Paeoniflorin promotes PPARγ expression to suppress HSCs activation by inhibiting EZH2-mediated histone H3K27 trimethylation.

BACKGROUND: The alleviating effect of paeoniflorin (Pae) on liver fibrosis has been established; however, the molecular mechanism and specific target(s) underlying this effect remain elusive. PURPOSE: This study was to investigate the molecular mechanism underlying the regulatory effect of Pae on hepatic stellate cells (HSCs) activation in liver fibrosis, with a specific focus on the role of Pae in modulating histone methylation modifications. METHODS: The therapeutic effect of Pae was evaluated by establishing in vivo and in vitro models of carbon tetrachloride (CCl4)-induced mice and transforming growth factor ß1 (TGF-ß1)-induced LX-2 cells, respectively. Molecular docking, surface plasmon resonance (SPR), chromatin immunoprecipitation-quantitative real time PCR (ChIP-qPCR) and other molecular biological methods were used to clarify the molecular mechanism of Pae regulating HSCs activation. RESULTS: Our study found that Pae inhibited HSCs activation and histone trimethylation modification in liver of CCl4-induced mice and LX-2 cells. We demonstrated that the inhibitory effect of Pae on the activation of HSCs was dependent on peroxisome proliferator-activated receptor γ (PPARγ) expression and enhancer of zeste homolog 2 (EZH2). Mechanistically, Pae directly binded to EZH2 to effectively suppress its enzymatic activity. This attenuation leaded to the suppression of histone H3K27 trimethylation in the PPARγ promoter region, which induced upregulation of PPARγ expression. CONCLUSION: This investigative not only sheds new light on the precise targets that underlie the remission of hepatic fibrogenesis induced by Pae but also emphasizes the critical significance of EZH2-mediated H3K27 trimethylation in driving the pathogenesis of liver fibrosis.

An integrated network pharmacology, molecular docking and experiment validation study to investigate the potential mechanism of Isobavachalcone in the treatment of osteoarthritis.

BACKGROUND: In many different plants, including Dorstenia and Psoralea corylifolia L., Isobavachalcone (IBC) is a naturally occurring flavonoid chemical having a range of biological actions, including anti-inflammatory, immunomodulatory, and anti-bacterial. The "Theory of Medicinal Properties" of the Tang Dynasty states that Psoralea corylifolia L. has the ability to alleviate discomfort in the knees and waist. One of the most widespread chronic illnesses, osteoarthritis (OA), is characterized by stiffness and discomfort in the joints. However, there hasn't been much research done on the effectiveness and underlying processes of IBC in the treatment of osteoarthritis. AIM OF THE STUDY: To investigate the potential efficacy and mechanism of IBC in treating osteoarthritis, we adopted an integrated strategy of network pharmacology, molecular docking and experiment assessment. MATERIALS AND METHODS: The purpose of this research was to determine the impact of IBC on OA and the underlying mechanisms. IBC and OA possible targets and processes were predicted using network pharmacology, including the relationship between IBC and OA intersection targets, Cytoscape protein-protein interaction (PPI) to obtain key potential targets, and GO and KEGG pathway enrichment analysis to reveal the probable mechanism of IBC on OA. Following that, in vitro tests were carried out to confirm the expected underlying processes. Finally, in vivo tests clarified IBC's therapeutic efficacy on OA. RESULTS: We anticipated and validated that the impact of IBC on osteoarthritis is mostly controlled by the PI3K-AKT-NF-κB signaling pathway by combining the findings of network pharmacology analysis, molecular docking and Experiment Validation. CONCLUSIONS: This study reveals the IBC has potential to delay OA development.

Yunpi Heluo decoction attenuates insulin resistance by regulating SIRT1-FoxO1 autophagy pathway in skeletal muscle of Zucker diabetic fatty rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Diabetes is a serious chronic metabolic disorder, and type 2 diabetes mellitus (T2DM) accounts for more than 90% of all diabetes cases. Insulin resistance (IR) is an early symptom, typical feature and main pathogenesis of T2DM due to the combined effects of genetic and environmental factors. Current evidence shows that IR is mainly caused by nutrient overload, systemic fatty acid excess, fatty tissue inflammation, endoplasmic reticulum stress, oxidative stress and abnormal autophagy. Autophagy plays an important role in the development of IR and decreased autophagy activity can cause IR through various ways. AIM OF THE STUDY: Yunpiheluo (YPHL) decoction is a Chinese herbal formula with unique advantages for the treatment of T2DM. The aim of the present study was to investigate the regulatory mechanism of YPHL on the autophagy pathway in the skeletal muscle of IR Zucker diabetic fatty (ZDF) rats. METHODS: T2DM ZDF rats were treated with YPHL or transfected with SIRT1 adeno-associated virus. Serum total cholesterol (TC), triglyceride (TG), high-density lipoprotein (HDL), low-density lipoprotein (LDL), insulin resistance index (IRI) and skeletal muscle TG levels were detected in a T2DM ZDF rat model. The skeletal muscle morphology was observed by histological analysis and Oil Red O Staining. Autophagosomes were observed by transmission electron microscopy (TEM). The skeletal muscle morphology and fat deposition were observed by histological examination and Oil Red O Staining. A rat skeletal muscle IR cell model was established and transfected with SIRT1 overexpression plasmids. Cell apoptosis was observed by DAPI staining. SIRT1 levels in skeletal muscle tissues and cells were detected by qRT-PCR. The protein expressions of SIRT1, FOXo1, LC3B and P62 were detected by Western blotting. RESULTS: Large numbers of lipid droplets and swollen mitochondria were observed in the skeletal muscle in both model group and negative control (NC) group receiving blank plasmid. Autophagosomes were seen in the skeletal muscle of YPHL and SIRT1 groups, with no significant structural abnormality. In addition, the protein expression of LC3B was decreased and the protein expression of p62 was increased significantly in the model group as compared with the NC group. After intervention with YPHL and SIRT1 overexpression, the protein expression of LC3B was significantly increased and p62 was significantly decreased. However, there was no significant difference in cell apoptosis between the two groups. CONCLUSION: The SIRT1-FoxO1 autophagy pathway may play a significant role in the pathogenesis of IR. YPHL could increase the autophagy level by regulating the SIRT1-FoxO1 signaling pathway in the skeletal muscle and improving the lipid metabolism, thereby attenuating IR.

Coptisine from Rhizoma coptidis exerts an anti-cancer effect on hepatocellular carcinoma by up-regulating miR-122.

With increasing incidence and mortality, hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide. In this study, microRNA-122 (miR-122) mimics and relevant control oligonucleotides were transfected into HepG2 cells in vitro, followed by coptisine (COP) and sorafenib treatments. Cell proliferation, migration, and apoptosis were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and colony formation assay, wound-healing assay, Hoechst 33258 staining and flow cytometry, respectively. Histopathology and miR-122 were analyzed by haemotoxylin and eosin (H&E) staining and real-time RT-PCR, respectively; whereas, the relevant protein expressions were detected by western blot. In vivo, COP enhanced the expression of miR-122 by 160% compared to control in male BALB/c nude mice; COP not only protected the liver morphology but also showed a significant anti-cancer effect. Further, there was no remarkable difference between the tumor weights in the COP and sorafenib groups, but there was a striking difference to the tumor control group (p < 0.05). Hence, COP inhibited the proliferation, migration and promoted apoptosis of HCC cells; moreover, it inhibited the tumor growth in nude mice by up-regulating the expression of miR-122.

Protective effect of Coptisine from Rhizoma Coptidis on LPS/D-GalN-induced acute liver failure in mice through up-regulating expression of miR-122.

Coptisine (COP), one of the main active ingredients of Rhizoma Coptidis, reportedly has anti-inflammatory, anti-colon cancer properties, but it remains elusive whether COP owns hepatoprotective activity. Mice were pretreated with COP for 7d prior to lipopolysaccharide/d-galactosamine (LPS/D-GalN) administration to detect the hepatic protective effects of COP. The mechanism was explored in using HepG2 cells with low level of miR-122 and LO2 cells with high level of miR-122, combining with miR-122 agomir transfection by means of detecting the expression of miR-122 and proteins, clinical index and apoptosis. COP ameliorated the LPS/D-GalN-induced liver failure by lowering serum levels of ALT and AST, raising hepatic GSH and SOD levels, and maintaining the morphology of hepatocytes, along with an increase in miR-122 expression in mice. The results in vitro indicated that, after miR-122 mimic administration, the alone treatment of COP and the co-treatment of COP and LPS transfection obviously promoted the apoptosis of HepG2, which was increased by 152.67% and 113.97% compared with NC (P < 0.05 vs NC). LPS significantly induced the apoptosis of L02 cells, but COP treatment attenuated that of L02 cells. Further analysis showed that COP increased the miR-122 level and the expression of Bax, cleaved-casp3 and decreased Bcl-2, Bcl-xL in LPS-treated HepG2 cells. COP increased the miR-122 level but decreased the expression of TLR4, Bcl-2, Bcl-xL in LPS-treated L02 cells. COP attenuated LPS/D-GalN-induced ALF by up-regulating the level of miR-122, synergistically promoting apoptosis, and suggesting COP which showed a potential protective effect on ALF.

Use of Estradiol Promotes Graft-Bone Healing in Rabbit Model of Anterior Cruciate Ligament Reconstruction With a Polyethylene Terephthalate Ligament.

The purpose of this study was to investigate whether the local use of estradiol after anterior cruciate ligament (ACL) reconstruction with a polyethylene terephthalate (PET) artificial ligament graft could promote graft-bone healing. A total of 45 New Zealand white rabbits underwent ACL reconstruction with a PET ligament graft. The experimental groups were administered a local estradiol injection at either a low dose after surgery or a high dose after surgery, and the control group did not receive an injection. Computed tomography (CT) scans and blood sample collection were routinely performed in all three groups. Over time, the serum estradiol levels increased in both experimental groups, and the CT images revealed a trend of a shrinking bone tunnel area in all three groups. The rabbits were randomly sacrificed at 2, 4, and 8 weeks after surgery. The load to failure and stiffness of the experimental groups were significantly higher than those of the control group at 4 and 8 weeks. The histological study identified more bone mineralization in the experimental groups at 4 weeks after surgery compared to the control group. This study showed that the use of estradiol is a promising approach in promoting graft-bone healing in rabbits undergoing ACL reconstruction with a PET ligament graft.