Pharmacological Study On Helicteres Angustifolia L. Ameliorating Pro-malignancy Shift In Ulcerative Colitis Via JAK/STAT Pathway

ObjectiveBased on cell proliferation test,cell inflammation model experiments and the AOM/DSS-induced malignant transformation models in UC mice,to observe the effects of Helicteres angustifolia L.(HA)on the proliferation of colorectal cancer cells and the secretion of inflammatory factors by macrophages,and to observe the effects of HA on the influence of the clinical symptoms of AOM/DSS-induced malignant transformation model in UC mice,explore the basic pharmacodynamics and mechanism of HA against UC malignant transformation,and provide basic theory and experimental basis for the utilization and clinical application of HA.Methods1.Observe the inhibition of HA on the proliferation of human colorectal cancer HCT-116 and Caco-2 cell lines in vitro drug experiments,and the effect of HA on the inflammatory factors secretion of RAW264.7 cells,which stimulated by LPS,in cell inflammation models.2.Via establishing the AOM/DSS induced UC malignant mouse model,to observe the weight,colorectal length,colorectal coefficient,organ coefficient of UC malignant mouse model mice by HA administration,colorectal histopathological changes and the influence of pathological scores,in order to evaluate the basic efficacy of HA against the UC malignancy transformation.The enzyme-linked immunosorbent assay(ELISA)was used to detect the pro-inflammatory cytokines interleukin 1β(IL-1β),interleukin 6(IL-6),and tumor necrosis factor-α(TNF-α)and anti-inflammatory cytokine interleukin 10(IL-10)content,Western Blot was used to determine the content of JAK2,STAT3,p-STAT3,NF-κB-P65,p-NF-κB-P65 in mouse colorectal tissues,to explore the anti-UC malignant mechanism of HA.Results1.HA has proliferation inhibitory effects on human colorectal cancer HCT-116 and Caco-2 cell lines,and its IC50 is 133.13μg/m L and 188.67μg/m L,respectively.HA has a significant inhibitory effect on the inflammatory response of RAW264.7 cells induced by LPS.Compared with the model group,HA administration can reduce the levels of pro-inflammatory cytokines IL-1β,IL-6 and TNF-α in the cell culture supernatant(P<0.05),while the level of anti-inflammatory factor IL-10 was increased(P<0.05).2.During the administration of DSS,the weight of the mice in the model group showed a significant downward trend,and the administration of HA could reduce the weight loss of the mice.In terms of organ coefficient,compared with the model group,the liver coefficient and spleen coefficient of the mice in each HA administration group were significantly decreased(P<0.05),while the thymus coefficient was significantly increased(P<0.05).In terms of colorectal histopathology,the malignant rate of mice in the model group was 87.5%(7/8),while the malignant rates of each HA administration group were12.5%(1/8),25%(2/8),and 25%(2/8),respectively.In terms of pathology scores,compared with the model group,the inflammatory pathology scores,dysplasia and cancer scores of mice at each doses of HA were decreased significantly(P<0.05).AOM/DSS induced malignant transformation of UC can increase the levels of pro-inflammatory cytokines IL-1β,IL-6 and TNF-α in the serum of mice(P<0.05),and decreased the content of anti-inflammatory factor IL-10(P < 0.05),compared with the model groups,the serum levels of pro-inflammatory cytokines IL-1β,IL-6 and TNF-α in the HA administration groups were decreased(P<0.05),and the anti-inflammatory factor IL-10 content was increased(P < 0.05).Compared with the model group,each HA administration group can significantly reduce the expression of JAK2,STAT3,p-STAT3,NF-κB-P65,and p-NF-κB-P65 in colorectal tissues(P<0.01).Conclusion1.HA has in vitro proliferation inhibitory activity on human colorectal cancer HCT-116 and Caco-2 cell lines;HA can inhibit the inflammatory response of the RAW264.7 cell line induced by LPS and reduce the pro-inflammatory cytokines IL-1β,IL-6,TNF-α and increase the secretion of anti-inflammatory cytokine IL-10,which means that HA have anti-inflammatory effects.2.AOM/DSS-induced malignant transformation of UC in mice can simulate the process of "inflammation" to "cancer",which is similar to CRC caused by repeated episodes of human UC.Weight loss,hematochezial,diarrhea,malignant transformation of colorectal tissues,elevated serum pro-inflammatory cytokines,and continuous activation of IL-6/JAK/STAT signaling pathways and NF-κB signaling pathways are some of the clinical symptoms and pathological changes of this model.HA can reduce weight loss,improve bloody stool and diarrhea and other clinical symptoms;it can maintain the organ coefficients of liver,spleen,and thymus;and can reduce the probability of malignant transformation of UC in mice;by down-regulating the pro-inflammatory cytokines IL-1βand IL-6,TNF-α and increase the content of anti-inflammatory cytokine IL-10 in serum,and down-regulate the expression of JAK2,STAT3,p-STAT3,NF-κB-P65,p-NF-κB-P65 protein in colorectal tissue,and regulate IL-6/JAK/STAT signaling pathway and NF-κB signaling pathway inhibit the body’s systemic inflammatory response,reduce the degree of inflammation and tissue damage in colorectal tissues,thereby inhibiting the development of UC to CRC.