Pollen typhae total flavone suppresses A7r5 cell proliferation promoted by insulin involving the MEK1/2-ERK1/2 cascades.

Pollen typhae, a traditional medicine in China, performs an anti-diabetic function and has anti-atherosclerosis effects involving suppression of vascular smooth muscle cell proliferation. However, the potential mechanisms keep to be revealed. The present study intended to investigate the influences of Pollen typhae extract named Pollen typhae total flavone (PTF) on A7r5 cell proliferation promoted by insulin and to uncover the underlying mechanisms. Proliferation and viability were evaluated by CCK-8 method. Western blotting was adopted to analyze the protein expression. Insulin promoted A7r5 cell proliferation, while PTF suppressed insulin-promoted proliferation in a concentration-dependent fashion. Although PTF did not change c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38MAPK) or MAPK kinase 1/2 (MEK1/2) protein expression and failed to affect the phosphorylation of JNK and p38MAPK, PTF remarkably inhibited extracellular signal-regulated kinase 1 and 2 (ERK1/2) protein expression and reduced ERK1/2 and MEK1/2 phosphorylation in A7r5 cells stimulated by insulin. Insulin-induced proliferation of A7r5 cells was abolished by inhibiting ERK1/2, which was in line with PTF. These findings indicate that PTF suppresses insulin-promoted proliferation of A7r5 cells involving the MEK1/2-ERK1/2 cascades, providing new insight into the potential uses of PTF for treatment of diabetic atherosclerosis.

Effect of Huangqin decoction on regulating intestinal flora in colitis mice characterized as inhibition of the NOD2-dependent pathway.

CONTEXT: Chinese herb Huangqin decoction (HQD) can regulate intestinal flora in ulcerative colitis (UC) mice. OBJECTIVE: Our study clarifies the mechanism of HQD in regulating the intestinal flora of UC mice. MATERIALS AND METHODS: Male C57BL/6 mice were randomly divided into six groups: Control, Model (3% DSS), Sulfasalazine (500 mg/kg), HQD-L (250 mg/kg), HQD-M (500 mg/kg), and HQD-H (1000 mg/kg) groups. Measurement of body weight, colon length, DAI, and haematoxylin-eosin staining were conducted. FISH and 16S rDNA detected colonic bacterial infiltration and intestinal flora changes. The expression of RegIIIγ and PRRs (NOD2, TLR5, TLR4) were detected by FCM and WB, respectively. In addition, WB, qPCR, or IHC were used to detect the expression of NOD2, MyD88, RIP2, and NF-κB p65 in the colon. ELISA was used to determine cytokines. RESULTS: Compared with the model group (DAI score, 2.38 ± 0.05; histological score, 4.08 ± 0.54), HQD treatment significantly reduced the DAI score (L, 2.16 ± 0.09; M, 1.45 ± 0.05; H, 1.18 ± 0.05) and histological score (L, 3.16 ± 0.82; M, 2.50 ± 0.81; H, 1.51 ± 0.76); restored the weight, the colonic length (p < 0.05). 16S rDNA identification showed HQD regulated the balance of intestinal flora. Moreover, HQD suppressed the expression of RegIIIγ (p < 0.05) and prevented colonic bacterial infiltration. Furthermore, WB results showed NOD2, and TLR4 were inhibited by HQD, especially NOD2 (p < 0.01). The data of WB, qPCR, and IHC demonstrated that the NOD2-dependent pathway was inhibited by HQD (p < 0.01). DISCUSSION AND CONCLUSIONS: HQD (1000 mg/kg) regulates the intestinal flora of colitis mice, mainly characterized as inhibition of the NOD2-dependent pathway. These results indicate that HQD has potential.

Rhubarb Peony Decoction ameliorates ulcerative colitis in mice by regulating gut microbiota to restoring Th17/Treg balance.

ETHNOPHARMACOLOGY RELEVANCE: Rhubarb Peony Decoction (RPD) is a formula of traditional Chinese medicine chronicled in Jin Gui Yao Lve, commonly used to treat ulcerative colitis (UC). However, the underlying mechanism of RPD treating UC remains elusive. In our study, we investigated the therapeutic efficacy of RPD and potential mechanism involved in inhibiting dextran sulfate sodium (DSS)-induced ulcerative colitis in mice. METHODS: The colitis was induced by DSS in mice for 5 days and estimated body weight loss, disease activity index (DAI) and colon length. Histological changes were observed by H&E staining. The number and abundance of gut mircrobiota were measured with 16 S rDNA sequencing. GC-MS was used to detect the concentration of short chain fatty acids (SCFAs) in cecum. Flow cytometry analyzed the proportion of Th17 and Treg cells in mesenteric lymph nodes (MLNs). IL-17A and Foxp3 in colon were determined by immunohistochemical analyses. The level of cytokine was determined by Multi-Analyte Flow Assay Kit. RESULTS: Administration of RPD significantly alleviated the pathological changes of UC mice, involving rescued the inflammation-related reduction of colon length, ameliorated body weight loss and damaged tissue. In addition, RPD altered the gut microbiota, involving restored α diversity, increased significantly the abundance of Firmicutes and Actinobacteria, decreased the Proteobacteria and Bacteroidetes. Furthermore, the number of Butyricicoccus pullicaecorum, a butyrate-producing bacterium, were augmented obviously by RPD. Besides, RPD restored the content of SCFA in intestinal tract. Additionally, the proportion of Th17 cells and Treg cells in mesenteric lymph nodes, likewise, the expression of IL-17A and Foxp3 in colon were regulated by RPD, contributing to the restoration of Th17/Treg balance. Moreover, RPD significantly decreased the level of IL-6, TNF-α, IFNγ, IL-10, IL-17A, IL-21, IL-22 in colon, simultaneously increased Treg-related cytokine TGF-ß at dose-dependently. CONCLUSIONS: These results demonstrated that RPD had effect on ulcerative colitis, which was related to regulating gut microbiota, especially Butyricicoccus pullicaecorum, and SCFAs to restore the gut Th17/Treg homeostasis.

[Shengqifuzheng Injection promotes the recovery of B cells in gut-associated lymphoid tissues of mice treated with cyclophosphamide].

Objective To investigate the effect of Shengqifuzheng Injection (SQFZ) on the number recovery of B cells in gut-associated lymphoid tissues (GALTs) of mice receiving cyclophosphamide-based chemotherapy. Methods BALB/c mice were randomly divided into control group, cyclophosphamide (Cy) group and SQFZ group. Mice in Cy group and SQFZ group were injected intraperitoneally with Cy (100 mg/kg), while the control mice were injected with an equal volume of normal saline. Twenty-four hours later, mice in SQFZ group were administrated intragastricly with 1 mL SQFZ once daily for 10 consecutive days, and mice in the other groups were given the same volume of normal saline. Body mass of all the mice was measured every day. Mice were killed on day 10, and the indexes of spleen and thymus were measured. Cell cycles of bone marrow cells and the percentage of B cells in lymphocytes in mesenteric lymph node (MLN) and Peyer's patch (PP) were detected by flow cytometry. In vitro, after being treated with SQFZ, activity of lymphocytes was evaluzed by MTT assay; expression of CD86 on B cell surface was analyzed by flow cytometry; and B cell proliferation was tested by carboxyfluorescein succinimidyl ester (CFSE)-based lymphocyte proliferation assay. Results SQFZ alleviated the loss of body mass caused by Cy and promoted the recovery of thymus indexes, spleen indexes and B cell number in MLN and PP. But it did not alleviate the bone marrow suppression of mice in this condition. In vitro, SQFZ enhanced lymphocyte activity, and improved the activation and proliferation of B cells. Conclusion SQFZ could accelerate the recovery of B cells in GALTs of mice receiving chemotherapy and it might act by promoting B cell proliferation.