Gga-miR-146b-3p promotes apoptosis and attenuate autophagy by targeting AKT1 in chicken granulosa cells.

The normal development of follicles determines the reproductive performance of females. Granulosa cells (GC) play crucial roles in follicular maturation. Numerous studies have shown that miRNAs are involved in the regulation of GC. According to our previous sequencing data, gga-miR-146b-3p was differentially expressed in normal and atretic chicken follicles. In this study, we verified that gga-miR-146b-3p attenuated proliferation and autophagy but promoted apoptosis in chicken GC. Threonine kinase1 (AKT1), a key member of the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway, was predicted to be a target gene of gga-miR-146b-3p via bioinformatic analysis. Dual-luciferase reporter gene assays were used to determine target relationships. Moreover, knockout of AKT1 decelerated proliferation and autophagy while accelerating the apoptosis of GC. However, overexpression of AKT1 reversed these results. In summary, our results demonstrated that gga-miR-146b-3p repressed the proliferation and autophagy of chicken GC while up-regulating apoptosis by targeting AKT1 through the PI3K/AKT signaling pathway. These findings may provide great insights for further exploration of the molecular regulation of gga-miR-146b-3p and AKT1 on the functions of GC during folliculogenesis.

[Research on autophagy induced by two xanthone compounds in HepG2 cells].

To investigate the inhibitory effects of two xanthone compounds, 1-hydroxy-2,3,4,8-4 methoxy xanthone(here in after referred to as Fr15) and 1-hydroxy-2,3,4,6-4 methoxy xanthone(here in after referred to as Fr17), on the proliferation of hepatocellular carcinoma cells HepG2, and to further investigate their mechanism in combination with transcriptomics. Cell counting was used to detect the effects of two kinds of xanthone compounds Fr15 and Fr17(0, 0.03, 0.15, 0.3 mmoL·L~(-1)) on the proliferation of HepG2 cells; the effects of the two compounds Fr15 and Fr17 on HepG2 cell cycle were detected by flow cytometry; the changes of autophagosomes count in cells were observed under fluorescence microscope; the expression of autophagy marker proteins autophagy marker proteins SQSTM 1(p62) and microtubule associated protein 1 light chain 3 Ⅰ/Ⅱ(LC3 Ⅰ/Ⅱ) in the cells was detected by Western blot; the differentially expressed genes between the control group and the experimental group were analyzed by RNA-seq transcriptome sequencing; qRT-PCR was used to verify the differentially expressed genes in sequencing. The results showed that compounds Fr15 and Fr17 inhibited the proliferation of HepG2 cells with the increase of drug concentration and time. Flow cytometry showed that compounds Fr15 and Fr17 had little effect on HepG2 cell cycle. Fluorescence microscopy results showed that the number of autophagosomes in cells increased with the increase of drug concentration. Western blot showed that the expression of p62 protein was decreased and the expression of LC3-Ⅱ protein was significantly increased after drug addition. The results of RNA sequencing showed that 26 102 and 52 351 differentially expressed genes were obtained in Fr15 and Fr17 respectively. Analysis of KEGG showed that drug treatment had a great effect on autophagy pathway. qRT-PCR verified that 6 up-regulated genes were related to autophagy, and their trend was consis-tent with sequencing results, where all 6 genes showed an up-regulated trend. Two xanthone compounds Fr15 and Fr17 may inhibit proliferation of HepG2 cells by inducing autophagy.

Immunomodulatory and neuroprotective mechanisms of Huangqi glycoprotein treatment in experimental autoimmune encephalomyelitis.

Previous studies have shown that Huangqi glycoprotein (HQGP) has an anti-inflammatory effect in vitro, and suppressed experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis; however, the mechanism underlying its effect is largely unknown. In this manuscript we investigated the mechanisms by which HQGP protect mice from EAE. HQGP was extracted from Astragalus membranaceus and purified by anion-exchange and gel filtration chromatography. HQGP delayed disease onset, reduced disease severity and alleviated inflammation and demyelination in the central nervous system (CNS). Moreover, HQGP reduced the infiltration of pathogenic immune cells and increased the expression of microtubule-associated protein 2 (MAP-2) and neuronal nuclei (NeuN) in the CNS. HQGP treatment also reduced the expression of chemokines such as CCL2 and CCL5 and the production of tumor necrosis factor α (TNF-α), interleukin (IL)-1ß, IL-6, but increased the level of IL-10. These results demonstrate that HQGP suppressed EAE development by modulating the immune system and the infiltration of leukocytes to the CNS as well as promoting axon and neural repair.

Triptolide, A Potential Autophagy Modulator.

As a major active component extracted from traditional Chinese herb Tripterygium wilfordii Hook F, triptolide exhibits multiple pharmacological effects. Autophagy is an evolutionary conserved intracellular catabolic process involved in cytoplasmic materials degradation. Autophagic dysfunction contributes to the pathologies of many human diseases, which makes it a promising therapeutic target. Recent studies have shown that triptolide exerts neuroprotection, anti-tumor activities, organ toxicity, and podocyte protection by modulating autophagy. This article highlights the current information on triptolide-modulated autophagy, analyzes the possible pathways involved, and describes the crosstalk between autophagy and apoptosis modulated by triptolide, in hope of providing implications for the roles of autophagy in pharmacological effects of triptolide and expanding its novel usage as an autophagy modulator.

[Immunomodulatory effect of Huangqi glycoprotein on mice with experimental autoimmune encephalomyelitis].

OBJECTIVE: To explore the therapeutic effect and possible mechanism of Huangqi glycoprotein (HQGP) on the development of experimental autoimmune encephalomyelitis (EAE). METHODS: Female C57BL/6 mice were immunized subcutaneously with myelin oligodendrocyte glycoprotein peptide-35-55 (MOG35-55) and divided into HQGP-treated group and EAE control group. Clinical score and body mass were recorded every other day. Inflammatory cell infiltrations of spinal cord were observed by HE and immunofluorescence staining. The cell viability of splenic mononuclear cells (MNCs) was detected by MTT assay. The release of NO was measured by Griess method. The levels of TNF-α, IL-6 and IFN-γ were determined by ELISA. The subtypes of CD4(+)T cells were analyzed by flow cytometry. RESULTS: HQGP treatment delayed the onset of EAE, attenuated the clinical symptoms and inhibited CD68(+) macrophage infiltration into the central nervous system. HQGP inhibited the viability of splenic MNCs, downregulated the secretion of NO, TNF-α, IL-6, and increased the secretion of IFN-γ. In addition, HQGP treatment effectively increased the numbers of CD4(+)CD25(+) T cells, CD4(+)IL-10(+) T cells and CD4(+)IFN-γ(+) T cells. CONCLUSION: HQGP relieve the inflammation of EAE via reducing the number of T cell subsets and inhibiting the secretion of inflammatory cytokines.