Search Results (20)

Epidural-related maternal fever (ERMF) occurs with significant incidence in women receiving local anesthetics such as ropivacaine via epidural catheter for pain relief during labor. The causal mechanism behind this phenomenon is still not fully resolved, but evidence suggests that these anesthetics cause sterile inflammation. In this observational study, we investigated a possible contributory role of the dual-specificity phosphatase-9 (DUSP9) controlling the activity of mitogen-activated protein kinases (MAPK), and also PH-domain and Leucine-rich repeat phosphatase (PHLPP) regulating AKT kinases. The data show that ropivacaine differentially affects the expression of these phosphatases in distinct cell types of the umbilical cord and placenta. The gene expression of DUSP9 was almost completely switched off in the presence of ropivacaine in HUVECs and extravillous trophoblasts for up to 6 h, while the expression of PHLPP1 was upregulated in HUVECs and syncytiotrophoblasts. Extravillous trophoblasts were identified as a source of pro-inflammatory mediators and regulatory miRNAs in response to ropivacaine. Placentae at term exhibited a distinct DUSP9 expression pattern, whether the patients belonged to the control group or received epidural analgesia with or without elevated body temperature. The observed data imply that ropivacaine induces complex effects on the MAPK and AKT pathways at the feto–maternal interface, which contribute to the ERMF phenomenon. Full article

In the domain of swine production, body weight (BW) and average daily gain (ADG) are recognized as the primary performance indicators. Nevertheless, the genetic architecture of ADG and BW in Dongliao black (DLB) pigs remains to be fully elucidated. In this study, we performed a genome-wide association analysis of BW, ADG, and body mass index (BMI) in 358 DLB pigs of different days of age. The genome-wide association study (GWAS) showed the following: (1) The most significant single nucleotide polymorphism (SNP) detected for BW was on Sus scrofa chromosome (SSC) 11:100,808 (p-value = 1.16 × 10⁻⁶) that was also the most significant SNP for ADG. (2) The most significant SNP associated with BMI was SSC17:51,463,521 (p-value = 5.16 × 10⁻⁸). (3) SNPs SSC10:6,523,844 and SSC17:23,852,682 were identified in both BW and ADG. A meta-analysis was conducted on BW at different days and demonstrated SSC5:39,028,335 (p-value = 8.37 × 10⁻⁶) which was not identified in the results of each single trait. The regions of two SNPs (SSC11:100,808, SSC4:10,703,277) exhibited considerable influence on both BW and ADG and the related regions were selected for linkage disequilibrium (LD) analyses that exhibited a notable linkage. In addition, several genes were identified that are associated with obesity and play roles in lipid metabolism, including MACROD2, PHLPP2, CYP2E1, and STT3B. Full article

Background: Exposure to hyperoxia is an important factor in the development of bronchopulmonary dysplasia (BPD) in preterm newborns. MicroRNAs (miRs) have been implicated in the pathogenesis of BPD and provide a potential therapeutic target. Methods: This study was conducted utilizing a postnatal animal model of experimental hyperoxia-induced murine BPD to investigate the expression and function of miR-195 as well as its molecular signaling targets within developing mouse lung tissue. Results: miR-195 expression levels increased in response to hyperoxia in male and female lungs, with the most significant elevation occurring in 40% O₂ (mild) and 60% O₂ (moderate) BPD. The inhibition of miR-195 improved pulmonary morphology in the hyperoxia-induced BPD model in male and female mice with females showing more resistance to injury and better recovery of alveolar chord length, septal thickness, and radial alveolar count. Additionally, we reveal miR-195-dependent signaling pathways involved in BPD and identify PH domain leucine-rich repeat protein phosphatase 2 (PHLPP2) as a novel specific target protein of miR-195. Conclusions: Our data demonstrate that high levels of miR-195 in neonatal lungs cause the exacerbation of hyperoxia-induced experimental BPD while its inhibition results in amelioration. This finding suggests a therapeutic potential of miR-195 inhibition in preventing BPD. Full article

Tunchang pigs are an indigenous pig population in China known for their high tolerance to roughage, delicious meat, and fecundity. However, the number of Tunchang pigs has been declining due to the influence of commercial breeds and African swine fever, which could potentially lead to inbreeding. To assess the inbreeding level and the genetic basis of important traits in Tunchang pigs, our research investigated the patterns in “runs of homozygosity” (ROHs) using whole genome resequencing data from 32 Tunchang pigs. The study aimed to determine the length, number, coverage, and distribution model of ROHs in Tunchang pigs, as well as genomic regions with high ROH frequencies. The results of the study revealed that a total of 20,499,374 single-nucleotide polymorphisms (SNPs) and 1953 ROH fragments were recognized in 32 individuals. The ROH fragments in Tunchang pigs were predominantly short, ranging from 0.5 to 1 megabases (Mb) in length. Furthermore, the coverage of ROHs varied across chromosomes, with chromosome 3 having the highest coverage and chromosome 11 having the lowest coverage. The genetic diversity of Tunchang pigs was found to be relatively high based on the values of H_E (expected heterozygosity), H_O (observed heterozygosity), pi (nucleotide diversity), Ne (effective population size), and MAF (minor allele frequency). The average inbreeding coefficients of Tunchang pigs, as determined by three different methods (F_HOM, F_GRM, and F_ROH), were 0.019, 0.0138, and 0.0304, respectively. These values indicate that the level of inbreeding in Tunchang pigs is currently low. Additionally, the study identified a total of 13 ROH islands on all chromosomes, which in total contained 38,913 SNPs and 120 genes. These ROH islands included genes associated with economically important traits, including meat quality (GYS1, PHLPP1, SLC27A5, and CRTC1), growth and development (ANKS1A, TAF11, SPDEF, LHB, and PACSIN1), and environmental adaptation (SLC26A7). The findings of this research offer valuable perspectives on the present status of Tunchang pig resources and offer a reference for breeding conservation plans and the efficient utilization of Tunchang pigs in the future. By understanding the inbreeding level and genetic basis of important traits in Tunchang pigs, conservation efforts can be targeted towards maintaining genetic diversity and promoting the sustainable development of this indigenous pig population. Full article

The circadian rhythm regulates biological processes that occur within 24 h in living organisms. It plays a fundamental role in maintaining biological functions and responds to several inputs, including food intake, light/dark cycle, sleep/wake cycle, and physical activity. The circadian timing system comprises a central clock located in the suprachiasmatic nucleus (SCN) and tissue-specific clocks in peripheral tissues. Several studies show that the desynchronization of central and peripheral clocks is associated with an increased incidence of insulin resistance (IR) and related diseases. In this review, we discuss the current knowledge of molecular and cellular mechanisms underlying the impact of circadian clock dysregulation on insulin action. We focus our attention on two possible mediators of this interaction: the phosphatases belonging to the pleckstrin homology leucine-rich repeat protein phosphatase family (PHLPP) family and the deacetylase Sirtuin1. We believe that literature data, herein summarized, suggest that a thorough change of life habits, with the return to synchronized food intake, physical activity, and rest, would doubtless halt the vicious cycle linking IR to dysregulated circadian rhythms. However, since such a comprehensive change may be incompatible with the demand of modern society, clarifying the pathways involved may, nonetheless, contribute to the identification of therapeutic targets that may be exploited to cure or prevent IR-related diseases. Full article

Purpose: In our study, the glucose and cell context-dependent impact of the BMI-1 inhibitor PTC-209 on the AKT pathway in endometrial cancer cells was determined. Methods: The expression of BMI-1 was inhibited by PTC-209 in endometrial cancer cells HEC-1A and Ishikawa stimulated with insulin and grown in different glucose concentrations. The migration, invasion, viability, and proliferative potential after PTC-209 treatment was assessed using wound-healing, Transwell assay, Matrigel-coated inserts, and MTT tests. Chromatin immunoprecipitation was used to determine the localization of BMI-1 protein at promoter sites of the genes tested. Results: BMI-1 inhibition caused an increase in PHLPP1/2 expression and a decrease in phospho-AKT level in both cell lines. The glucose concentration and insulin stimulation differentially impact the AKT pathway through BMI-1 in cells differing in PTEN statuses. The expression of BMI-1 is dependent on the glucose concentration and insulin stimulation mostly in PTEN positive HEC-1A cells. In high glucose concentrations, BMI-1 affects AKT activity through PHLPPs and in hypoglycemia mostly through PTEN. BMI-1 inhibition impacts on genes involved in SNAIL, SLUG, and CDH1 and reduces endometrial cancer cells’ migratory and invasive potential. Conclusions: Our results indicate that the relationship between BMI-1 and phosphatases involved in AKT regulation depends on the glucose concentration and insulin stimulation. Full article

Pleckstrin homology domain and leucine rich repeat protein phosphatase (PHLPP) knockout mice have improved outcomes after a stroke, traumatic brain injury (TBI), and decreased maladaptive vascular remodeling following vascular injury. Thus, small-molecule PHLPP inhibitors have the potential to improve neurological outcomes in a variety of conditions. There is a paucity of data on the efficacy of the known experimental PHLPP inhibitors, and not all may be suited for targeting acute brain injury. Here, we assessed several PHLPP inhibitors not previously explored for neuroprotection (NSC13378, NSC25247, and NSC74429) that had favorable predicted chemistries for targeting the central nervous system (CNS). Neuronal culture studies in staurosporine (apoptosis), glutamate (excitotoxicity), and hydrogen peroxide (necrosis/oxidative stress) revealed that NSC74429 at micromolar concentrations was the most neuroprotective. Subsequent testing in a rat model of asphyxial cardiac arrest, and in a mouse model of severe TBI, showed that serial dosing of 1 mg/kg of NSC74429 over 3 days improved hippocampal survival in both models. Taken together, NSC74429 is neuroprotective across multiple insult mechanisms. Future pharmacokinetic and pharmacodynamic (PK/PD) studies are warranted to optimize dosing, and mechanistic studies are needed to determine the percentage of neuroprotection mediated by PHLPP1/2 inhibition, or potentially from the modulation of PHLPP-independent targets. Full article

Colon cancer is considered as a complex disease that consists of metastatic seeding in early stages. Such disease is not simply caused by the action of a single RNA, but is associated with disorders of many kinds of RNAs and their regulation relationships. Hence, it is of great significance to study the complex regulatory roles among mRNAs, miRNAs and lncRNAs for further understanding the pathogenic mechanism of colon cancer. In this study, we constructed a heterogeneous network consisting of differentially expressed mRNAs, miRNAs and lncRNAs. This contains three kinds of vertices and six types of edges. All RNAs were re-divided into three categories, which were “related”, “irrelevant” and “unlabeled”. They were processed by dynamic excitation restart random walk (RW-DIR) for identifying colon cancer-related RNAs. Ten RNAs were finally obtained related to colon cancer, which were hsa-miR-2682-5p, hsa-miR-1277-3p, ANGPTL1, SLC22A18AS, FENDRR, PHLPP2, hsa-miR-302a-5p, APCDD1, MEX3A and hsa-miR-509-3-5p. Numerical experiments have indicated that the proposed network construction framework and the following RW-DIR algorithm are effective for identifying colon cancer-related RNAs, and this kind of analysis framework can also be easily extended to other diseases, effectively narrowing the scope of biological experimental research. Full article

Corn silk (Stigma Maydis) has been utilized as an important herb against obesity by Chinese, Korean, and Native Americans, but its phytochemicals and mechanisms(s) against obesity have not been deciphered completely. This study aimed to identify promising bioactive constituents and mechanism of action(s) of corn silk (CS) against obesity via network pharmacology. The compounds from CS were identified using Gas Chromatography Mass Spectrometry (GC-MS) and were confirmed ultimately by Lipinski’s rule via SwissADME. The relationships of the compound-targets or obesity-related targets were confirmed by public bioinformatics. The signaling pathways related to obesity, protein-protein interaction (PPI), and signaling pathways-targets-bioactives (STB) were constructed, visualized, and analyzed by RPackage. Lastly, Molecular Docking Test (MDT) was performed to validate affinity between ligand(s) and protein(s) on key signaling pathway(s). We identified a total of 36 compounds from CS via GC-MS, all accepted by Lipinski’s rule. The number of 36 compounds linked to 154 targets, 85 among 154 targets related directly to obesity-targets (3028 targets). Of the final 85 targets, we showed that the PPI network (79 edges, 357 edges), 12 signaling pathways on a bubble chart, and STB network (67 edges, 239 edges) are considered as therapeutic components. The MDT confirmed that two key activators (β-Amyrone, β-Stigmasterol) bound most stably to PPARA, PPARD, PPARG, FABP3, FABP4, and NR1H3 on the PPAR signaling pathway, also, three key inhibitors (Neotocopherol, Xanthosine, and β-Amyrone) bound most tightly to AKT1, IL6, FGF2, and PHLPP1 on the PI3K-Akt signaling pathway. Overall, we provided promising key signaling pathways, targets, and bioactives of CS against obesity, suggesting crucial pharmacological evidence for further clinical testing. Full article

Transmembrane prostate androgen-induced protein (TMEPAI), also known as PMEPA1, is highly expressed in many types of cancer and promotes oncogenic abilities. However, the mechanisms whereby TMEPAI facilitates tumorigenesis are not fully understood. We previously established TMEPAI-knockout (KO) cells from human triple-negative breast cancer (TNBC) cell lines and found that TMEPAI-KO cells showed reduced tumorigenic abilities. Here, we report that TMEPAI-KO cells upregulated the expression of pleckstrin homology (PH) domain and leucine-rich repeat protein phosphatase 1 (PHLPP1) and suppressed AKT Ser473 phosphorylation, which was consistent with TCGA dataset analysis. Additionally, the knockdown (KD) of PHLPP1 in TMEPAI-KO cells partially but significantly rescued AKT Ser473 phosphorylation, as well as in vitro and in vivo tumorigenic activities, thus showing that TMEPAI functions as an oncogenic protein through the regulation of PHLPP1 subsequent to AKT activation. Furthermore, we demonstrated that TMEPAI PPxY (PY) motifs are essential for binding to NEDD4-2, an E3 ubiquitin ligase, and PHLPP1-downregulatory ability. Moreover, TMEPAI enhanced the complex formation of PHLPP1 with NEDD4-2 and PHLPP1 polyubiquitination, which leads to its proteasomal degradation. These findings indicate that the PY motifs of TMEPAI suppress the amount of PHLPP1 and maintain AKT Ser473 phosphorylation at high levels to enhance the tumorigenic potentiality of TNBC. Full article

Prior work demonstrated that Phlpp1 deficiency alters trabecular bone mass and enhances M-CSF responsiveness, but the cell types and requirement of Phlpp1 for this effect were unclear. To understand the function of Phlpp1 within myeloid lineage cells, we crossed Phlpp1 floxed mice with mice harboring LysM-Cre. Micro-computed tomography of the distal femur of 12-week-old mice revealed a 30% increase in bone volume per total volume of Phlpp1 female conditional knockouts, but we did not observe significant changes within male Phlpp1 cKO_LysM mice. Bone histomorphmetry of the proximal tibia further revealed that Phlpp1 cKO_LysM females exhibited elevated osteoclast numbers, but conversely had reduced levels of serum markers of bone resorption as compared to littermate controls. Osteoblast number and serum markers of bone formation were unchanged. In vitro assays confirmed that Phlpp1 ablation enhanced osteoclast number and area, but limited bone resorption. Additionally, reconstitution with exogenous Phlpp1 suppressed osteoclast numbers. Dose response assays demonstrated that Phlpp1^−/− cells are more responsive to M-CSF, but reconstitution with Phlpp1 abrogated this effect. Furthermore, small molecule-mediated Phlpp inhibition enhanced osteoclast numbers and size. Enhanced phosphorylation of Phlpp substrates—including Akt, ERK1/2, and PKCζ—accompanied these observations. In contrast, actin cytoskeleton disruption occurred within Phlpp inhibitor treated osteoclasts. Moreover, Phlpp inhibition reduced resorption of cells cultured on bovine bone slices in vitro. Our results demonstrate that Phlpp1 deficiency within myeloid lineage cells enhances bone mass by limiting bone resorption while leaving osteoclast numbers intact; moreover, we show that Phlpp1 represses osteoclastogenesis and controls responses to M-CSF. Full article

Maternal obesity and gestational diabetes mellitus (GDM) are increasing worldwide, representing risk factors for both mother and child short/long-term outcomes. Oxidative stress, lipotoxicity and altered autophagy have already been reported in obesity, but few studies have focused on obese pregnant women with GDM. Antioxidant and macro/chaperone-mediated autophagy (CMA)-related gene expressions were evaluated herein in obese and GDM placentas. A total of 47 women with singleton pregnancies delivered by elective cesarean section were enrolled: 16 normal weight (NW), 18 obese with no comorbidities (OB GDM(–)), 13 obese with GDM (OB GDM(+)). Placental gene expression was assessed by real-time PCR. Antioxidant gene expression (CAT, GPX1, GSS) decreased, the pro-autophagic ULK1 gene increased and the chaperone-mediated autophagy regulator PHLPP1 decreased in OB GDM(–) vs. NW. On the other hand, PHLPP1 expression increased in OB GDM(+) vs. OB GDM(–). When analyzing results in relation to fetal sex, we found sexual dimorphism for both antioxidant and CMA-related gene expressions. These preliminary results can pave the way for further analyses aimed at elucidating the placental autophagy role in metabolic pregnancy disorders and its potential targetability for the treatment of diabetes outcomes. Full article

Chaperone-mediated autophagy (CMA) is a catabolic pathway fundamental for cell homeostasis, by which specific damaged or non-essential proteins are degraded. CMA activity has three main levels of regulation. The first regulatory level is based on the targetability of specific proteins possessing a KFERQ-like domain, which can be recognized by specific chaperones and delivered to the lysosomes. Target protein unfolding and translocation into the lysosomal lumen constitutes the second level of CMA regulation and is based on the modulation of Lamp2A multimerization. Finally, the activity of some accessory proteins represents the third regulatory level of CMA activity. CMA’s role in oncology has not been fully clarified covering both pro-survival and pro-death roles in different contexts. Taking all this into account, it is possible to comprehend the actual complexity of both CMA regulation and the cellular consequences of its activity allowing it to be elected as a modulatory and not only catabolic machinery. In this review, the role covered by CMA in oncology is discussed with a focus on its relevance in glioma. Molecular correlates of CMA importance in glioma responsiveness to treatment are described to identify new early efficacy biomarkers and new therapeutic targets to overcome resistance. Full article

Canine malignant mammary gland tumors present with a poor prognosis due to metastasis to other organs, such as lung and lymph node metastases. Unlike in human studies where obesity has been shown to increase the risk of breast cancer, this has not been well studied in veterinary science. In our preliminary study, we discovered that leptin downregulated cathepsin A, which is responsible for lysosomal-associated membrane protein 2a (LAMP2a) degradation. LAMP2a is a rate-limiting factor in chaperone-mediated autophagy and is highly active in malignant cancers. Therefore, in this study, alterations in metastatic capacity through cathepsin A by leptin, which are secreted at high levels in the blood of obese patients, were investigated. We used a canine inflammatory mammary gland adenocarcinoma (CHMp) cell line cultured with RPMI-1640 and 10% fetal bovine serum. The samples were then subjected to real-time polymerase chain reaction, Western blot, immunocytochemistry, and lysosome isolation to investigate and visualize the metastasis and chaperone-mediated autophagy-related proteins. Results showed that leptin downregulated cathepsin A expression at both transcript and protein levels, whereas LAMP2a, the rate-limiting factor of chaperone-mediated autophagy, was upregulated by inhibition of LAMP2a degradation. Furthermore, leptin promoted LAMP2a multimerization through the lysosomal mTORC2 (mTOR complex 2)/PH domain and leucine rich repeat protein phosphatase 1 (PHLPP1)/AKT1 (Serine/threonine-protein kinase 1) pathway. These findings suggest that targeting leptin receptors can alleviate mammary gland cancer cell metastasis in dogs. Full article

Recently, we have reported that blockade/deletion of P2X7 receptor (P2X7R), an ATP-gated ion channel, exacerbates heat shock protein 25 (HSP25)-mediated astroglial autophagy (clasmatodendrosis) following kainic acid (KA) injection. In P2X7R knockout (KO) mice, prolonged astroglial HSP25 induction exerts 5′ adenosine monophosphate-activated protein kinase/unc-51 like autophagy activating kinase 1-mediated autophagic pathway independent of mammalian target of rapamycin (mTOR) activity following KA injection. Sustained HSP25 expression also enhances AKT-serine (S) 473 phosphorylation leading to astroglial autophagy via glycogen synthase kinase-3β/bax interacting factor 1 signaling pathway. However, it is unanswered how P2X7R deletion induces AKT-S473 hyperphosphorylation during autophagic process in astrocytes. In the present study, we found that AKT-S473 phosphorylation was increased by enhancing activity of focal adhesion kinase (FAK), independent of mTOR complex (mTORC) 1 and 2 activities in isolated astrocytes of P2X7R knockout (KO) mice following KA injection. In addition, HSP25 overexpression in P2X7R KO mice acted as a chaperone of AKT, which retained AKT-S473 phosphorylation by inhibiting the pleckstrin homology domain and leucine-rich repeat protein phosphatase (PHLPP) 1- and 2-binding to AKT. Therefore, our findings suggest that P2X7R may be a fine-tuner of AKT-S473 activity during astroglial autophagy by regulating FAK phosphorylation and HSP25-mediated inhibition of PHLPP1/2-AKT binding following KA treatment. Full article