Therapeutic Mechanism of Zhishi Decoction Regulating P38/MAPK Signaling Pathway on Functional Constipation (FC)

Background: Zhishi decoction (ZSD) is one of the most common herb decoctions in traditional Chinese medicine (TCM), and it is used for the treatment of FC. However, its main therapeutic mechanism is not yet clear. This study aims to explore the possible pharmacodynamic material basis and potential molecular mechanism from network pharmacology and molecular docking and verify them through animal experiments. Methods: Firstly, the effective ingredients, potential targets, and key targets of ZSD in the treatment of FC were screened through network pharmacology. Go and KEGG analyses were performed for potential targets. Secondly, molecular docking was used to link the main active components of ZSD with target genes to predict their possible molecular mechanisms. Finally, 30 male BALB/c mice (20±2 g) were randomly divided into five groups (n=6), including the blank group, ZSD groups with two dosages (7.15, 14.3 g/kg), FC model group, and positive group (lactulose group). All the mice were given difenoxate tablets for 14 days to establish FC model except the blank group. Moreover, the mice in the blank group were given the same volume of normal saline. After admination for 14 days, the whole colon tissues were obtained for the analysis of small intestinal propulsion rate, and the expression of P38MAPK in colon tissues of mice was observed via immunohistochemistry and WesterBlot. Results: In this study, 43 active ingredients in ZSD were identified. Four hundred and thirty potential therapeutic targets were selected, among which AKT1, MAPK12, and MAPK14 were key targets. 164 GO biological processes and 123 KEGG signaling pathways were identified after analysis， such as MAPK signaling pathway, TNF signaling pathway etc. The molecular docking results showed that Prangenin, 4-Hydroxyhomopterocarpin, isoponcimarin, and AKT1, MAPK12, MAPK14 had good binding degree. Additionally, ZSD could relieve the symptoms of FC in mice significantly. Compared with the model group, p38/MAPK positive expression cells and protein expression levels in the colon tissues of ZSD groups significantly increased in a dose-dependent manner (p<0.01). Conclusion: This study confirmed that ZSD could act on AKT1, MAPK12, and MAPK14 targets to activate the p38/MAPK signaling pathway to relieve FC induced by defenoxate tablets. The further development of ZSD provided a theoretical basis. result: In this study, 43 active ingredients and 640 targets in the ZSD were identified , and 7172 targets related to FC were screened. In the final screening, 430 drug-disease common targets were obtained, among which AKT1, MAPK12 and MAPK14 were the key targets. 164 GO biological processes and 123 KEGG signaling pathways were identified after analysis，such as MAPK signaling pathway, TNF signaling pathway, et al. Molecular docking results showed that Prangenin, 4-Hydroxyhomopterocarpin, isoponcimarin and AKT1, MAPK12, MAPK14 had good binding degree. ZSD could relieve the symptoms of FC in mice significantly. Compared with model group, p38/MAPK positive expression cells and protein expression levels in colon tissues of ZSD groups increased as dose-depandant manner signifitantly (p<0.01) .