Si-Ni-San reduces lipid droplet deposition associated with decreased YAP1 in metabolic dysfunction-associated fatty liver disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Metabolic dysfunction-associated fatty liver disease (MAFLD) is the most common chronic liver disease worldwide. However, its complex pathogenesis and lack of effective drugs for treating it present significant challenges. Si-Ni-San (SNS) is one of the representative formulas for treating patients with MAFLD in traditional Chinese medicine (TCM) clinics. According to our previous work, SNS reduces lipid droplet (LD) deposition in livers of mice with MAFLD. AIM OF THE STUDY: To elucidate the mechanism of SNS in reducing LD deposition in MAFLD. MATERIALS AND METHODS: First, LD areas were detected with Oil red O staining in HepG2 cells induced by oleic acid (OA). Cell Counting Kit-8 (CCK-8) assay was used to test cell viability after treatment with different concentrations of SNS serum. The expression of Yes-associated protein 1 (YAP1) was monitored by Western blot. Second, C57BL/6 mice were fed a high-fat diet (HFD) for 12 weeks and gavaged with SNS decoction during the 11th and 12th weeks. Then, the weight of the body and the liver was examined. LD numbers and their locations in the liver were detected by triglyceride (TG) assay and hematoxylin and eosin staining (H&E). The expression levels of YAP1 and perilipin2 (PLIN2) were detected using Western blot and immunohistochemistry (IHC) in liver tissues. Finally, active ingredients of SNS decoction and SNS serum were identified by liquid chromatography-mass spectrometry (LC-MS). Finally, molecular docking was performed between the compounds in SNS and YAP1 to analyze their active interaction. RESULTS: Cellular experiments showed that SNS serum reduced LD vacuoles and YAP1 expression in OA-induced HepG2 cells. Animal experiments confirmed that LD vacuoles, PLIN2 expression (3.16-fold), and YAP1 expression (2.50-fold) were increased in the HFD group compared with the normal diet (ND) group. SNS reduced LD vacuoles, TG content (0.84-fold), PLIN2 expression (0.33-fold), and YAP1 expression (0.27-fold) compared with the normal saline (NS) group in Yap1Flox mice with MAFLD. In SNS, baicalein-6-glucuronide, desoxylimonin, galangin-7-glucoside, glycyrrhizic-acid, licoricesaponin-K2, and nobiletin showed a high binding effect with YAP1. Knockout of hepatocyte YAP1 reduced LD vacuoles, TG content (0.40-fold), and PLIN2 expression (0.62-fold) in mice. Meanwhile, SNS reduced LD vacuoles, TG content (0.70-fold), and PLIN2 expression (0.19-fold) in Yap1LKO mice with MAFLD. The effect of SNS in reducing TG and PLIN2 was diminished in Yap1LKO mice compared with Yap1Flox mice. CONCLUSION: SNS reduced LD deposition and YAP1 expression in MAFLD liver cells both in vivo and in vitro. YAP1 was highly expressed in livers with MAFLD, and knockout of hepatocellular YAP1 reduced LD deposition in mice. SNS reduced LD deposition associated with decreased YAP1 in MAFLD liver cells.

Modulation of Oral Microflora during Helicobacter Pylori Eradication.

OBJECTIVE: To analyse the saliva microbial abundance and composition by 16s rRNA sequence during Helicobacter pylori (H.pylori) eradication. STUDY DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: Hebei University of Chinese Medicine, Hebei Provincial Hospital of Traditional Chinese Medicine, from March 2019 to January 2020. METHODOLOGY: The saliva microbial were analysed before and after the bismuth-containing quadruple therapy. A total of ten saliva samples (three groups) were enrolled in the study. The authors used the linear discriminant analysis effect size (LEfSe) method and Welch's t-test for comparative analysis to identify which taxa could be significantly affected in three groups. RESULTS: H.pylori 16S rRNA gene sequence was not detected in the ten saliva samples. The abundance of Prevotella_sp._oral_clone_P4PB_83_P2 from healthy adults was higher than H.pylori-positive patients. Moreover, after the bismuth-containing quadruple therapy, the diversity and richness of saliva bacteria reduced. Lautropia, Burkholderiales, uncultured bacterium, Burkholderiaceae, and Actinomyces were enriched in H.pylori-positive patient samples after the bismuth-containing quadruple therapy. CONCLUSION: The diversity and richness of salivary microbiome were reduced in H.pylori-positive patient, and bismuth-containing quadruple therapy affected oral microbiota. Key Words: Helicobacter pylori, Saliva, Microbiota, RNA, Bismuth.

[N-acetylcysteine inhibits the proliferation of hydrogen peroxide treated fibroblast-like synoviocytes in rats with adjuvant arthritis (AA) via blocking Nrf2/Keap1 pathway].

Objective To investigate the effect of N-acetylcysteine (NAC) on the proliferation of fibroblast-like synoviocytes (FLS) treated with low concentration of hydrogen peroxide (H2O2) in rats with adjuvant arthritis (AA) and its mechanism. Methods Twenty SD rats were divided into a normal group and a model group (10 rats in each group). The model group was established by subcutaneous injection of Freund's complete adjuvant into the toe of rats, and the rats were sacrificed 28 days later. The contents of serum malondialdehyde (MDA) were detected by thiobarbituric acid method; the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were determined by hydroxylamine method and colorimetry respectively; and Nrf2 and Keap1 proteins in ankle synovial tissues of AA rats were detected by immunohistochemistry. AA-FLS were isolated, cultured, and identified by digestion of ankle joint slides of AA rats in vitro. The effects of NAC at different concentrations (final concentration 0, 0.3, 0.9, 3, 10, 30, 90, 180 µmol/L) on the activity of AA-FLS treated with H2O2 at low concentration (5 µmol/L) were detected by CCK-8 assay. The content of mitochondrial reactive oxygen species (ROS) in AA-FLS was detected by MitoSOX fluorescent probe. The effects of NAC (final concentration 0, 3, 10, 30 µmol/L) on Nrf2 and Keap1 protein expressions in AA-FLS treated with H2O2 at low concentration were detected by Western blotting. Results Compared with those in the control group, in AA model, the MDA level increased and SOD and GSH-Px levels decreased in serum, and the Nrf2 protein increased and the Keap1 protein decreased in synovial tissue. Immunocytochemical staining confirmed that the isolated and cultured cells were AA-FLS; NAC inhibited the proliferation of AA-FLS treated with H2O2 in a concentration-dependent manner, and the mitochondrial ROS content and the protein expressions of Nrf2 and Keap1 decreased. Conclusion NAC can inhibit the proliferation of AA-FLS treated with H2O2, which may be related to blocking Nrf2/Keap1 pathway.