| Objective:1.To investigate the role of RAGE/NF-κB signaling pathway in intestinal mucositis model.2.Observe the protective effect of moxibustion on chemotherapyinduced intestinal mucositis.3.To explore the mechanism of moxibustion in the intervention of chemotherapy-induced intestinal mucositis by regulating the RAGE/NF-κB signaling pathway.Methods:(1)Stimulate Caco-2 cells by LPS to simulate intestinal mucosal inflammation in vivo,using CCK-8 to detect the effect of LPS and FPS-ZM1 on the proliferation of Caco-2 cells;Western blot was used to detect the protein expression of ZO-1、Occludin,RAGE、p-NF-κBp65、NF-κBp65、P-IKBa and IKBa;RT-PCR was used to detect RAGE m RNA 、 NF-κB m RNA、IL-6 m RNA expression levels;ELISA was used to detect IL-1β and IL-6 expression in Caco-2 secretion,to explore the role of RAGE/NF-κB signaling pathway in intestinal mucositis.(2)Mild moxibustion on rats at Guan yuan(CV4),Tian shu(both sides,ST25)and Qi hai(CV6),a rat model of chemotherapeutic intestinal mucositis is established by intraperitoneal injection of 5-fluorouracil.General situation(such as body weight、food intake、occult blood test、stool characteristics,etc)of rats in each group is recorded;ELISA was used to detect IL-1β 、 IL-6 expression;HE staining and transmission electron microscopy experimental techniques were used to observe the protective effect of moxibustion on chemotherapy-induced intestinal mucositis,to explore the protective effect of moxibustion on chemotherapy-induced intestinal mucositis.(3)Western blot was used to detect the protein expression of R AGE,p-NF-κBp65,NF-κBp65 in ileal tissue;immunohistochemistry,i mmunofluorescence was used to detect RAGE expression in ileal tiss ue;RT-PCR was used to detect RAGE m RNA,NF-κB m RNA expres sion levels;ELISA was used to detect IL-1β、IL-6、RAGE expressio n in serum,to explore the effect of moxibustion on the RAGE/NF-κB signaling pathway of chemotherapy-induced intestinal mucositis.Result:(1)The results showed that,compared with the blank group,LPS stimulated Caco-2 cells had no effect on cell proliferation within 24 hours(p>0.05),when LPS stimulated Caco-2 cells for 48 hours,the LPS concentration was greater than or equal to 1 μg/ml,cells were significantly inhibited Proliferation(p<0.05);the expression of tight junction proteins ZO-1 and Occludin was significantly down-regulated(p<0.05),the content of IL-1β and IL-6 increased(p<0.05);compared with the LPS group,inhibition of the RAGE/NF-κB signaling pathway can reduce the expression of RAGE,p-NF-κBp65,and p-IKBa proteins(p<0.05),and the expression of IL-6 m RNA,RAGE m RNA and NF-κB m RNA decreased(p<0.05),the content of IL-1β and IL-6 in the cell supernatant decreased(p<0.05).(2)After a single intraperitoneal injection of 5-Fu,the weight and food intake of rats in each group decreased to varying degrees,and the intestinal mucosa showed different degrees of damage;the weight and food intake of the moxibustion group were higher than that of the chemotherapy group at the same time,and the DAI score was lower than that of the chemotherapy group at the same time,the expression of IL-1β and IL-6 in serum were significantly lower than those in the chemotherapy group(p<0.05).Histopathology showed that the mucosal damage was alleviated after moxibustion treatment.(3)After a single intraperitoneal injection of 5-Fu,the RAGE expression was significantly increased in ileum.Compared with the chemotherapy group,moxibustion down-regulated the RAGE and p-NF-κBp65 protein expression in rats with chemotherapy-induced intestinal mucositis(p<0.05),the expression levels of RAGE m RNA and NF-κB m RNA decreased(p<0.05),and the levels of RAGE、IL-1β and IL-6 in serum decreased(p<0.05).Conclusion:Inhibition of RAGE/NF-κB signaling pathway can reduce intestinal mucositis inflammation,Moxibustion pretreatment can improve 5-Fuinduced chemotherapy intestinal mucosal inflammation in rats,The mechanism may be through regulating RAGE/NF-κB signaling pathway. |
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