摘要: Objectives The transformation of cytotrophoblasts into mesenchymal-like extravillous trophoblasts is necessary for successful embryo implantation, and the inadequate transformation may cause abortion. Epiregulin, which is a new growth factor, plays important roles in the reproductive processes. The glycosylation of many proteins in reproduction processes is critical. Protein O-fucosyltransferase 1 (poFUT1) is the key enzyme for the biosynthesis of O-fucosylation on the specific glycoproteins. Urokinase-type plasminogen activator (uPA) contains O-fucosylated domain on Thr(18). However, the functions of epiregulin and poFUT1 in the trophoblast epithelial-mesenchymal transition (EMT) process, the regulatory mechanism of epiregulin on poFUT1 and the resulting O-fucosylated uPA remain unclear. Materials and methods We employed ELISA and Western blot to detect serum levels of epiregulin and poFUT1 from non-pregnancy women, pregnancy women and abortion patients. Using two trophoblast cell lines and a mouse pregnancy model, we investigated the underlying mechanisms of epiregulin and poFUT1 in trophoblast EMT process. Results Serum levels of epiregulin and poFUT1 were higher in pregnant women compared with non-pregnant women, and their levels were significantly decreased in abortion patients compared with pregnant women. The results showed that epiregulin upregulated poFUT1 expression and increased O-fucosylation on uPA, which further activated the PI3K/Akt signalling pathway, facilitating EMT behaviour of trophoblast cells and embryo implantation in the mouse pregnant model. Conclusions Level of epiregulin and poFUT1 is lower in abortion patients than early pregnancy women. Epiregulin promotes trophoblast EMT through O-fucosylation on uPA catalysed by poFUT1. Epiregulin and poFUT1 may be suggested as the potential diagnostic biomarkers and useful treatment targets for abortion.

摘要: Objectives Chemoresistance induced by cisplatin has become the major impediment to lung cancer chemotherapy. This study explored the potential chemoresistant genes and underlying mechanisms of chemoresistance in NSCLC. Materials and methods Gene expression profile was integrated with DNA methylation profile to screen the candidate chemoresistant genes. Bioinformatic analysis and immunohistochemistry were used to analyse the association of a candidate gene with the characteristics of NSCLC patients. Recombinant lentivirus vectors were utilized to overexpress or silence candidate gene. Microarrays and immunoblotting were applied to explore the downstream targets of candidate gene. Xenograft models were established to validate the findings in vitro. Results An increased ZNF300 expression was detected in three chemoresistant cell lines of NSCLC, and the higher expression of ZNF300 was associated with poor OS of NSCLC patients. Cells with upregulated ZNF300 presented chemoresistance and enhanced aggressive growth compared to cells with downregulated ZNF300. ZNF300 inhibited MAPK/ERK pathways and activated CDK1 through inhibiting WEE1 and MYT1 and modulating MYC/AURKA/BORA/PLK1 axis. ICA and ATRA improved the anti-tumour effect of cisplatin on chemoresistant cells by inducing differentiation. Conclusions ZNF300 promotes chemoresistance and aggressive behaviour of NSCLC through regulation of proliferation and differentiation by downregulating MAPK/ERK pathways and regulation of slow-cycling phenotype via activating CDK1 by inhibiting WEE1/MYT1 and modulating MYC/AURKA/BORA/PLK1 axis. Cisplatin, combined with ATRA and ICA, might be beneficial in chemoresistant cases of NSCLC.

摘要: Objectives The level of histone H3 lysine 79 methylation is regulated by the cell cycle and involved in cell proliferation. KDM2B is an H3K79 demethylase. Proliferating cell nuclear antigen (PCNA) is a component of the DNA replication machinery. This study aimed at elucidating a molecular link between H3K79me recognition of PCNA and cell cycle control. Materials and methods We generated KDM2B-depleted 293T cells and histone H3-K79R mutant-expressing 293T cells. Western blots were primarily utilized to examine the H3K79me level and its effect on subsequent PCNA dissociation from chromatin. We applied IP, peptide pull-down, isothermal titration calorimetry (ITC) and ChIP experiments to show the PCNA binding towards methylated H3K79 and DNA replication origins. Flow cytometry, MTT, iPOND and DNA fibre assays were used to assess the necessity of KDM2B for DNA replication and cell proliferation. Results We revealed that KDM2B-mediated H3K79 demethylation regulated cell cycle progression. We found that PCNA bound chromatin in an H3K79me-dependent manner during S phase. KDM2B was responsible for the timely dissociation of PCNA from chromatin, allowing to efficient DNA replication. Depletion of KDM2B aberrantly enriched chromatin with PCNA and caused slow dissociation of residual PCNA, leading to a negative effect on cell proliferation. Conclusions We suggested a novel interaction between PCNA and H3K79me. Thus, our findings provide a new mechanism of KDM2B in regulation of DNA replication and cell proliferation.

摘要: Objectives Postflight orthostatic intolerance has been regarded as a major adverse effect after microgravity exposure, in which cerebrovascular adaptation plays a critical role. Our previous finding suggested that dedifferentiation of vascular smooth muscle cells (VSMCs) might be one of the key contributors to cerebrovascular adaptation under simulated microgravity. This study was aimed to confirm this concept and elucidate the underlying mechanisms. Materials and Methods Sprague Dawley rats were subjected to 28-day hindlimb-unloading to simulate microgravity exposure. VSMC dedifferentiation was evaluated by ultrastructural analysis and contractile/synthetic maker detection. The role of T-type Ca(V)3.1 channel was revealed by assessing its blocking effects. MiR-137 was identified as the upstream of Ca(V)3.1 channel by luciferase assay and investigated by gain/loss-of-function approaches. Calcineurin/nuclear factor of activated T lymphocytes (NFAT) pathway, the downstream of Ca(V)3.1 channel, was investigated by detecting calcineurin activity and NFAT nuclear translocation. Results Simulated microgravity induced the dedifferentiation and proliferation in rat cerebral VSMCs. T-type Ca(V)3.1 channel promoted the dedifferentiation and proliferation of VSMC. MiR-137 and calcineurin/NFATc3 pathway were the upstream and downstream signalling of T-type Ca(V)3.1 channel in modulating the dedifferentiation and proliferation of VSMCs, respectively. Conclusions The present work demonstrated that miR-137 and its target T-type Ca(V)3.1 channel modulate the dedifferentiation and proliferation of rat cerebral VSMCs under simulated microgravity by regulating calcineurin/NFATc3 pathway.

摘要: Objectives Mechanical force plays an important role in modulating stem cell fate and behaviours. However, how periodontal ligament stem cells (PDLSCs) perceive mechanical stimulus and transfer it into biological signals, and thereby promote alveolar bone remodelling, is unclear. Materials and Methods An animal model of force-induced tooth movement and a compressive force in vitro was used. After force application, tooth movement distance, mesenchymal stem cell and osteoclast number, and proinflammatory cytokine expression were detected in periodontal tissues. Then, rat primary PDLSCs with or without force loading were isolated, and their stem cell characteristics including clonogenicity, proliferation, multipotent differentiation and immunoregulatory properties were evaluated. Under compressive force in vitro, the effects of the ERK signalling pathway on PDLSC characteristics were evaluated by Western blotting. Results Mechanical force in vivo induced PDLSC proliferation, which was accompanied with inflammatory cytokine accumulation, osteoclast differentiation and TRPV4 activation; the force-stimulated PDLSCs showed greater clonogenicity and proliferation, reduced differentiation ability, improved induction of macrophage migration, osteoclast differentiation and proinflammatory factor expression. The biological changes induced by mechanical force could be partially suppressed by TRPV4 inhibition. Mechanistically, force-induced activation of TRPV4 in PDLSCs regulated osteoclast differentiation by affecting the RANKL/OPG system via ERK signalling. Conclusions Taken together, we show here that TRPV4 activation in PDLSCs under mechanical force contributes to changing their stem cell characteristics and modulates bone remodelling during tooth movement.