腸道菌群介導潰瘍性結腸炎大鼠腸CYP3A和P-糖蛋白的變化及機制
腸道菌群是一個非常龐大、複雜、多樣、持續動態變化的微生態系統。它與宿主通過底物的代謝交換與共代謝等方式相互作用, 深刻影響宿主代謝表型, 決定宿主的營養、健康和疾病狀態。本文作者構建潰瘍性結腸炎模型, 研究了腸道菌群介導潰瘍性結腸炎大鼠腸CYP3A和P-糖蛋白的變化及機制。
腸道細胞色素P450 3A (cytochrome P450 3A, CYP3A) 和P-糖蛋白 (P-glycoprotein, P-gp) 是腸屏障的重要組成部分。炎症性腸病中腸CYP3A和P-gp下調伴隨腸道菌群紊亂。但兩者是否關聯？機制如何？尚不清楚。本研究採用5% 葡聚糖硫酸鈉誘導大鼠潰瘍性結腸炎 (ulcerative colitis, UC), 並對正常動物分別灌胃正常及UC動物糞便, 發現糞便移植改變了受體動物腸道菌組成, 而移植UC糞便組腸CYP3A2和P-gp mRNA的表達顯著下調。外膜囊泡 (outer-membrane vesicles, OMVs) 是革蘭陰性菌產生、進行群體行為及與環境通信的關鍵結構。不同處理組的OMVs均能下調人結腸腺癌細胞Caco-2中CYP3A4和P-gp的mRNA表達, 而UC組以及UC糞便處理組的OMVs的抑制作用更強, 且相對分子質量3～5萬的OMVs組分的作用更顯著。細胞經toll樣受體4 (toll like receptor 4, TLR4) 抑製劑瑞沙托維處理或轉染TLR4 siRNA能夠阻斷OMVs對CYP3A4和P-gp的下調。本研究證實, UC腸道菌部分通過分泌OMVs活化TLR4受體通路下調腸道CYP3A和P-gp表達。
1、DSS誘導及糞便移植對大鼠糞便菌組成的影響
Figure 1 Alterations of total and six main bacteria in fecal samples from normal and ulcerative colitis (UC) rats, and rats receiving normal or UC rat feces. mRNA levels were measured by QPCR. Animals received water (Nor), 5% DSS (UC), feces from normal (NN) or ulcerative colitis rats (NU), respectively, for 7 days. Data were expressed as mean ± SD of triplicate determinations. The relative level of each bacterium at each time point was obtained by comparing with the level of the same group on day 0
2、DSS誘導及糞菌移植對大鼠不同腸段CYP3A2及P-gp表達的影響
Figure 2 mRNA expression of CYP3A2 and P-gp in different intestinal segments of rats. mRNA expression was measured by QPCR. Data were expressed as mean ± SD (n = 6). *P < 0.05 vs Nor; #P < 0.05 vs NN
3、腸道菌OMVs對Caco-2細胞中CYP3A4和P-gp表達的影響
Figure 3 Effects of rat OMVs on mRNA expression of CYP3A4 and P-gp in Caco-2 cells. Cells were treated by OMVs from normal (NOMVs) and UC (UCOMVs) rats, and rats receiving normal (NNOMVs) or UC (NUOMVs) rat feces, respectively. mRNA expression was measured by QPCR. Data were expressed as mean ± SD of triplicate determination of one representative experiment. *P < 0.05 vs control; #P < 0.05 vs NOMVs or NNOMVs
4、不同OMVs組分對Caco-2細胞中CYP3A4和P-gp表達的影響
Figure 4 Effects of OMVs different fractions on mRNA expression of CYP3A4 and P-gp in Caco-2 cells. Cells were treated with or without different OMVs fractions from normal or UC rats. mRNA expression was measured by QPCR. Data were expressed as mean ± SD of triplicate determination of one representative experiment. *P < 0.05 vs control; #P < 0.05 vs NOMVs
5、siRNA對Caco-2細胞中TLR4和GAPDH表達的影響
Figure 5 Effects of TLR4 siRNA transfection on TLR4 mRNA expression in Caco-2 cells. Cells were transfected with TLR4 siRNA for 48 h. Cells treated with negative siRNA (NC) or GAPDH siRNA (PC) were processed in parallel to serve as controls. mRNA expression was determined by QPCR. Data was expressed as mean ± SD of triplicate determination of one representative experiment. *P < 0.05 vs control, #P < 0.05 vs NC. TLR4: Toll like receptor 4; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase
6、抑制或沉默TLR4受體對Caco-2細胞中CYP3A4和P-gp表達的影響
Figure 6 Effects of TLR4 inhibitor and siRNA transfection on mRNA expression of CYP3A4 and P-gp in Caco-2 cells. Cells were treated with TLR4 inhibitor (A), transfected with TLR4 or negative siRNA (NC) (B) followed by treatment of OMVs from normal or UC rats. mRNA expression was determined by QPCR. Data were expressed as mean ± SD of triplicate determination of one representative experiment. *P < 0.05 vs control; #P < 0.05 vs NOMVs or UCOMVs. TAK-242: Resatorvid
參考文獻：高雪姣, 李婷, 魏斌, 嚴志祥, 燕茹. 腸道菌群介導潰瘍性結腸炎大鼠腸CYP3A和P-糖蛋白的變化及機制 [J]. 藥學學報, 2017, 52(1): 34-43.
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