ABSTRACT
Aim To investigate the feasibility and mechanism of rhynchophylline in the treatment of in-rhynchophylline flammatory bowel disease (IBD) based on network pharmacology combined with in vivo and in vitro experiments. Methods The target of rhynchophylline-IBD intersection was obtained from the database, and GO and KEGG enrichment analysis were performed. The binding of key target proteins was screened by molecular docking. In vivo the IBD model of mice was induced by sodium dextran sulfate (DSS). After seven days of rhynchophylline intervention, the signs of mice in each group were observed and DAI scores were recorded. The levels of interleukin-1β (3 (IL-1 β), my-eloperoxidase (MPO) and other inflammatory factors in colon tissue of mice were detected by ELISA. The intestinal permeability of each group was detected. In vitro experiments were conducted to establish the inflammatory model of Caco2 cells induced by DSS, and to clarify the regulatory effect of leptosinine on key targets. Results A total of 70 rhynchophylline-IBD intersection targets were screened, and enrichment analysis showed that they were related to the inflammatory prooess, PI3K-Akt and Hippo signaling pathway s. Molecular docking results showed that was most stable in binding with JAK2 and JAK1. In vivo experiment results showed that compared with model group, body weight, colon length and weight of rhynchophylline group significantly increased (P < 0. 05). DAI score, IL-1β, MPO and other inflammatory factors in colon tissue and intestinal permeability significantly decreased (P < 0. 01). In vitro experiment results showed that compared with model group, rhynchophylline group significantly promoted the proliferation of Caco2 cells (P < 0. 05). The levels of IL-6 and NO were significantly reduced (P < 0. 05). Western blot results showed that rhynchophylline could decrease the expressions of JAK2 and JAK1 (P < 0. 05). Conclusion Rhynchophylline may play a role in the treatment of IBD by inhibiting the expression of JAK2 and JAK1 proteins and reducing inflammatory response in body.
ABSTRACT
Gemcitabine (GEM) is a commonly used drug in the clinical treatment of non-small cell lung cancer. Due to the accumulation of cells mediating immune escape and T cell depletion after chemotherapy, tumor microenvironment (TME) tends to be immunosuppressive status, which ultimately leads to tumor metastasis. The experimental protocol was approved by the Medical Laboratory Animal Ethics Committee of Jiangsu Provincial Academy of Chinese Medicine. Therefore, we observed the immunomodulatory effects of micro-particulate Ganoderma lucidum spore β-glucan (PGSG) on macrophages in vitro experiments. Next, mice subcutaneous Lewis lung cancer models were established to observe the anti-tumor effects of PGSG through oral administration of PGSG combined with GEM. Flow cytometry analysis was used to analyze the ratio of anti-tumor T cells in tumors and spleen, as well as the proportion of myeloid-derived suppressor cells (MDSC), tumor-associated macrophages (TAM) and regulatory cells (Tregs). The results showed that PGSG can up-regulate the expression of major histocompatibility antigens (MHC-II), CD40, CD86 and CD80 on the surface of macrophages, enhance the ability to phagocytosis of neutral red and further mediate the release of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interleukin-4 (IL-4) and interleukin-10 (IL-10). In vivo experiments, combined administration can significantly decrease the volume and weight of tumors, reduce the ratio of MDSC (CD11b+Gr-1+), M-MDSC (CD11b+Ly6G-Ly6Chigh) and Treg (CD4+Foxp3+). At the same time, PGSG promoted the conversion of M2 (F4/80+CD206+) to M1 (F4/80+MHC-II+) and enhanced the response of helper T cell-1 (Th1) (CD4+IFN-γ+) and cytotoxic T lymphocyte (CTL) (CD8+IFN-γ+), which is of great significance for killing tumors. These results suggest that PGSG can regulate innate and adaptive antitumor immune responses, reshape the immunosuppressive microenvironment and enhance the anti-lung cancer effect of GEM.
ABSTRACT
To investigate the changes in activation and proliferation of Tregs after targeted RNA inter-ference of Kvl. 3 channel genes and in vitro administration of eplerenone (EPL). Methods After lenti-virus vector was transfected to regulatory T cells (Tregs) of rats, qPCR and whole-cell patch-clamp methods were used to detect gene knockout efficiency, and EOSA method was used to detect cytokine secretion IL-10 and TGF-p levels of Tregs group,Tregs + EPL group,RNAi-Tregs group,and RNAi-Tregs + EPL group. Results Lentivirus vector was successfully transfected into Tregs cells, and the mRNA level and current density of Kvl. 3 channel was 78% and 71.3% respectively; compared with Tregs group, extracellular and intracellular TGF-p levels in RNAi-Tregs group were significantly reduced (P < 0. 01), and extracellular and intracellular TGF-(3 levels in Tregs + EPL group were also reduced (P < 0. 05 ) ; compared with RNAi-Tregs group, extracellular and in-tracellular TGF-fJ levels in RNAi-Tregs + EPL group showed no change. However, IL-10 levels in Tregs group,Tregs + EPL group, RNAi-Tregs group, RNAi-Tregs + EPL group showed no significant change. Conclusions Kvl.3 channel mediates the activation and proliferation of Tregs cells, while EPL can reduce the activation and proliferation of Tregs cells by directly inhibiting Kvl.3 channel and reducing the secretion of TGF-fi levels, further indicating that EPL is a specific blocker of Kvl. 3 channel.