Expression Of PCNA Research Articles

Overexpressed microRNA (miR)-382-3p promoted vascular remodeling via suppressing autophagy-related protein 7 (ATG7) in chronic thromboembolic pulmonary hypertension.

This research has investigated the role of miR-382-3p in chronic thromboembolic pulmonary hypertension (CTEPH). Human pulmonary artery smooth muscle cells (hPASMCs) were treated with PDGF-BB to induce proliferation, and then transfected with miR-382-3p mimic, miR-382-3p inhibitor, ATG7 overexpression plasmid, and siATG7. MiR-382-3p, ATG7, VEGF, PCNA, p62, and LC3-Ⅱ/LC3-I levels were detected by qRT-PCR and Western blotting. Cell viability and migration were tested through CCK-8 and wound healing assays, respectively. Target genes of miR-382-3p were predicted by Targetscan and starBase, and pathway analysis was implemented through WebGestalt. The binding relationship between miR-382-3p and ATG7 was analyzed by the dual-luciferase reporter and RIP assays. A CTEPH model was constructed in rats with the treatment of miR-382-3p antagomir or agomir, and mean pulmonary artery pressure (mPAP) was measured. Lung tissue was observed through the HE staining assay. MiR-382-3p level in hPASMCs was obviously upregulated with the increasing dose of PDGF-BB. MiR-382-3p mimic promoted yet miR-382-3p inhibitor suppressed hPASMC proliferation. MiR-382-3p directly targeted ATG7. ATG7 overexpression repressed hPASMC proliferation and migration, whereas siATG7 exerted the opposite effects. ATG7 overexpression partly neutralized the effects of miR-382-3p mimic on proliferation, migration, and autophagy-related proteins (ATG7, p62, and LC3-Ⅱ/LC3-I) in hPASMCs, whereas siATG7 partly offset the impacts of miR-382-3p inhibitor. MiR-382-3p antagomir reversed CTEPH-induced mPAP elevation, miR-382-3p upregulation, thickening of the pulmonary artery wall, and increased expressions of VEGF, PCNA, and autophagy-related proteins in rats, while miR-382-3p agomir potentiated these effects induced by CTEPH. Overexpressed miR-382-3p promotes vascular remodeling via ATG7 inhibition in CTEPH.

Itraconazole inhibits proliferation, induces apoptosis, and reduces angiogenesis of hemangioma endothelial cells by downregulating the hedgehog signaling pathway

Infantile hemangioma (IH) is among the most prevalent benign vascular tumours in infants. The pathogenesis of IH mainly involves abnormal proliferation of vascular endothelial cells and the formation of new vessels. Itraconazole was shown to be effective in treating IH; however, the mechanism underlying its action is still unclear. The purpose of this study was to examine the effects of itraconazole on the proliferation, apoptosis, and angiogenesis of hemangioma endothelial cells (HemECs); human umbilical vein endothelial cells served as the control group. The expression of genes involved in the hedgehog (HH) signaling pathway (SHH, PTCH1, SMO, and GLI1) was determined using real-time quantitative polymerase chain reaction. Western blotting was used to determine the expression of related proteins. In this study, itraconazole significantly dose- and time-dependently inhibited the viability of HemECs. Itraconazole suppressed the expression of PCNA, Ki67, and vascular endothelial growth factor (VEGF), demonstrating that this treatment inhibited cell proliferation and angiogenesis. Moreover, itraconazole induced apoptosis of HemECs by activating the expression of BAX and inhibiting the expression of BCL2. Itraconazole inhibited SHH, PTCH1, SMO, and GLI1 expression. Activation of the HH pathway by recombinant human sonic hedgehog (rhSHH) protein attenuated the effect of itraconazole on HemECs. In conclusion, itraconazole inhibits proliferation, induces apoptosis, and reduces angiogenesis of HemECs via the downregulation of the HH signaling pathway. Therefore, itraconazole may be an alternative choice for the treatment of IH.

Long-term abuse of caffeine sodium benzoate induces endothelial cells injury and leads to coagulation dysfunction.

Our hospital admitted a patient who had difficulty in coagulation even after blood replacement, and the patient had abused caffeine sodium benzoate (CSB) for more than 20 years. Hence, we aimed to explore whether CSB may cause dysfunction in vascular endothelial cells and its possible mechanism. Low, medium, and high concentrations of serum of long-term CSB intake patients were used to treat HUVECs, with LPS as the positive control. MTT and CCK8 were performed to verify CSB's damaging effect on HUVECs. The expression of ET-1, ICAM-1, VCAM-1, and E-selectin were measured by ELISA. TUNEL assay and Matrigel tube formation assay were carried out to detect apoptosis and angiogenesis of HUVECs. Flow cytometry was applied to analyze cell cycles and expression of CD11b, PDGF, and ICAM-1. Expression of PDGF-BB and PCNA were examined by western blot. The activation of MAPK signaling pathway was detected by qRT-PCR and western blot. Intracellular Ca2+ density was detected by fluorescent probes. CCK8 assay showed high concentration of CSB inhibited cell viability. Cell proliferation and angiogenesis were inhibited by CSB. ET-1, ICAM-1, VCAM-1, and E-selectin upregulated in CSB groups. CSB enhanced apoptosis of HUVECs. CD11b, ICAM-1 increased and PDGF reduced in CSB groups. The expression level and phosphorylation level of MEK, ERK, JUN, and p38 in MAPK pathway elevated in CSB groups. The expression of PCNA and PDGF-BB was suppressed by CSB. Intracellular Ca2+ intensity was increased by CSB. Abuse of CSB injured HUVECs and caused coagulation disorders.

Obesity Contributes to Transformation of Myometrial Stem-Cell Niche to Leiomyoma via Inducing Oxidative Stress, DNA Damage, Proliferation, and Extracellular Matrix Deposition.

Leiomyomas (fibroids) are monoclonal tumors in which myometrial stem cells (MSCs) turn tumorigenic after mutation, abnormal methylation, or aberrant signaling. Several factors contribute to metabolic dysfunction in obesity, including abnormal cellular proliferation, oxidative stress, and DNA damage. The present study aims to determine how adipocytes and adipocyte-secreted factors affect changes in MSCs in a manner that promotes the growth of uterine leiomyomas. Myometrial stem cells were isolated from the uteri of patients by fluorescence-activated cell sorting (FACS) using CD44/Stro1 antibodies. Enzyme-linked immunosorbent assay (ELISA), Western blot, and immunocytochemistry assays were performed on human adipocytes (SW872) co-cultured with MSCs and treated with leptin or adiponectin to examine the effects of proliferation, extracellular matrix (ECM) deposition, oxidative damage, and DNA damage. Co-culture with SW872 increased MSC proliferation compared to MSC culture alone, according to 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) results. The expressions of PCNA and COL1A increased significantly with SW872 co-culture. In addition, the expression of these markers was increased after leptin treatment and decreased after adiponectin treatment in MSCs. The Wnt/β-catenin and TGF-β/SMAD signaling pathways promote proliferation and ECM deposition in uterine leiomyomas. The expression of Wnt4, β-catenin, TGFβ3, and pSMAD2/3 of MSCs was increased when co-cultured with adipocytes. We found that the co-culture of MSCs with adipocytes resulted in increased NOX4 expression, reactive oxygen species production, and γ-H2AX expression. Leptin acts by binding to its receptor (LEP-R), leading to signal transduction, resulting in the transcription of genes involved in cellular proliferation, angiogenesis, and glycolysis. In MSCs, co-culture with adipocytes increased the expression of LEP-R, pSTAT3/STAT3, and pERK1/2/ERK/12. Based on the above results, we suggest that obesity may mediate MSC initiation of tumorigenesis, resulting in leiomyomas.

Carvacrol showed a curative effect on reproductive toxicity caused by Bisphenol AF via antioxidant, anti-inflammatory and anti-apoptotic properties

Bisphenol AF (BPAF) is an endocrine disruptor, and human exposure to these chemicals is growing in industrialized nations. BPAF has been demonstrated in studies to have toxic effects on reproductive health. This study examined the effects of oral exposure to BPAF on the reproductive system and the protective effects of carvacrol in rats. From 32 Wistar albino rats, four separate groups were set up for this purpose. Carvacrol 75 mg/kg and BPAF 200 mg/kg were administered by oral gavage method. Rat sperm parameters and serum testosterone levels were measured after 28 days of administration. The study looked at the MDA in the testis tissues, as well as CAT, GPx, and GSH as antioxidants parameters, NF-κB and TNF-α as inflammatory markers, caspase-3 and Bcl-2 as apoptosis parameters, and PCNA as cell proliferation markers. In addition, testis tissues underwent histological evaluation. As a result, in rats exposed to only BPAF, sperm counts declined, testosterone levels reduced, oxidative stress, inflammation, and apoptosis increased, and cell proliferation decreased. Furthermore, severe disruptions in tissue architecture and decreased spermatogenesis were reported. In contrast, sperm parameters improved, testosterone levels increased, oxidative stress and inflammation decreased, and apoptosis was prevented in the carvacrol-treated group compared to the BPAF-only group. It was also found that spermatogenesis was maintained, and structural abnormalities in testicular tissue were mostly avoided with an increase in PCNA expression. According to the findings, despite BPAF-induced testicular and reproductive toxicity, carvacrol had therapeutic potential due to its anti-inflammatory, antioxidant, cell proliferation-increasing, and anti-apoptotic activities.

Human umbilical cord mesenchymal stem cells (hUC-MSCs) alleviate paclitaxel-induced spermatogenesis defects and maintain male fertility

Chemotherapeutic drugs can cause reproductive damage by affecting sperm quality and other aspects of male fertility. Stem cells are thought to alleviate the damage caused by chemotherapy drugs and to play roles in reproductive protection and treatment. This study aimed to explore the effects of human umbilical cord mesenchymal stem cells (hUC-MSCs) on alleviating paclitaxel (PTX)-induced spermatogenesis and male fertility defects. An in vivo PTX-induced mice model was constructed to evaluate the reproductive toxicity and protective roles of hUC-MSCs in male fertility improvement. A 14 day PTX treatment regimen significantly attenuated mice spermatogenesis and sperm quality, including affecting spermatogenesis, reducing sperm counts, and decreasing sperm motility. hUC-MSCs treatment could significantly improve sperm functional indicators. Mating experiments with normal female mice and examination of embryo development at 7.5 days post-coitum (dpc) showed that hUC-MSCs restored male mouse fertility that was reduced by PTX. In IVF experiments, PTX impaired sperm fertility and blastocyst development, but hUC-MSCs treatment rescued these indicators. hUC-MSCs’ protective role was also displayed through the increased expression of the fertility-related proteins HSPA2 and HSPA4L in testes with decreased expression in the PTX-treated group. These changes might be related to the PTX-induced decreases in expression of the germ cell proliferation protein PCNA and the meiosis proteins SYCP3, MLH1, and STRA8, which were restored after hUC-MSCs treatment. In the PTX-treated group, the expression of testicular antioxidant proteins SIRT1, NRF2, CAT, SOD1, and PRDX6 was significantly decreased, but hUC-MSCs could maintain these expressions and reverse PTX-related increases in BAX/BCL2 ratios. hUC-MSCs may be a promising agent with antioxidant and anti-apoptosis characteristics that can maintain sperm quality following chemotherapy treatment.

Regulation of survival, growth performance, and intestinal health of large yellow croaker (Larimichthys crocea) larvae by sodium decanoate supplementation

A 30-day feeding trial was undertaken to elucidate the impact of diets supplemented with sodium decanoate (Sd) on survival, growth performance, and intestinal health in Larimichthys crocea larvae. A series of experimental diets consisting of graduated levels of Sd were formulated, including 0.0% (the control), 0.1%, 0.2%, 0.4%, or 0.8%. The study revealed no significant variations in survival among dietary groups. The growth performance (final body weight and specific growth rate) exhibited significant linear and quadratic trends, with the highest values observed in larvae fed a diet containing 0.2% Sd, which was significantly higher than that of the control group. Notably, Larvae fed diets with 0.2%–0.8% Sd had significantly higher intestinal fold height than the control group. A finding that was substantiated by the increased mRNA expression of pcna and odc, which are known to reflect processes of epithelial proliferation and differentiation. Meanwhile, the significant increase in the activity of the intestinal brush border membrane enzyme (Leucine-aminopeptidase), observed in diets supplemented with 0.1%–0.8% Sd, provides compelling evidence supporting the promotion of intestinal development and maturation. In terms of digestive and absorptive capacity, dietary supplementation with 0.4% Sd significantly increased the activity of trypsin and creatine kinase, while 0.1%–0.4% Sd specifically enhanced the activity of Na+-K+-Adenosinetriphosphatase than the control group. Importantly, larvae consuming Sd-supplemented diets at appropriate levels exhibited increased mRNA expression of tight junction-associated genes. Additionally, diets supplemented appropriate Sd improved the antioxidant capacity by modulating antioxidant enzymes and genes, antioxidant substances, and lipid peroxidation products. In summary, the diet incorporating 0.2% Sd could potentially boost the growth of larvae, which may be connected to Sd promoting intestinal health by regulating intestinal morphology and development, digestion and absorption capacity, intestinal barrier function, and antioxidant capacity. This study sheds light into the application of medium-chain fatty acids in marine fish larvae and contributes to the development of nutritional regulation strategies aimed at enhancing larval development.

SYST-23 CDK1 AS AN INDEPENDENT PROGNOSTIC INDICATOR AND POTENTIAL THERAPEUTIC TARGET IN GLIOMA

Abstract Glioma is the most common primary malignant tumor in the brain with extremely poor prognosis. Firstly, in order to find the important genes and mechanisms involved in the initiation and progression of glioma, we selected four groups of gene expression profiles from GEO dataset include GSE29796, GSE4290, GSE50161 and GSE7696 for comprehensive data analysis. We identified 360 differentially expressed genes (DEGs), then we screened out core gene which is CDK1 by protein-protein interaction (PPI) network analysis. So as to clarify whether CDK1 is an independent prognostic factor for glioma, we performed expression analysis and prognostic analysis. Meanwhile, in order to verify the expression and survival of CDK1, we used Western blot to detect CDK1 expression of 15 glioma specimens and corresponding adjacent tissues. Moreover, we divided the expression values of CDK1 gene in the CGGA database into high and low groups based on the median for differential expression analysis. Finally, to explore the molecular mechanism of action of CDK1 in gliomas, we conducted gene ontology (GO) enrichment analysis and gene collection enrichment analysis (GSEA),and performed gene co-expression analysis. The results revealed that the expression level of CDK1 in glioma was significantly higher than normal tissues, its high expression was closely associated with poor survival of glioma. And the prognostic analysis showed that CDK1 was an independent prognostic index of glioma. Through the mechanism analysis of GO and GSEA, it is demonstrated that CDK1 plays an important role in the mitosis cell cycle of glioma, and inhibiting the expression of CDK1 can affect the proliferation of glioma cells. Coexpression analysis displayed that the high expression of PCNA, MCM2, MCM3, MCM4, MCM6, PLK1, TTK and MAD2L1 are also correlated to the poor survival and prognosis of glioma. Hence CDK1 is an independent prognostic factor and potential therapeutic target of glioma. Furthermore ,we can affect the proliferation of glioma cells through inhibiting the expression of CDK1.

BFNB Enhances Hair Growth in C57BL/6 Mice through the Induction of EGF and FGF7 Factors and the PI3K-AKT-β-Catenin Pathway.

The objective of this study was to investigate the potential effects of a formulation derived from the bioactive fraction of nanostructured Bacopa procumbens (BFNB) on the promotion of hair growth in C57BL/6 mice. The characterization of the follicular phases and histomorphological analysis showed that the topical application of the formulation for 15 days significantly increased pigmentation and hair growth on the dorsum and head of the mice. Additionally, an acceleration of the follicular cycle phases was observed, along with an increase in the number of follicles, hair length, and diameter, compared to mice treated with minoxidil. In silico analysis and molecular characterization demonstrated that BFNB enhances the expression of epidermal growth factor (EGF) and fibroblast growth factor 7 (FGF7), activating the PI3K-AKT-β-catenin signaling pathway, as well as the expression of PCNA, KI-67, Cyclin D1, and Cyclin E, regulating the cell cycle and cell proliferation, crucial events for hair regeneration. Our results strongly suggest the utility of BFNB as a therapeutic alternative to stimulate hair growth and promote hair health.

FOXO1 regulates the formation of bovine fat by targeting CD36 and STEAP4

Intramuscular fat content is closely related to the quality of beef, where the forkhead box protein O1 (FOXO1) is involved in adipocyte differentiation and lipid metabolism, but the specific mechanism of its involvement is still unclear. In this study, interfering with FOXO1 promoted the G1/S transformation of bovine adipocytes by enhancing the expression of proliferation marker genes PCNA, CDK1, CDK2, CCNA2, CCNB1, and CCNE2, thereby positively regulating the proliferation of bovine adipocytes. Additionally, interfering with FOXO1 negatively regulated the expression of adipogenic differentiation marker genes PPARG and CEBPA, as well as lipid anabolism marker genes ACC, FASN, SCD1, SREBP1, FABP4, ACSL1, LPL, and DGAT1, thus reducing triglyceride (TG) content and inhibiting the generation of lipid droplets in bovine adipocytes. A combination of transcriptomic and metabolomics analyses revealed that FOXO1 could regulate the lipogenesis of cattle by influencing the AMPK and PI3K/AKT pathways. Importantly, chromatin immunoprecipitation (ChIP) and site-directed mutagenesis revealed that FOXO1 could regulate bovine lipogenesis by binding to the promoter regions of the CD36 and STEAP4 genes and affecting their transcriptional activities. These results provide a foundation for studying the role and molecular mechanism of FOXO1 in the bovine adipogenesis.

Based on network pharmacology, gastrodin attenuates hypertension-induced vascular smooth muscle cell proliferation and PI3K/AKT pathway activation

The effects and underlying mechanisms of gastrodin treatment on hypertensive vascular dysfunction and proliferation of vascular smooth muscle cells (VSMCs) were determined in vitro and in vivo. Using a pharmacological target network interaction analysis, 151 common targets and a PPI network were identified containing the top 10 hub genes. Kyoto encyclopedia of genes and genomes (KEGG) analysis identified the PI3K/AKT pathway as a significantly enriched pathway. Both spontaneous hypertensive rats (SHRs) and Wistar Kyoto rats were used to assess the therapeutic effects of gastrodin on hypertension. Gastrodin treatment of the SHRs resulted in a marked attenuation of elevated blood pressure, pulse wave velocity, and pathological changes in the abdominal aorta. Moreover, gastrodin treatment significantly inhibited cell growth and downregulated the expression of PCNA as well as the p-PI3K/PI3K and p-AKT/AKT levels in angiotensin II-stimulated VSMCs. Taken together, gastrodin treatment attenuates blood pressure elevation, vascular dysfunction, and proliferation of VSMCs and inhibits the activation of the PI3K/AKT pathway.

Knockdown of DEC2 expression inhibits the proliferation of mesangial cells through suppressed glycolysis and p38 MAPK/Erk pathway in lupus nephritis

BackgroundTo elucidate the mechanism by which DEC2 modulates the proliferation of mesangial cells (MCs) in lupus nephritis (LN).MethodsThe 32-week-old female Fcgr2b−/− mice and their serum-treated MCs were used as in vivo and in vitro LN model. MCs knocked down of DEC2 and overexpressed with DEC2 were also established. The expression of DEC2 was measured in the kidneys of Fcgr2b−/− mice and LN serum-treated MCs using RT-qPCR and Western blot. MCs proliferation was detected by 5-ethynyl-2′-deoxyuridine (EdU) labeling assay and PCNA expression using immunofluorescence. The glucose metabolism was evaluated in LN serum-treated MCs, and the levels of lactate production, glucose consumption, ATP production and mitochondrial membrane potential were assayed. The glycolysis and mitochondrial respiration function of the MCs were measured using the Extracellular Flux Analyzer. The extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were dynamically monitored and multiple important bioenergetic parameters can be calculated. The expression of Toll like receptor 4 (TLR4) and glucose transporter 1 (GLUT1) were detected in the MCs. Multiple signaling proteins were screened.ResultsDEC2 was found overexpressed in the kidney of Fcgr2b−/− LN mice. Knockdown of DEC2 inhibited LN serum-induced MCs proliferation. DEC2 was associated with the glucose metabolism in LN serum-treated MCs. DEC2 regulated glycolysis in LN serum-treated MCs. DEC2 was associated with mitochondrial fitness in LN serum treated MCs. DEC2 activated MCs glycolysis through TLR4 and glucose transporter 1 (GLUT1) regulation. DEC2 regulated MCs proliferation through two signaling pathways including dependent and independent of glycolysis, which located in the downstream of TLR4 signaling.ConclusionKnockdown of DEC2 expression inhibits the proliferation of MCs through suppressed glycolysis and p38 MAPK/ERK pathway in LN.

Modulation of the Proliferative Pathway, Neuroinflammation and Pain in Endometriosis.

Endometriosis is a chronic disease characterized by pelvic inflammation. This study aimed at investigating the molecular mechanisms underlying the pathology and how they can be modulated by the administration of a natural compound, Actaea racemosa (AR). We employed an in vivo model of endometriosis in which rats were intraperitoneally injected with uterine fragments from donor animals. During the experiment, rats were monitored by abdominal high-frequency ultrasound analysis. AR was able to reduce the lesion's size and histological morphology. From a molecular point of view, AR reduced hyperproliferation, as shown by Ki-67 and PCNA expression and MAPK phosphorylation. The impaired apoptosis pathway was also restored, as shown by the TUNEL assay and RT-PCR for Bax, Bcl-2, and Caspase levels. AR also has important antioxidant (reduced Nox expression, restored SOD activity and GSH levels, and reduced MPO activity and MDA levels) and anti-inflammatory (reduced cytokine levels) properties. Moreover, AR demonstrated its ability to reduce the pain-like behaviors associated with the pathology, the neuro-sensitizing mediators (c-FOS and NGF) expression, and the related central astrogliosis (GFAP expression in the spinal cord, brain cortex, and hippocampus). Overall, our data showed that AR was able to manage several pathways involved in endometriosis suppression.

Aqueous M. oleifera leaf extract alleviates DSS-induced colitis in mice through suppression of inflammation

Ethnopharmacological relevanceMoringa oleifera Lam. (M. oleifera) is a perennial deciduous tree with considerable agricultural and pharmacological value. Nearly all parts of the tree are edible, and nearly all parts are used in traditional medicine. Leaves of M. oleifera have the functions of hypoglycemic (antidiabetic), anti-cancer and anti-oxidant stress, but less research pay attention to the anti-inflammatory effect of M. oleifera leaves. Aim of the studyInflammatory bowel disease (IBD) is a chronic and relapsing inflammatory disorder of the gut with no ideal medication. Here, we investigated the anti-inflammatory effects of aqueous extract of M. oleifera leaves. Materials and methodsIntestinal organoids and mice as in vitro and in vivo models to investigate the effects of aqueous extract of M. oleifera leaves on inflammation induced by TNF-α and dextran sulfate sodium (DSS) respectively. The expression of inflammatory cytokines and proliferation-related genes were evaluated by RT-qPCR, respectively. The compounds in the leaf extract were determined by LC/MS, and network pharmacology approach was employed to predict 54 anti-IBD potential targets of quercetin-3-galactoside (QG) and isoquercitrin (IS). ResultsWe found that the extract protected against damage to intestinal organoids caused by tumor necrosis factor (TNF-α), and significantly down-regulated the expression of inflammatory cytokines. The extract also suppressed the TNF-α-induced expression of Pcna, c-Myc, and c-Jun. Additionally, oral administration of the extract also ameliorated DSS-induced colon damage (colonic shortening, loss of goblet cells and overall abnormal cellularity), and inhibited the expression of inflammatory cytokines and proliferation-related genes in colitis. By LC/MS we identified nearly 2000 of the compounds in the leaf extract, of the flavonoids identified, QG and IS made up the largest percentage; both have been shown to have anti-inflammatory properties. Moreover, network pharmacology approach was employed to predict 54 anti-IBD potential targets of QG and IS. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that the overlapping targets participated in response to oxidative stress and PI3K−Akt signaling pathway respectively. ConclusionsThe present study demonstrated the anti-inflammatory capability, in vitro and in vivo, of the aqueous extract of M. oleifera leaves and suggests its potential phytotherapeutic treatment for IBD.

Thymoquinone attenuates doxorubicin-induced lung damage via heat shock proteins, inflammation and endoplasmic reticulum stress

The aim of this study is to determine possible therapeutic effects of THQ on doxorubicin induced lung damage in rats. Rats were divided into five groups (n = 8): control, THQ, olive oil, DOX (a single dose of 15 mg/kg intraperitoneally (i.p.) on seventh day of the experiment), and DOX+THQ (10 mg/kg THQ per day and 15 mg/kg DOX i.p. on seventh day). Animals were euthanized, and lung tissues were evaluated histopathologically. Caspase3, GRP78, GADD153, HSP90, HSP70, PCNA, COX2 and TNFα immunostaining were performed to determine the expression levels of these proteins among groups. TUNEL method was used for evaluation of apoptotic index. Moreover, total antioxidant status (TAS), total oxidant status (TOS) and IL6 in lung tissue were measured by ELISA assay. The DOX group had histopathological deterioration compared to the control group. There was an increase in apoptotic index, caspase 3, GRP78, GADD153, HSP90, HSP70, PCNA, COX2 and TNFα expressions in the DOX group. While TAS level of the DOX group decreased, IL6 and TOS level increased when compared with the other groups. However, there was improvement in lung tissue in DOX + THQ group compared to the DOX group. There was a decrease in apoptotic index, caspase 3, GRP78, GADD153, HSP90, HSP70, PCNA, COX2, TNFα expressions and TOS, IL6 in DOX+THQ group compared to the DOX group. We suggest that THQ can be used as a protective agent to reduce the toxic effects of DOX.

Involvement of Ataxin-3 (ATXN3) in the malignant progression of pancreatic cancer via deubiquitinating HDAC6

BackgroundPancreatic cancer is a common digestive system cancer and one of the most lethal malignancies worldwide. Ataxin-3 (ATXN3) protein is a deubiquitinating enzyme implicated in the occurrence of diverse human cancers. The potential role of ATXN3 in pancreatic cancer still remains unclear. MethodsATXN3 was screened from differentially-upregulated genes of GSE71989, GSE27890 and GSE40098 datasets. The mRNA and protein levels of ATXN3 was evaluated in pancreatic cancer samples and cell lines. Through the gain- and loss-of-function experiments, the effects of ATXN3 on cell proliferation, migration and invasion were evaluated using cell counting kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU) staining, wound healing and Transwell assays. Subsequently, the interaction between ATXN3 and HDAC6 was confirmed using double immunofluorescence staining, co-immunoprecipitation (co-IP) and proximity ligation assay (PLA). The underlying mechanism of ATXN3 was determined by knockdown of HDAC6 in ATXN3-upregulated pancreatic cancer cells. The function of ATXN3 in vivo was verified through xenograft assay. ResultsHigh expression of ATXN3 was found in pancreatic cancer tissues. Increased ATXN3 expression dramatically promoted cell proliferation, migration, and invasion. The malignant phenotypes were suppressed in ATXN3-silenced pancreatic cancer cells. ATXN3 was proved to interact with HDAC6 and regulate its degradation through deubiquitination. Downregulation of HDAC6 inhibited ATXN3-induced development of pancreatic cancer cells through regulating the expression of PCNA, vimentin and E-cadherin. ATXN3 facilitated tumor growth of pancreatic cancer and increased HDAC6 expression in vivo. ConclusionsThis study confirmed that ATXN3 facilitated malignant phenotypes of pancreatic cancer via reducing the ubiquitination of HDAC6.

Toxicity of methomyl insecticides to testicular cells and protective effect of folic acid.

Methomyl is a carbamate insecticide with confirmed testicular toxicity. This study intended to observe the effect of methomyl on testicular cells and the protective effect of folic acid through in vitro experiments. The GC-1 spermatogonia, TM4 Sertoli cells, and TM3 Leydig cells were treated with methomyl (0, 250, 500, and 1000μM) with or without folic acid (0, 10, 100, and 1000nM) for 24h. It was found that methomyl increased cytotoxicity to testicular cells in a dose-dependent manner. In spermatogonia, methomyl significantly inhibited the expression of proliferation genes Ki67 and PCNA at 1000μM, and increased the expression of apoptosis genes Caspase3 and Bax at each dose. In Sertoli cells, methomyl dose-dependently inhibited the expression of blood-testis barrier function genes TJP1, Cx43, and N-cadherin, but did not affect Occludin and E-cadherin. In Leydig cells, methomyl inhibited the expression of steroid synthase P450scc, StAR, Hsd3b1 and down-regulated the level of testosterone, but did not affect Cyp17a1 and Hsd17b1. Further, folic acid could basically reduce the damage caused by methomyl. This study provided new insights into the toxicity of methomyl and the protective effect of folic acid.

Yin Yang 1-Induced Long Noncoding RNA DUXAP9 Drives the Progression of Oral Squamous Cell Carcinoma by Blocking CDK1-Mediated EZH2 Degradation.

LncRNAs play a critical role in oral squamous cell carcinoma (OSCC) progression. However, the function and detailed molecular mechanism of most lncRNAs in OSCC are not fully understood. Here, a novel nuclear-localized lncRNA, DUXAP9 (DUXAP9), that is highly expressed in OSCC is identified. A high level of DUXAP9 is positively associated with lymph node metastasis, poor pathological differentiation, advanced clinical stage, worse overall survival, and worse disease-specific survival in OSCC patients. Overexpression of DUXAP9 significantly promotes OSCC cell proliferation, migration, invasion, and xenograft tumor growth and metastasis, and upregulates N-cadherin, Vimentin, Ki67, PCNA, and EZH2 expression and downregulates E-cadherin in vitro and in vivo, whereas knockdown of DUXAP9 remarkably suppresses OSCC cell proliferation, migration, invasion, and xenograft tumor growth in vitro and in vivo in an EZH2-dependent manner. Yin Yang 1 (YY1) is found to activate the transcriptional expression of DUXAP9 in OSCC. Furthermore, DUXAP9 physically interacts with EZH2 and inhibits EZH2 degradation via the suppression of EZH2 phosphorylation, thereby blocking EZH2 translocation from the nucleus to the cytoplasm. Thus, DUXAP9 can serve as a promising target for OSCC therapy.

Chk1/2 inhibitor AZD7762 blocks the growth of preantral follicles by inducing apoptosis, suppressing proliferation, and interfering with the cell cycle in granulosa cells.

Checkpoint kinases 1/2 (Chk1/2) have an important role in somatic cell development and oocyte meiotic maturation. However, the role of Chk1/2 in folliculo-genesis has not been fully elucidated. The aim of this study was to assess the effects of Chk1/2 inhibition on ovarian folliculogenesis and granulosa cell development in mice. Preantral follicles (100-120 μm) and granulosa cells from pre-ovulatory follicles (pre-GCs) of mice were isolated and cultured with or without Chk1/2 inhibitor AZD7762. Preantral follicles were cultured for 96h. Then, follicle morphology and fol-licular growth were assessed every 48h. Granulosa cells were cultured for 48h with or without AZD7762, after which cell apoptosis, cell proliferation, and cell cycle analysis were assessed; meanwhile, the mRNA expression of PCNA and Bax were measured by real-time RT-PCR, and PCNA and Bax protein were measured by Western blot. Compared with control follicles, AZD7762 inhibited growth of preantral folli-cles (P<0.05). Furthermore, inhibition of Chk1/2 significantly induced apoptosis (P<0.05) and inhibited the proliferation of granulosa cells (P<0.01), arrested cell cycle at S and G2/M phases, and decreased G1 phase fraction (P<0.001). Also, the expres-sion of PCNA mRNA and protein were reduced (P<0.01), while Bax mRNA and pro-tein were increased (P<0.05) post AZD7762 treatment in granulosa cells. This study revealed that Chk1 and Chk2 have a crucial role during preantral follicular development by regulating the proliferation and apoptosis of granu-losa cells.

Urotensin II promotes the proliferation and secretion of vascular endothelial growth factor in rat dermal papilla cells by activating the Wnt-β-catenin signaling pathway

Introduction. Urotensin II (U II) is a kind of active peptide with a variety of biological effects, such as promoting cell proliferation and endocrine effects. The aim of this study is to investigate the effect of urotensin II on the proliferation and secretion of vascular endothelial growth factor (VEGF) in cultured rat dermal papilla cells (DPCs), and to explore its molecular mechanism. Materials and Methods. We used the DPCs isolated from the thoracic aortas of Wistar-Kyoto rats to run the CCK8 and ELISA assay, RC-PCR and Western blotting techniques to identify the effect of Urotensin II on the proliferation and secretion of VEGF in DPCs, data were analyzed by one-way ANOVA or t-test. Results. U II can increase the mRNA expression of proliferation markers Ki67 and PCNA. In addition, the Wnt/β-catenin pathway was activated by U II, but Wnt inhibitor DKK1 reversed the effect of U II. Conclusions. U II promoted the proliferation and secretion of VEGF in rat DPCs through activation of the Wnt-β-catenin signaling pathway.