Shaanxi University of Traditional Chinese Medicine

This study aimed to investigate the possible mechanism through which acupuncture protects ovaries with Poor Ovarian Response (POR) in rats based on microRNA (miRNA).

Thirty-six SPF SD female non-pregnant rats aged 8 weeks were randomly divided into the blank group, model group, and acupuncture group, with 12 rats in each group. According to the group, the rats were given gavage of Tripterygium wilfordii polyglycosides suspension for 14 days to establish the model of POR, and then the rats were treated with acupuncture for 2 weeks, once a day, for 20 minutes.

Afterward, their hormone levels were measured, and HE staining was performed on the ovaries after the intervention. Three rats from each group were randomly selected for ovarian tissue miRNA sequencing, and differential miRNAs were subjected to Gene Ontology (GO) enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway analysis, and Quantitative Polymerase Chain Reaction (QPCR) verification. By using TargetScan to predict the target genes of differential miRNAs, we validated the results with a dual luciferase reporter gene assay.

Compared with the blank group, the estrus cycle of the model group was significantly prolonged (P<0.01). Anti-Müllerian Hormone (AMH) (P<0.01) and Estrogen (E2) were significantly decreased (P<0.05). Follicle-Stimulating Hormone (FSH) (P<0.05) and Luteinizing Hormone (LH) increased sharply (P<0.01). Compared with the model group, the estrus cycle was significantly shortened in the acupuncture group (P<0.01). AMH and E2 were markedly raised (P<0.05). FSH (P<0.05) and LH (P<0.01) were significantly declined. miRNA sequencing showed that there were 23 miRNAs significantly different between the model group and the blank group (P<0.05), and 30 miRNAs significantly different between the acupuncture group and the model group (P<0.05). GO demonstrated that the network was mainly involved in cellular components, cells, cellular metabolic processes, binding, and single biological processes; KEGG signaling pathway enrichment showed that it was mainly related to MAPK, adhesion junction, calcium signaling pathways, etc. Q-PCR results showed that after modeling, the expression rose, and after acupuncture, the expression of the following miRNA decreased: miR-154-5p (P<0.01), miR-300-5p (P < 0.05), miR-376c-5p (P<0.05). The dual luciferase reporter assay showed that the relative luciferase activity of the miR-300-5p mimics+MAP3K1-WT group was significantly lower than that of the NC+MAP3K1-WT group (P<0.01). HE results demonstrated that the number of primordial follicles and primary follicles in the model group was significantly lower than that in the blank group (P<0.05). Moreover, the morphology was poorer, and the granulosa cell layer was thinner. Compared with the model group, the number of primary follicles in the acupuncture group increased (P<0.05); the morphology and granulosa cell structure of the ovary were improved to different degrees, and mature follicles could be seen.

Acupuncture may improve the ovarian responsiveness of POR rats by regulating miR-154- 5p,miR-300-5p, and miR-376c-5p. Furthermore, miR-300-5p can specifically bind to the 3'-UTR of MAP3K1, and MAP3K1 may be the target of miR-300-5p.

To observe whether electroacupuncture stimulation of auricular concha region (EA-ACR) on behavior changes of depression by loss of control over stress model (LOC), and whether its effect is improved by regulating the expression levels of hydroxytryptamine (serotonin, 5-HT) 1A receptor (5-HT_1AR)/ hydroxytryptamine (serotonin, 5-HT) 2A receptor (5-HT_2AR) in hippocampus.

LOC was prepared using a Skinner box, and EA-ACR to observe behavioral changes, and Western Blot was used to detect the changes of 5-HT_1AR/5-HT_2AR in the hippocampus, and then observe the changes of EA-ACR behavior after microinjection of 5-HT_1AR/5-HT_2AR antagonist into the hippocampus.

EA-ACR improve depressive-like behavior, up-regulated 5-HT_1AR expression and down-regulated 5-HT_2AR expression in hippocampal brain area. EA-ACR did not improve depression-like behavior after hippocampal microinjection of 5-HT_1AR antagonist, while injection of 5-HT_2AR antagonists can improve depression-like behaviors.

EA-ACR can improve depressive-like behaviors. Loss of control over stress leads to up-regulation of 5-HT_1AR and down-regulation of 5-HT_2AR in the hippocampus, while EA-ACR mainly improves depressive behavior by regulating 5-HT_1AR in Hip.

To investigate the effect of 1α,25(OH)2D3 on hepatic stellate cells and the mechanism of the TGF-β1/Smad signaling pathway.LX2 cells were treated with TGF-β1 and different concentrations of 1α,25(OH)2D3. Cell proliferation was assessed using the CCK8 assay to determine the optimal concentration of 1α,25(OH)2D3 activity. The cell cycle and apoptotic rates were evaluated using flow cytometry. The expressions of Samd2, Samd3, Samd4, and Samd7 was assessed by western blotting, whereas the expression of MMP1, MMP13, and TIMP-1 was detected by qPCR.Compared with the control group, the 1α,25(OH)2D3 group had a higher apoptotic rate of LX2 cells, the cell cycle was blocked from the G1 stage to the S stage, the expressions of Samd2, Samd7, MMP1, and MMP13 increased, while the expressions of Samd3, Samd4, and TIMP-1 decreased.1α,25(OH)2D3 inhibits hepatic stellate cell activation and exerts anti-hepatic fibrosis effects by downregulating the expression of Samd3, Samd4, TIMP-1 and upregulating the expression of Samd2, Samd4, MMP1, and MMP13.