Efficacy of Zhizhuwan Decoction and Its Ingredient-contained Serums on Proliferation and Apoptosis of Rat Colon Interstitial Cells of Cajal / 中国实验方剂学杂志

ABSTRACT

Objective:To explore the efficacy and molecular mechanism of Zhizhuwan decoction and its ingredient-contained serums on the proliferation and apoptosis of rat colon interstitial cells of cajal (ICC), and make a molecule-level analysis of the possible mechanism of traditional Chinese medicine (TCM) purgation-tonifying therapy in treating slow transit constipation (STC). Method:A total of 40 rats were divided into Atractylodis Macrocephalae Rhizoma(AMR) group, Aurantii Fructus Immaturus（AFI） group, Zhizhuwan group and blank serum group on random basis, with 10 in each group. Baizhu group was given 17.7 g·kg-1·d-1 of AMR decoction by gavage, AFI group was given 8.9 g·kg-1·d-1 AFI decoction by gavage, Zhizhuwan group was given 26.4 g·kg-1·d-1 Zhizhuwan decoction by gavage, and blank serum group was given 3 mL sterile distilled water for 7 consecutive days, once a day. Drug-contained serums and blank serum were collected from blood of the above groups and diluted to 5%, 10%, 15% and 20% concentrations. Each concentration was intervened for 24 h and 48 h, and the amount and status of ICC were observed. The best intervening concentration and time for each group with cell counting kit-8 (CCK-8) were determined. Rat colon ICC was divided into blank control group, blank serum group, AMR group, AFI group and Zhizhuwan group. ICC proliferation for each group was detected with EdU, ICC apoptosis for each group was detected by flow cytometry, and expressions of X-linked inhibitor of apoptosis protein (XIAP) and proliferating cell nuclear antigen (PCNA) were detected by Western blot. Result:Compared with the normal group, the best intervention concentration for blank serum group, AMR group and AFI group was 10%, while that for Zhizhuwan group was 5%. The best intervention times for the above groups were all 24 h. No distinct difference between the effect of blank control group and blank serum group on the proliferation and apoptosis of ICC was observed. In comparison with blank control group and blank serum group, AMR group, AFI group and Zhizhuwan group showed significant changes in ICC proliferation rate (P<0.05，P<0.01). There was a greater increase in ICC proliferation rate of Zhizhuwan group than that of AMR group and Zhizhu group (P<0.05，P<0.01), with no distinct difference between the changes of ICC proliferation rates in AMR group and AFI group. There was no significant difference between the changes of ICC apoptosis rates in AMR group, AFI group and Zhizhuwan group than in blank control group and blank serum group. There were significant increases in the expressions of XIAP and PCNA in AMR group, AFI group and Zhizhuwan group than in blank control group and blank serum group (P<0.05，P<0.01)， but with little difference among the three groups. Conclusion:At certain concentrations, Zhizhuwan, AFI and AMR all have the effect in improving ICC proliferation by increasing XIAP and PCNA expressions, with no evident effect on the apoptosis of ICC, based on TCM purgation-tonifying therapy, Zhizhuwan has the effect in improving ICC proliferation, with a better effect than single administration with AFI or AMR.