目的 研究葡萄籽提取物（GSPE）與柳氮磺吡啶（SASP）聯(lián)合用藥治療葡聚糖硫酸鈉（DSS）誘導(dǎo)的小鼠潰瘍性結(jié)腸炎的效果及作用機制。方法 將SPF級C57小鼠隨機分為對照組、模型組、GSPE （250 mg/kg）組、SASP （250 mg/kg）組、聯(lián)合用藥（SASP 250 mg/kg+SASP 250 mg/kg）組，對照組給予正常飲用水，其他組均自由飲用3% DSS水溶液，連續(xù)飲用7 d，誘發(fā)小鼠潰瘍性結(jié)腸炎模型；造模第1天同步開始ig給藥，每天記錄小鼠體質(zhì)量、便血、便型等癥狀變化；7 d后斷頭取血，收集結(jié)腸和脾臟，記錄結(jié)腸長度、脾臟質(zhì)量變化情況。HE染色評估小鼠結(jié)腸黏膜組織病理變化，ELISA法檢測血清和結(jié)腸組織中炎癥因子腫瘤壞死因子（TNF-α）、白細胞介素（IL）-1β、IL-6和NO的表達變化以及丙二醛（MDA）、超氧化物歧化酶（SOD）細胞因子的含量變化，免疫組化分析結(jié)腸組織上皮細胞中NF-κB-p65、Nrf2、HO-1和Keap-1含量的變化。應(yīng)用金氏Q值法分析聯(lián)合用藥的作用效果。結(jié)果 與模型組比較，各給藥組的小鼠體質(zhì)量下降幅度顯著降低（P<0.01），小鼠腹瀉、便血癥狀改善明顯，其中聯(lián)合用藥組小鼠體質(zhì)量較單獨給藥組下降更緩，小鼠腹瀉、便血程度改善更明顯（P<0.05）。各給藥組小鼠血清及結(jié)腸組織中IL-1β、IL-6、TNF-α、NO、MDA含量較模型組顯著降低，SOD含量則顯著升高（P<0.01）；病理組織切片分析發(fā)現(xiàn)，各給藥組小鼠結(jié)腸黏膜病理損傷較模型組顯著減輕，其中聯(lián)合用藥組小鼠結(jié)腸黏膜病理損傷較單獨給藥組降低更為顯著（P<0.05）。免疫組化分析結(jié)果也顯示，各給藥組小鼠的NF-κB-p65、Keap-1蛋白表達與模型組比較顯著下調(diào)（P<0.01），Nrf2、HO-1蛋白表達顯著上調(diào)（P<0.01）。聯(lián)合給藥組與單獨給藥組比較，NF-κB-p65、Keap-1蛋白表達顯著降低（P<0.05），Nrf2、HO-1蛋白表達顯著升高（P<0.05）。金氏Q值法評價兩藥合用后對IL-1β、IL-6、TNF-α、NO、MDA、NF-κB-p65、Keap-1的作用效果均大于1.15，說明GSPE與SASP具有協(xié)同作用。結(jié)論 GSPE與SASP聯(lián)合用藥治療實驗性潰瘍性結(jié)腸炎效果優(yōu)于單獨給藥，聯(lián)合用藥增強GSPE和SASP的抗氧化和抗炎作用，GSPE與SASP配伍治療潰瘍性結(jié)腸炎可進一步發(fā)揮協(xié)同增效的作用。

Objective To study the effect and mechanism of grape seed extract combined with sulfasalazine on DSS-induced ulcerative colitis (UC) in mice. Methods SPF grade C57 mice were randomly divided into five groups:control group, model group, grape seed extract group (GSPE, 250 mg/kg), sulfasalazine group (SASP, 250 mg/kg) and the combination medicine group[GSPE (250 mg/kg) + SASP (250 mg/kg)]. The control group was given normal drinking water, and the other groups were free to drink 3% DSS aqueous solution for 7 consecutive days to induce a model of UC in mice. The symptoms such as body weight, blood in the stool, and stool type were recorded every day. All mice were sacrificed after drug intervention for 7 consecutive days. Blood, colons and spleens were collected, respectively. The changes in colon length and spleen weight were recorded. HE staining was used to evaluate the pathological changes of mouse colonic mucosa. ELISA was used to detect the expression of inflammatory factors tumor necrosis factor:TNF-α, interleukin 1β (IL-1β), interleukin 6 (IL-6) and NO in serum and colon tissues, as well as the changes of MDA, SOD cytokines. Immunohistochemical analysis of the changes of content of NF-κB-p65, Nrf2, HO-1, Keap-1 were carried out in colon tissue epithelial cells. Effect of combined drug was evaluated by the Jin's Q-value method. Results Compared with model group, the body weight of mice in the GSPE group, SASP group and combination medicine group decreased slowly, and the symptoms of diarrhea and blood in the stool were significantly improved. The weight loss of the combination medicine group was slower than that of GSPE group and SASP group. Meanwhile, compared with the single administration groups, the diarrhea and bloody stools of the combination medicine group improved significantly. After administration, the contents of IL-1β, IL-6, TNF-α, NO, and MDA in the serum and colon tissues of mice were significantly lower than that in the model group, and the SOD content was significantly increased (P<0.01); pathological tissue analysis revealed that the pathological damage of the colonic mucosa of the mice in each administration group was significantly reduced, and the pathological damage of the colonic mucosa of the mice in the combination medicine extract was significantly lower than that of the other two groups (P<0.05). Immunohistochemical analysis showed that the expression of NF-κB and Keap-1 protein in each administration group was significantly reduced (P<0.01), and the expression of Nrf2 and HO-1 protein was significantly increased (P<0.05). Among them, the expression of NF-κB-p65 and Keap-1 protein in the combination medicine group was significantly lower than that in SASP group and GSPE group (P<0.05). The expression of Nrf2 and HO-1 protein in the combination medicine group were significantly higher than that in single administration groups (P<0.05). The Jin's Q-values of the effect of combined drug on IL-1β, IL-6, TNF-α, NO, MDA, NF-κB-p65, and Keap-1 were all greater than 1.15, which indicated that there is quite obviously synergy between GSPE and SASP. Conclusion Compared with single administration, the combined use of SASP and GSPE will enhance the antioxidant and anti-inflammatory effects of GSPE and SASP. The combined use of the two drugs has a synergistic effect on the treatment of UC.

R285.5

國家自然科學基金資助項目（81673693）；武警后勤學院創(chuàng)新團隊基金項目（2019）；武警后勤學院博士啟動金（WHB201710）；武警后勤學院基礎(chǔ)研究項目（WHJ201905）