Metformin reduces myogenic contracture and myofibrosis induced by rat knee joint immobilization via AMPK-mediated inhibition of TGF-ß1/Smad signaling pathway.

PURPOSE: The two structural components contributing to joint contracture formation are myogenic and arthrogenic contracture, and myofibrosis is an important part of myogenic contracture. Myofibrosis is a response to long-time immobilization and is described as a condition with excessive deposition of endomysial and perimysial connective tissue components in skeletal muscle. The purpose of this study was to confirm whether metformin can attenuate the formation of myogenic contracture and myofibrosis through the phosphorylation level of adenosine monophosphate-activated protein kinase (AMPK) and inhabitation of subsequent transforming growth factor beta (TGF-ß) 1/Smad signaling pathway. MATERIALS AND METHODS: An immobilized rat model was used to determine whether metformin could inhibit myogenic contracture and myofibrosis. The contents of myogenic contracture of knee joint was calculated by measuring instrument of range of motion (ROM), and myofibrosis of rectus femoris were determined by ultrasound shear wave elastography and Masson staining. Protein expression of AMPK and subsequent TGF-ß1/Smad signaling pathway were determined by western blot. Subsequently, Compound C, a specific AMPK inhibitor, was used to further clarify the role of the AMPK-mediated inhibition of TGF-ß1/Smad signaling pathway. RESULTS: We revealed that the levels of myogenic contracture and myofibrosis were gradually increased during immobilization, and overexpression of TGF-ß1-induced formation of myofibrosis by activating Smad2/3 phosphorylation. Activation of AMPK by metformin suppressed overexpression of TGF-ß1 and TGF-ß1-induced Smad2/3 phosphorylation, further reducing myogenic contracture and myofibrosis during immobilization. In contrast, inhibition of AMPK by Compound C partially counteracted the inhibitory effect of TGF-ß1/Smad signaling pathway by metformin. CONCLUSION: Notably, we first illustrated the therapeutic effect of metformin through AMPK-mediated inhibition of TGF-ß1/Smad signaling pathway in myofibrosis, which may provide a new therapeutic strategy for myogenic contracture.

LncRNA KCNQ1OT1 regulates the invasion and migration of hepatocellular carcinoma by acting on S1PR1 through miR-149.

The aim of this study was to investigate the effect of lncRNA KCNQ1OT1 on HCC and to explore the possible underlying mechanisms. The expression levels of KCNQ1OT1, miR-149 and S1PR1 were detected by qRT-PCR assay. A dual luciferase reporter assay was used to detect the interaction between KCNQ1OT1 and miR-149, as well as miR-149 and S1PR1. The interaction between KCNQ1OT1 and miR-149 was further investigated by RNA pull-down assay. Wound healing assays and Transwell assays were carried out to determine cell migration and invasion. A xenograft tumour assay was used to validate the role of KCNQ1OT1 in vivo. KCNQ1OT1 and S1PR1 were significantly increased, but miR-149 was decreased in HCC cells. Luciferase reporter assays and RNA pull-down assays revealed that KCNQ1OT1 directly targeted miR-149. In addition, miR-149 bound to the 3'-UTR of S1PR1. Knockdown of KCNQ1OT1 or overexpression of miR-149 inhibited the invasion and migration of HCC cells. However, suppression of miR-149 could abrogate the effect of KCNQ1OT1 knockdown on the invasion and migration abilities of HCC cells. In vivo assays showed that KCNQ1OT1 knockdown suppressed tumour growth. This work suggests that lncRNA KCNQ1OT1 might act as a potential therapeutic target in HCC.

miR-34a regulates HDAC1 expression to affect the proliferation and apoptosis of hepatocellular carcinoma.

miR-34a is an important molecule that can inhibit the tumor growth. This study aimed to investigate the functional role of miR-34a in hepatocellular carcinoma (HCC) and explore the interaction between miR-34a and histone deacetylase 1 (HDAC1). RT-qPCR was employed to detect the mRNA expression of miR-34a and HDAC1 in 60 HCC tissues. Results showed miR-34a expression in HCC tissues was significantly lower than in normal tissues (P<0.05), but HDAC1 expression in HCC tissues was markedly higher than in normal tissues (P<0.05). In addition, miR-34a expression was negatively related to HDAC1 expression. miR-34a mimic was transfected into HCC cell lines (HepB3 and HepG2). CCK8 assay, colony formation assay and flow cytometry showed miR-34a over-expression could inhibit the proliferation of HCC cells and induce their apoptosis. Western blotting indicated miR-34a over-expression down-regulated the expression of Bcl-2, procaspase-3, procaspase-9 and c-Myc, but up-regulate p21 expression. Bioinformatics analysis indicated HDAC1 was a target gene of miR-34a. Dual Luciferase Reporter Gene Assay and retrieval assay showed miR-34a could act at the 3'UTR of HDAC1 gene to regulate its expression. Thus, miR-34a may inhibit the proliferation of HCC cells and induce their apoptosis via regulating HDAC1 expression. Our findings provide evidence for the diagnosis and therapeutic target of HCC.

[Antagonistic effects of vitamin E on the testicular injury by cyclophosphamide in mice].

OBJECTIVE: To observe the protective effects of vitamin E on the testicular injury by cyclophosphamide in mice, and the correlative mechanism. METHODS: Fifty sexually mature male mice were randomly divided into five groups: the cyclophosphamide group (the CP group), the low-dose vitamin E group (the low-dose group), the middle-dose vitamin E group (the middle-dose group), the high-dose vitamin E group (the high-dose group), the matched control group (the control group). The first four groups were given cyclophosphamide by gavage at a dose of 5 mg/(kg x d). The low-dose group, the middle-dose group and the high-dose group were given vitamin E by subcutaneous injection at doses of 30 mg/(kg x d), 50 mg/(kg x d) , 70 mg/(kg x d) after 4 h of cyclophosphamide treatment. The control group was gavaged with equivalent normal saline. The treatment period for all groups was 28 days. The level of plasma FSH, LH, T and the activity of testicular SOD, GSHPx, CAT and the level of testicular MDA were detected. The histological structure and the ultrastructure of the testis were examined by light microscope and electron microscope. RESULTS: As compared with the CP group, the plasma FSH, LH, T level and the SOD, GSHPx, CAT activity in the middle-dose group and the high-dose group were higher (P< 0.05, P< 0.01), MDA level significantly lower(P<0.01). The histological structure and the ultrastructure of the testis were in the normal range. CONCLUSION: Vitamin E has protective effects on the testicular injury by cyclophosphamide in mice. The possible mechanism of vitamin E may be its scavenging free radical and antioxidant effects, as well as it may have some stimulatory effects on gonadotrophin releasing of pituitary anterior lobe.