摘要: Busulfan is an antineoplastic, which is always accompanied with the abnormal of spermatogonia self-renewal and differentiation. It has been demonstrated that the omega-3 polyunsaturated fatty acids (PUFAs) benefits mature spermatozoa. However, whether omega-3 can protect endogenous spermatogonia and the detailed mechanisms are still unclear. Evaluate of spermatogenesis function (in vivo) were examined by histopathological analysis, immunofluorescence staining, and western blotting. The levels of lipid metabolites in testicular tissue were determined via liquid chromatography. We investigated the effect of lipid metabolites on Sertoli cells provided paracrine factors to regulate spermatogonia proliferation and differentiation using co-culture system. In our study, we showed that omega-3 PUFAs significantly improved the process of sperm production and elevated the quantity of both undifferentiated Lin28+ spermatogonia and differentiated c-kit+ spermatogonia in a mouse model where spermatogenic function was disrupted by busulfan. Mass spectrometry revealed an increase in the levels of several omega-3 metabolites in the testes of mice fed with omega-3 PUFAs. The eicosapentaenoic acid metabolite 12-hydroxyeicosapentaenoic acid (12-HEPE) up-regulated bone morphogenic protein 4 (BMP4) expression through GPR120-ERK1/2 pathway activation in Sertoli cells and restored spermatogonia proliferation and differentiation. Our study provides evidence that omega-3 PUFAs metabolite 12-HEPE effectively protects spermatogonia and reveals that GPR120 might be a tractable pharmacological target for fertility in men received chemotherapy or severe spermatogenesis dysfunction.

摘要: Scaffold protein AF4/FMR2 family member 4 (AFF4) has been found to play a role in osteogenic commitment of stem cells. However, function of AFF4 in human periodontal ligament stem cells (hPDLSCs) has not been studied yet. This present study aims to investigate the biological effect of AFF4 on osteogenic differentiation of hPDLSCs and potential mechanistic pathway. First, AFF4 expression profile was evaluated in conditions of periodontitis and osteogenic differentiation of hPDLSCs by immunohistochemical staining, western blot and qRT-PCR. Next, si-RNA mediated knockdown and lentiviral transduction mediated overexpression of AFF4 were adopted to explore impact of AFF4 on osteogenic capacity of hPDLSCs. Then, possible mechanistic pathway was identified. At last, pharmacological agonist of autophagy, rapamycin, was utilized to affirm the role of autophagy in AFF4-regulated osteogenesis of hPDLSCs. First, AFF4 expressions were significantly lower in inflamed periodontal tissues and lipopolysaccharides-treated hPDLSCs than controls, and were up-regulated during osteogenic differentiation of hPDLSCs. Next, osteogenic potential of hPDLSCs was impaired by AFF4 knockdown and potentiated by AFF4 overexpression. Moreover, AFF4 was found to positively regulate autophagic activity in hPDLSCs. At last, rapamycin treatment was shown to be able to partly restore AFF4 knockdown-suppressed osteogenic differentiation. Our study demonstrates that AFF4 regulates osteogenic potential of hPDLSCs via targeting autophagic activity. The involvement of AFF4 in periodontal homeostasis was identified for the first time. The present study shows that AFF4 protein expression was significantly lower in periodontitis tissues than healthy periodontal tissues. And we further revealed that AFF4 regulates osteogenic differentiation of hPDLSCs. The above findings provide a clue for possible engagement of AFF4 in local dyshomeostasis in periodontitis.image

摘要: The liver is the most tolerogenic of transplanted organs. However, the mechanisms underlying liver transplant tolerance are not well understood. The comparison between liver transplantation tolerance and heart/kidney transplantation rejection will deepen our understanding of tolerance and rejection in solid organs. Here, we built a mouse model of liver, heart and kidney allograft and performed single-cell RNA sequencing of 66,393 cells to describe the cell composition and immune cell interactions at the early stage of tolerance or rejection. We also performed bulk RNA-seq of mouse liver allografts from Day 7 to Day 60 post-transplantation to map the dynamic transcriptional variation in spontaneous tolerance. The transcriptome of lymphocytes and myeloid cells were characterized and compared in three types of organ allografts. Cell-cell interaction networks reveal the coordinated function of Kupffer cells, macrophages and their associated metabolic processes, including insulin receptor signalling and oxidative phosphorylation in tolerance induction. Cd11b+ dendritic cells (DCs) in liver allografts were found to inhibit cytotoxic T cells by secreting anti-inflammatory cytokines such as Il10. In summary, we profiled single-cell transcriptome analysis of mouse solid organ allografts. We characterized the immune microenvironment of mouse organ allografts in the acute rejection state (heart, kidney) and tolerance state (liver). Here, we established mouse models of liver, heart and kidney transplantation. We performed single-cell analysis of the allografts at the early phase of rejection/tolerance as well as bulk RNA analysis of long-term tolerance in liver allograft at different time point. We compared gene expression patterns of the immune microenvironment in organ allografts. We identified coordinated function of macrophages and associated metabolic process including insulin receptor signaling and oxidative phosphorylation process in liver transplantation tolerance induction.image

摘要: Orofacial muscle defect due to congenital anomalies, tumour ablation or traumatic accident that exceeds endogenous regeneration capacity may lead to sustained deficits in masticatory function and nutrition intake. Functional recovery has always been the goal of muscle tissue repair, but currently, there is no suitable model for quantitative analyses of either functional consequences or treatment efficacy of orofacial muscle defect. This study proposed a critical size volumetric muscle loss (VML) model in mouse masseter with impaired mastication on nutrition. Full-thickness VML defects in diameter of 1.0, 1.5, 2.0 and 3.0 mm were generated in the centre of the mouse masseter using a biopsy punch to determine the critical size for functional impairment. In the VML region, myogenesis was dampened but fibrogenesis was activated, as long with a reduction in the density of the neuromuscular junction and an increase in vascular density. Accordingly, persistent fibrosis was observed in the centre region of VML in all diameters. The 2.0 mm diameter was the critical threshold to masticatory function impairment after VML in the masseter. VML of 3.0 mm diameter led to a significant impact on nutrition intake and body weight gain. Autologous muscle graft effectively relieved the fibrosis and functional deficit after VML injury in the masseter. This model serves as a reliable tool in studying functional recovery strategies for orofacial muscle defects. In this study, we presented for the first time an injury model in the orofacial masticatory muscle with function impairment and systemic impact, along with comprehensive quantitative histological analysis, muscle repair effector cell proliferative activity and masticatory function analysis, providing a powerful tool for testing and developing tissue engineering and regenerative medicine approaches for orofacial muscle repair. image

摘要: Glaucoma and other optic neuropathies lead to progressive and irreversible vision loss by damaging retinal ganglion cells (RGCs) and their axons. Cell replacement therapy is a potential promising treatment. However, current methods to obtain RGCs have inherent limitations, including time-consuming procedures, inefficient yields and complex protocols, which hinder their practical application. Here, we have developed a straightforward, rapid and efficient approach for directly inducing RGCs from mouse embryonic fibroblasts (MEFs) using a combination of triple transcription factors (TFs): ASCL1, BRN3B and PAX6 (ABP). We showed that on the 6th day following ABP induction, neurons with molecular characteristics of RGCs were observed, and more than 60% of induced neurons became iRGCs (induced retinal ganglion cells) in the end. Transplanted iRGCs had the ability to survive and appropriately integrate into the RGC layer of mouse retinal explants and N-methyl-D-aspartic acid (NMDA)-damaged retinas. Moreover, they exhibited electrophysiological properties typical of RGCs, and were able to regrow dendrites and axons and form synaptic connections with host retinal cells. Together, we have established a rapid and efficient approach to acquire functional RGCs for potential cell replacement therapy to treat glaucoma and other optic neuropathies. The transcription factor (TF) combination ASCL1 + BRN3B + PAX6 could efficiently reprogram mouse embryonic fibroblasts (MEFs) into induced retinal ganglion cells (iRGCs). These iRGCs not only expressed several molecular markers of native RGCs, but also exhibited typical RGC electrophysiological properties. Upon transplantation into mouse retinal explants or NMDA-damaged retinas, iRGCs could integrate into the ganglion cell layer, survive for a prolonged period, grow long neuron fibres, form synapses with host cells, and exhibit electrophysiological functions.image