Study On The Effect Of Butyl Alcohol Extract Of Baitouweng Decoction On Ulcerative Colitis Colonized By Candida Albicans

Objective: To study the effect of Butyl alcohol extract of Bai Tou Weng decoction and Butanol Extract on the Ulcerative Colitis colonized by Candida albicans.Methods: At First,the total extract of Bai TouWeng decoction was extracted by 80%ethanol refluxing method,and the total extract of Bai Tou Weng decoction was extracted with different polar solvents.In combination with this study,the n-butanol extract was only suspended by rotary evaporator.(BAEB),and the activity of BAEB against Candida albicans was confirmed by XTT method.The model of acute ulcerative colitis in mice was induced by 3.5% DSS.After observing the symptoms of acute exacerbation of ulcerative colitis for seven days,the strain of Candida albicans(CA)standard strain SC5314 with a concentration of 1.6×109CFU/m L was given 0.5ml daily to replicate the intestines Candida albicans was used to treat ulcerative colitis mice model.After 5 days of modeling,BAEB was used to treat the mice.The morphological changes of the mice were observed.The levels of IL-6,IL-8,IL-1β,HBD-2 and HBD-3 in colonic mucosa of mice were detected by ELISA,and the changes of serum ASCA contents were measured by ELISA.The expression of Toll-like receptors 1,2,4,5(TLR1,TLR2,TLR4,TLR5)and Dectin-1 in colonic mucosa were detected by real-time quantitative PCR(q RT-PCR)Candida albicans was used to control the effect of Candida albicans on the yeast phase(in this paper,the culture of human colon cancer epithelial cells(Caco-2 cell line)was incubated with Candida albicans(CA)for 24 h,and the Caco-2cells survived by MTT assay.The levels of IL-6,IL-8,IL-1β,HBD-2 and HBD-3 in Caco-2 cell supernatants were detected by ELISA.Real-time quantitative PCR was performed by Toll-like receptors 1,2,4,5 were detected by Caco-2 cells and Dectin-1gene expression.Results : 1.The results showed that the antibacterial activity of the extract was 64~256μg/m L,and it was proved that the BAEB was better than that of the control(BAEB)Of the antifungal effect.2.Establishment of acute ulcerative colitis model in mice with DSS colonization(1)By observing the general activity of mice,weight,appearance changes.It was found that the mice in the DSS model group showed symptoms such as loss of appetite,slow activity and weight loss with the increase of the number of free drinking days,and accompanied by dilute stools.A small amount of blood was also found in the mouse anus,which was reported in the literature with ulcerative colitis In addition to the above state,the body size is more thin,almost no activity.(2)The changes of colonic tissue in mice showed that the increase of the number of days of free drinking 3.5% DSS showed that the phenomenon of colonic shortening and swelling was observed by observing the naked eye of the mice after the modeling,and the intestinal contents In the model group for the dilute,the normal group for the black granular,and ulcerative colitis colonic pathology is also basically consistent.(3)The model of ulcerative colitis was established in C.A,and the contents of intestinal contents were rinsed and collected.The results showed that C.A were grown in DSS+CA group,CA group and BAEB group(P<0.05).The C.A colonies in the treatment group was significantly lower than in the control group(P<0.05).The colonies in the DSS+CA group was significantly higher than that in the CA group(P<0.05).(4)Pathological observation: DSS model group and DSS+C.A model group,the surface of the colon mucosa can be seen a large number of inflammatory cell infiltration,glandular atrophy,colonic mucosal surface can be seen uneven ups and downs,ulcers and mucosal surface defects.After BAEB gavage treatment,the above pathological phenomenon was significantly reduced.(5)The q RT-PCR was used to detect the expression of TLR1,TLR2,TLR4,TLR5 and Dectin-1 in the intestinal mucosa homogenate of different groups.Compared with the control group,the DSS model group and Candida albicans infected mice The expression levels of TLR1,TLR2,TLR4,TLR5 and Dectin-1 in tissue homogenatewere significantly increased(P<0.05),while BAEB significantly reduced TLR1,TLR2,TLR4,TLR5,And the expression level of Dectin-1(P <0.05).(6)The expression of ASCA in the serum of DSS+C.A group was significantly higher than that of DSS group(P>0.05),but there was no significant difference between the two groups(P>0.05)。(7)The levels of IL-6,IL-8,IL-1β,HBD-2 and HBD-3 in the supernatant of intestinal tissue homogenate of mice were detected by ELISA.Compared with the blank control group,DSS and DSS+C.A groups,The levels of IL-8,IL-6,IL-1β,HBD-2、3were significantly higher than those in DSS+CA group.The BAEB group IL-6,IL-1β,HBD-2 and HBD3 were significantly decreased,indicating that BAEB high group can reduce the level of inflammatory expression.(8)MTT assay was used to screen the Caco-2 cells to protect the ideal intervention concentration of BAEB under BAEB pre-protection of Candida albicans heat inactivated spores.Caco-2 cells were significantly inhibited by Caco-2 cells treated with 80 μg/m L BAEB for 24 h and 3×106CFU/m L C.A for Caco-2 cells for 24 h.(9)The levels of IL-8,IL-6,IL-1β,HBD-2 and HBD3 in the supernatant of Caco-2 cells treated with BAEB were detected by ELISA.The results showed that compared with the control group,IL-8,IL-6,IL-1β,HBD-2 and HBD3 were significantly increased,suggesting that the Caco-2 cell model was successfully established by CA.Compared with the model group,the high dose group of BAEB The above expression in the supernatant of the cells was significantly reduced,indicating that BAEB high dose can reduce the level of inflammatory expression.(10)The effect of BAEB on the expression of TLRs and Dectin-1 in Caco-2 cells was detected by q RT-PCR.The results showed that the expression levels of TLR1,TLR2,TLR4,TLR5 and Dectin-1 were significantly up-regulated in Caco-2 cells when cultured with heat-inactivated Candida albicans,while BAEB was significantly lower in BAEB TLR1,TLR2,TLR4,TLR5,Dectin-1.This is consistent with the experimental results in vivo.Conclusion: 3.5% DSS free drinking seven days after induction of acute ulcerative colitis in mice,Candida albicans can be replicated after Candida albicans-induced ulcerative colitis model and aggravate the expression of ulcerative colitis,by BAEB treatment Can effectively alleviate the expression of inflammatory factors.Candida albicans inactivated spores can stimulate the expression of inflammatory factors,defensins and TLRs in Caco-2 cells,while BAEB preconditions can reduce the expression of inflammatory factors,defensins and TLRs receptors.Indicating that BAEB not only can directly inhibit the growth of Candida albicans,and Candida albicans caused by the inherent immune process involved in the intestinal mucosal inflammatory injury also improved.