Prolonged administration of total glucosides of paeony improves intestinal immune imbalance and epithelial barrier damage in collagen-induced arthritis rats based on metabolomics-network pharmacology integrated analysis.

Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease characterized by synovial inflammation and joint damage with complex pathological mechanisms. In recent years, many studies have shown that the dysregulation of intestinal mucosal immunity and the damage of the epithelial barrier are closely related to the occurrence of RA. Total glucosides of paeony (TGP) have been used clinically for the treatment of RA in China for decades, while the pharmacological mechanism is still uncertain. The purpose of this study was to investigate the regulatory effect and mechanism of TGP on intestinal immunity and epithelial barrier in RA model rats. The results showed that TGP alleviated immune hyperfunction by regulating the ratio of CD3+, CD4+ and CD8+ in different lymphocyte synthesis sites of the small intestine, including Peyer's patches (PPs), intraepithelial lymphocytes (IELs), and lamina propria lymphocytes (LPLs). Specially, TGP first exhibited immunomodulatory effects on sites close to the intestinal lumen (IELs and LPLs), and then on PPs far away from the intestinal lumen as the administration time prolonged. Meanwhile, TGP restores the intestinal epithelial barrier by upregulating the ratio of villi height (V)/crypt depth (C) and expression of tight junction proteins (ZO-1, occludin). Finally, the integrated analysis of metabolomics-network pharmacology was also used to explore the possible regulation mechanism of TGP on the intestinal tract. Metabolomics analysis revealed that TGP reversed the intestinal metabolic profile disturbance in CIA rats, and identified 32 biomarkers and 163 corresponding targets; network pharmacology analysis identified 111 potential targets for TGP to treat RA. By intersecting the results of the two, three key targets such as ADA, PNP and TYR were determined. Pharmacological verification experiments showed that the levels of ADA and PNP in the small intestine of CIA rats were significantly increased, while TGP significantly decreased their ADA and PNP levels. In conclusion, purine metabolism may play an important role in the process of TGP improving RA-induced intestinal immune imbalance and impaired epithelial barrier.

[miR-153 aggravates lung injury induced by lipopolysaccharide via inhibiting activated protein C (APC) in rats with sepsis and its mechanism].

Objective To investigate the mechanism of miR-153 targeting activated protein C (APC) to regulate lipopolysaccharide (LPS)-induced lung injury in rats with sepsis. Methods Female SD rats were divided into control group, LPS group, LPS combined with miR-153 inhibitor (miR-153 inhibitor) group, LPS combined with miR-153 inhibitor negative control (inhibitor NC) group, LPS and miR-153 inhibitor combined with APC small interfering RNA (si-APC) group, LPS and miR-153 inhibitor combined with APC small interfering RNA negative control (si-NC) group. Except for the control group, the other groups were given corresponding treatments, and then LPS were given to establish rat sepsis injury models. Real-time quantitative PCR was used to detect the expression of miR-153 and Western blot analysis to detect the protein expression of APC, B-cell lymphoma 2 (Bcl2) and cleaved caspase-3 (c-caspase-3). The rat alveolar epithelial cells were isolated and cultured, and their cell viability was detected by CCK-8 assay, along with cell apoptosis detected by flow cytometry. ELISA was performed to test the expression levels of superoxide dismutase (SOD), malondialdehyde (MDA), interleukin 6 (IL-6), IL-1ß and tumor necrosis factor α (TNF-α) in rat serum; and the dual fluorescein reporter experiment detects the relationship between miR-153 and APC. Results Compared with the control group, rat model of sepsis lung injury showed significantly increased expression of miR-153 and reduced APC. The cell viability, Bcl2 protein expression and SOD activity of the LPS group were significantly reduced, and the apoptosis rate, c-caspase-3 protein expression, MDA, IL-6, IL-1ß, and TNF-α content significantly escalated. Compared with the LPS combined with inhibitor NC group, the cell viability, Bcl2 protein expression and SOD activity of the LPS combined miR-153 inhibitor group significantly increased, while the apoptosis rate, c-caspase-3 protein expression, the content of MDA, IL-6, IL-1ß and TNF-α dropped considerably. miR-153 can target and regulate the expression of APC. Compared with the LPS and miR-153 inhibitor combined with si-NC group, the cell viability, Bcl2 protein expression and SOD activity of the LPS and miR-153 inhibitor combined with si-APC group significantly down-regulated, and the apoptosis rate, c-caspase-3 protein expression, MDA, IL-6, IL-1ß, and TNF-α contents increased significantly. Conclusion LPS induces increased expression of miR-153 in lung tissue, inhibits APC expression, promotes apoptosis, oxidative stress and inflammatory response in lung tissue of septic rats, and aggravates damage.

Dynamic Alterations in the Gut Microbiota of Collagen-Induced Arthritis Rats Following the Prolonged Administration of Total Glucosides of Paeony.

Rheumatoid arthritis (RA) is a common autoimmune disease linked to chronic inflammation. Dysbiosis of the gut microbiota has been proposed to contribute to the risk of RA, and a large number of researchers have investigated the gut-joint axis hypothesis using the collagen-induced arthritis (CIA) rats. However, previous studies mainly involved short-term experiments; very few used the CIA model to investigate changes in gut microbiota over time. Moreover, previous research failed to use the CIA model to carry out detailed investigations of the effects of drug treatments upon inflammation in the joints, hyperplasia of the synovium, imbalance in the ratios of Th1/Th2 and Th17/Treg cells, intestinal cytokines and the gut microbiota following long-term intervention. In the present study, we carried out a 16-week experiment to investigate changes in the gut microbiota of CIA rats, and evaluated the modulatory effect of total glucosides of paeony (TGP), an immunomodulatory agent widely used in the treatment of RA, after 12 weeks of administration. We found that taxonomic differences developed in the microbial structure between the CIA group and the Control group. Furthermore, the administration of TGP was able to correct 78% of these taxonomic differences, while also increase the relative abundance of certain forms of beneficial symbiotic bacteria. By the end of the experiment, TGP had reduced body weight, thymus index and inflammatory cell infiltration in the ankle joint of CIA rats. Furthermore, the administration of TGP had down-regulated the synovial content of VEGF and the levels of Th1 cells and Th17 cells in CIA rats, and up-regulated the levels of Th2 cells and Treg cells. The administration of TGP also inhibited the levels of intestinal cytokines, secretory immunoglobulin A (SIgA) and Interferon-γ (IFN-γ). In conclusion, the influence of TGP on dynamic changes in gut microbiota, along with the observed improvement of indicators related to CIA symptoms during 12 weeks of administration, supported the hypothesis that the microbiome may play a role in TGP-mediated therapeutic effects in CIA rats. The present study also indicated that the mechanism underlying these effects may be related to the regulation of intestinal mucosal immunity remains unknown and deserves further research attention.