risk. We propose that functional FSHR signaling in the remnant follicles of the aged multiparous ovary somehow counteracts the oxidative stress and subsequent chronic inflammation typically observed in the senescent ovary. This mechanism would minimize DNA damage, thereby lowering the probability of neoplastic transformation in the aged mammalian ovary. The precise mechanism by which pregnancy imprints such a long-term follicle–OSE crosstalk warrants further investigation. Full article

Toll-like receptors (TLRs) play crucial roles in innate immunity, but their function in reproduction remains poorly understood. This study investigated the expression patterns and localization of TLR1-9 in the reproductive system of Hu sheep and their potential association with prolificacy. All TLRs were expressed in the oviduct, uterus, and ovary, with TLR6 showing significantly higher expression in the oviduct, while TLR3, TLR6, and TLR7 were predominantly expressed in the ovary. Following this initial screening, we focused on TLR2, TLR6, and TLR7 for detailed analysis. Immunohistochemical analysis revealed that TLR2, TLR6, and TLR7 were localized in the luminal epithelium and circular muscle of the oviduct, the luminal and superficial glandular epithelium of the uterus, and in ovarian follicles at all developmental stages. A comparative analysis between high-prolificacy (HP) and low-prolificacy (LP) Hu sheep demonstrated significantly lower TLR2 expression in the reproductive organs of HP sheep, while TLR6 expression was higher and TLR7 expression was lower in HP ovaries compared to LP ovaries. Notably, TLR7 was observed around apoptotic bodies of granulosa cells, suggesting a potential role in follicular development through the regulation of granulosa cell apoptosis. These findings establish a novel link between innate immunity and reproductive function in sheep, suggesting that TLRs, particularly TLR2, TLR6, and TLR7, may serve dual roles as immune sentinels and reproductive regulators influencing ovine fertility. Full article

Cannabidiol (CBD), a major non-psychoactive cannabinoid of the Cannabis sativa L. plant, has demonstrated anti-inflammatory effects in various studies. However, the therapeutic use of CBD is still limited. Despite its potential, little is known about the molecular mechanisms of CBD on epithelial integrity, particularly concerning effects in native intestinal tissue. To accomplish this, our study aimed to investigate the effects of CBD ex vivo on the follicle-associated epithelium of Peyer’s Patches (PP) and villus epithelium (VE) from porcine intestine. To measure the epithelial barrier, the Ussing chamber technique was employed, followed by immunoblotting and confocal laser-scanning immunofluorescence microscopy of tight junction proteins and specific receptors. The results revealed that CBD significantly strengthens the epithelial barrier of PP by upregulation of sealing tight junction proteins, including occludin, claudin-1, -3, and -7. Additionally, the study showed the potential of CBD to decrease the expression of Tumor necrosis factor alpha (TNFɑ) receptor 1 (TNFR-1) in PP that plays a key role in chronic inflammatory diseases. The study highlights the potential of CBD in the prevention of inflammatory conditions and underlines the important role of PP as a target for bioactive compounds. Full article

Follicle-associated epithelium (FAE) covering the lymphoid follicles of Peyer’s patches (PPs) plays a central role in mucosal immunity. Here, we investigated FAE-derived intrafollicular epithelial bodies (IFEBs) that apparently detach from the FAE and sink deep into the lymphoid tissue of the PPs. Analysis of rabbit PP FAE was carried out by a variety of microscopy and immunohistochemistry techniques using a panel of specific antibodies to determine the nature of the IFEBs. IFEBs displayed the typical features of the FAE, with cytokeratin (CK)+ epithelial cells and CK+/vimentin+ M-cell-like cells. Serial sections of PP tissues showed that the IFEBs are formations frequently separated by the overlying FAE that maintains its integrity. Further, IFEBs showed the presence of junction-associated molecules like zonulin-1 and desmoplakins. Also, IFEBs apparently disaggregate within the lymphoid tissue, as demonstrated by basement membrane disappearance and the finding of isolated epithelial cells that acquire the features of non-polarized epithelial cells. Segments of the FAE in rabbit PPs can detach, forming IFEBs that migrate inside the lymphoid tissue. Although the biological relevance of the newly described IFEBs remains to be determined, we interpreted these data as showing the highly dynamic nature of the PP-associated FAE. Full article

T cell phenotypes and kinetics are emerging as crucial factors associated with immunotherapeutic responses in a wide range of solid cancer types. However, challenges remain in understanding the spatial and temporal profiles of T cells with differential phenotypes due to difficulties in single-cell analysis with preserved tissue structures. Here, we provide an optimized 12-marker multiplex immunohistochemical (IHC) panel and single-cell-based quantitative assessment to identify the spatial distributions of T cell phenotypes in formalin-fixed paraffin-embedded sections. This panel revealed differential T cell populations with spatial localizations in human tonsil tissue, where the percentages of CD8⁺ T cell-expressing programmed death receptor-1 (PD-1), T cell immunoglobulin and mucin domain 3 (TIM3), and other T cell phenotypic markers vary by tonsillar tissue components such as follicles, parenchyma, and epithelium. A specimen from salivary gland adenocarcinoma during hyper-progression, followed by anti-PD-1 treatment, exhibited the exclusion of CD8⁺ T cells from the intratumoral regions. Although the vast majority of peritumoral CD8⁺ T cells exhibited proliferative effector T cell phenotypes with PD-1⁻TIM3⁻Ki67⁺CD45RA⁺, intratumoral CD8⁺ T cells showed exhausted phenotypes with PD-1⁺TIM3⁻ and increased Eomes expression, which might be related to poor therapeutic response in this case. To verify these findings in the context of temporal changes, we analyzed six longitudinal samples from a single patient with maxillary sinus cancer, observing increased T cell exhaustion along with metastasis and progression. Together, highly multiplexed IHC can be applied to analyze the spatiotemporal phenotypes of T cells, potentially contributing to the understanding of the mechanisms of resistance to immunotherapy. Full article

Bovine gammaherpesvirus 6 (BoHV-6) is endemic in cattle in Europe, with a high prevalence. There is evidence that the virus is a commensal and not associated with disease processes. For other gammaherpesviruses, it is known that they have a rather specific target cell spectrum, generally including B cells and, at least in the early phase of infection, the epithelium of the respiratory tract. In a previous study we detected BoHV-6 by quantitative PCR for the gB gene sequence of BoHV-6 in lung, bronchial lymph nodes, spleen and tongue with variable loads, suggesting cells in these tissues as target cells. In the present study, formalin-fixed, paraffin embedded samples of the same tissues from 10 cattle, with high overall BoHV-6 copy numbers, were examined by RNA in situ hybridization for BoHV-6 ORF73. This revealed extremely limited viral ORF73 transcription. A signal was only detected in individual lymphocytes within lymphatic follicles in bronchial lymph nodes, and within very rare alveolar epithelial cells and interstitial cells in the lungs, without any evidence of pathological changes in the tissues. No signal was detected in the spleen or in the oral mucosa of the tongue. The results are consistent with previous findings with other gammaherpesviruses, murine herpesvirus-68, ovine herpesvirus-2 and/or Epstein–Barr virus. They provide further evidence that BoHV-6 is without any consequence to the host and can indeed represent a commensal in cattle. Full article

Fiber diameter is an important characteristic that determines the quality and economic value of rabbit wool. This study aimed to investigate the genetic determinants of wool fiber diameter through an integration analysis using transcriptomic and proteomic datasets from hair follicles of coarse and fine wool from Angora rabbits. Using a 4D label-free technique, we identified 423 differentially expressed proteins (DEPs) in hair follicles of coarse and fine wool in Angora rabbits. Eighteen DEPs were examined using parallel reaction monitoring, which verified the reliability of our proteomic data. Functional enrichment analysis revealed that a set of biological processes and signaling pathways related to wool growth and hair diameter were strongly enriched by DEPs with fold changes greater than two, such as keratinocyte differentiation, skin development, epidermal and epithelial cell differentiation, epidermis and epithelium development, keratinization, and estrogen signaling pathway. Association analysis and protein–protein interaction network analysis further showed that the keratin (KRT) family members, including KRT77, KRT82, KRT72, KRT32, and KRT10, as well as CASP14 and CDSN, might be key factors contributing to differences in fiber diameter. Our results identified DEPs in hair follicles of coarse and fine wool and promoted understanding of the molecular mechanisms underlying wool fiber diameter variation among Angora rabbits. Full article

Peyer’s patches (PPs) are part of the gut-associated lymphatic tissue (GALT) and represent the first line of the intestinal immunological defense. They consist of follicles with lymphocytes and an overlying subepithelial dome with dendritic cells and macrophages, and they are covered by the follicle-associated epithelium (FAE). A sealed paracellular pathway in the FAE is crucial for the controlled uptake of luminal antigens. Quercetin is the most abundant plant flavonoid and has a barrier-strengthening effect on tight junctions (TJs), a protein complex that regulates the paracellular pathway. In this study, we aimed to analyze the effect of quercetin on porcine PPs and the surrounding villus epithelium (VE). We incubated both tissue types for 4 h in Ussing chambers, recorded the transepithelial electrical resistance (TEER), and measured the unidirectional tracer flux of [³H]-mannitol. Subsequently, we analyzed the expression, protein amount, and localization of three TJ proteins, claudin 1, claudin 2, and claudin 4. In the PPs, we could not detect an effect of quercetin after 4 h, neither on TEER nor on the [³H]-mannitol flux. In the VE, quercetin led to a higher TEER value, while the [³H]-mannitol flux was unchanged. The pore-forming claudin 2 was decreased while the barrier-forming claudin 4 was increased and the expression was upregulated. Claudin 1 was unchanged and all claudins could be located in the paracellular membrane by immunofluorescence microscopy. Our study shows the barrier-strengthening effect of quercetin in porcine VE by claudin 4 upregulation and a claudin 2 decrease. Moreover, it underlines the different barrier properties of PPs compared to the VE. Full article

Wool fiber diameter (WFD) is an important index of wool traits and the main determinant of wool quality and value. However, the genetic determinants of fiber diameter have not yet been fully elucidated. Here, coarse and fine wool of Wan strain Angora rabbits and their hair follicle traits were characterized. The results indicated significant differences in the diameters of wool fibers and their hair follicles. The RNA sequencing (RNA-Seq) technique was used to identify differences in gene expression in hair follicles between coarse and fine wool. In total, 2574 differentially expressed genes (DEGs) were found between the two hair follicle groups. Transcription factors, keratin-associated protein (KAP) and keratin (KRT) families, and ECM-related genes may control the structure of fine fibers in rabbits. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that skin development, epidermal cell and keratinocyte differentiation, epithelium development, and Notch and ribosome signaling pathways were significantly enriched, respectively. GSEA further filtered six important pathways and related core genes. PPI analysis also mined functional DEGs associated with hair structure, including LEF1, FZD3, SMAD3, ITGB6, and BMP4. Our findings provide valuable information for researching the molecular mechanisms regulating wool fiber and could facilitate enhanced selection of super-fine wool rabbits through gene-assisted selection in the future. Full article

The primary definition of ovarian aging refers to the loss of follicles. Moreover, the aging of the microenvironment in ovaries, specifically affecting the follicles, may reveal deterioration with advancing age. Besides aging, metabolic disorders associated with hypercaloric diets may affect ovarian health and manifest characteristics associated with premature aging. In this study, we used 10-week-old chinchilla rabbits fed with a high-fat and high-carbohydrate diet (HFCD) until 25 weeks of age to explore hallmarks of reminiscent ovarian aging. The HFCD diet appeared to affect the ovarian reserve, reflected in a significant decrease in primordial follicles. Likewise, Sudan black stain detection revealed substantial differences in the deposits of lipofuscin in the interstitial glands of HFCD-fed rabbits compared to controls, constituting a “hallmark” of aging. The HFCD showed no induced changes in the expression of SOD 2 in the interstitial gland; however, surface epithelium cells were greater expressed. Besides this, the HFCD induced nuclear translocation of NF-ΚΒ p65 factor transcription in surface epithelium cells. We conclude that an HFCD induces a greater accumulation of senescence cells in the interstitial gland, promoting characteristics reminiscent of ovarian aging. However, the activation mechanism of NF-KB caused by an HFCD, which may be stress-responsive and generated by the interstitial gland, requires further study. Full article

Long urethral strictures or even recurrent urethral strictures, mostly with scar tissue showing insufficient healing tendencies, are defined as complex and represent a big challenge in modern reconstructive urology. Initially, the treatment of complicated urethral strictures was associated with a high failure rate (20–40%) due to the growth of hair in the neourethra and a lack of sufficient suitable epithelium when scrotal skin was used. Although much effort was put into tissue engineering recently, harvesting and transplanting autologous tissue represent the standard of care for urethral substitution or augmentation. Since 1977, two-staged urethroplasty with the usage of free foreskin or 0.1 mm thick meshed skin from the upper leg was performed in complicated cases and was initially described in 1984 and 1989 by Schreiter and Schreiter and Noll, respectively. In stage 1, the graft is harvested by cutting the skin thinly above the hair follicles and transplanted as a plate around the opened urethra. In stage 2, after 8–12 weeks, the neourethra is formed. Success rates of up to 84% are described. Considering the complexity of the strictures in which mesh graft urethroplasty is usually performed, the reachable success rates are outstanding, especially considering that this surgery is most likely the last opportunity to prevent perineostomy or even urinary diversion. This article describes the surgical technique and embeds the mesh graft urethroplasty in today’s literature to underline its importance in the surgical management of complex urethral strictures. Full article

Purpose: Precise molecular and cellular mechanisms of radiation-induced dermatitis are incompletely understood. Histone variant H2A.J is associated with cellular senescence and modulates senescence-associated secretory phenotype (SASP) after DNA-damaging insults, such as ionizing radiation (IR). Using ex vivo irradiated cultured foreskin, H2A.J was analyzed as a biomarker of radiation-induced senescence, potentially initiating the inflammatory cascade of radiation-induced skin injury. Methods: Human foreskin explants were collected from young donors, irradiated ex vivo with 10 Gy, and cultured in air-liquid interphase for up to 72 h. At different time-points after ex vivo IR exposure, the foreskin epidermis was analyzed for proliferation and senescence markers by immunofluorescence and immunohistochemical staining of sectioned tissue. Secretion of cytokines was measured in supernatants by ELISA. Using our mouse model with fractionated in vivo irradiation, H2A.J expression was analyzed in epidermal stem/progenitor cell populations localized in different regions of murine hair follicles (HF). Results: Non-vascularized foreskin explants preserved their tissue homeostasis up to 72 h (even after IR exposure), but already non-irradiated foreskin epithelium expressed high levels of H2A.J in all epidermal layers and secreted high amounts of cytokines. Unexpectedly, no further increase in H2A.J expression and no obvious upregulation of cytokine secretion was observed in the foreskin epidermis after ex vivo IR. Undifferentiated keratinocytes in murine HF regions, by contrast, revealed low H2A.J expression in non-irradiated skin and significant radiation-induced H2A.J upregulations at different time-points after IR exposure. Based on its staining characteristics, we presume that H2A.J may have previously underestimated the importance of the epigenetic regulation of keratinocyte maturation. Conclusions: Cultured foreskin characterized by highly keratinized epithelium and specific immunological features is not an appropriate model for studying H2A.J-associated tissue reactions during radiation-induced dermatitis. Full article

Iron deficiency is one of the most prevalent nutritional disorders worldwide. The standard treatment involves iron supplementation, but this task is challenging because of poor solubility and organoleptic issues. Moreover, the need to increase iron bioavailability represents a challenge for treating iron-related disorders. In this study, gastroresistance and iron intestinal absorption of an innovative granular formulation composed of ferric pyrophosphate, modified starch and phospholipids branded as Ferro Fosfosoma^® was investigated. Gastroresistant properties were studied using standard protocols, and a bioaccessible fraction was obtained by exposing a food supplement to in vitro digestion. This fraction was used for investigating iron absorption in Caco-2 and human follicle-associated intestinal epithelium (FAE) models. Ferro Fosfosoma^® showed an improved resistance to gastric digestion and higher intestinal absorption than ferric pyrophosphate salt used as a control in both models. In the FAE model, Ferro Fosfosoma^® induces larger iron absorption than in the Caco-2 monolayer, most likely due to the transcytosis ability of M cells. The larger iron absorption in the Ferro Fosfosoma^®-treated FAE model corresponds to higher ferritin level, proving physiological iron handling that was once delivered by granular formulation. Finally, the formulation did not induce any alterations in viability and barrier integrity. To conclude, Ferro Fosfosoma^® favors iron absorption and ferritin expression, while preserving any adverse effects. Full article

Gut-associated lymphoid tissue is one of the most diverse and complex immune compartments in the human body. The subepithelial compartment of the gut consists of immune cells of innate and adaptive immunity, non-hematopoietic mesenchymal cells, and stem cells of different origins, and is organized into secondary (and even tertiary) lymphoid organs, such as Peyer’s patches, cryptopatches, and isolated lymphoid follicles. The function of isolated lymphoid follicles is multifaceted; they play a role in the development and regeneration of the large intestine and the maintenance of (immune) homeostasis. Isolated lymphoid follicles are also extensively associated with the epithelium and its conventional and non-conventional immune cells; hence, they can also function as a starting point or maintainer of pathological processes such as inflammatory bowel diseases or colorectal carcinogenesis. These relationships can significantly affect both physiological and pathological processes of the intestines. We aim to provide an overview of the latest knowledge of isolated lymphoid follicles in colonic inflammation and colorectal carcinogenesis. Further studies of these lymphoid organs will likely lead to an extended understanding of how immune responses are initiated and controlled within the large intestine, along with the possibility of creating novel mucosal vaccinations and ways to treat inflammatory bowel disease or colorectal cancer. Full article

Enteric glial cells (EGC) are known to regulate gastrointestinal functions; however, their role in Crohn’s disease (CD) is elusive. Microscopic erosions over the ileal Peyer’s patches are early signs of CD. The aim of this work was to assess the localization of EGC in the follicle and interfollicular region of the Peyer’s patches and in the lamina propria and study the effects of EGC mediators on barrier function in CD patients and non-inflammatory bowel disease (non-IBD) controls. EGC markers, glial fibrillary acidic protein (GFAP), and S100 calcium-binding protein β (S100β) were quantified by immunofluorescence and Western blotting. Both markers showed significantly more EGC in the Peyer’s patches and lamina propria of CD patients compared to the non-IBD controls. In CD patients there were significantly more EGC in Peyer’s patches compared to lamina propria, while the opposite pattern was seen in controls. Barrier function studies using Ussing chambers showed increased paracellular permeability by EGC mediators in CD patients, whereas permeability decreased by the mediators in controls. We show the accumulation of EGC in Peyer’s patches of CD patients. Moreover, EGC mediators induced barrier dysfunction in CD patients. Thus, EGC might have harmful impacts on ongoing inflammation and contribute to the pathophysiology of the disease. Full article