高精料对奶山羊盲肠中SCFAs-GPR41/43介导的炎症反应的影响
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摘要
为了探讨高精料长期饲喂过程中,反刍动物肠道内的短链脂肪酸(SCFAs)在奶山羊盲肠中通过其特异性受体G蛋白偶联受体41/43(GPR41/43)对其下游的炎症信号通路是否有影响,选用12只萨能奶山羊随机分为2组,对照组(LC)饲喂低精料(精∶粗=4∶6,n=6),试验组(HC)饲喂高精料(精∶粗=6∶4,n=6)日粮,经10周饲喂试验,取盲肠组织样及盲肠内容物进行气相色谱短链脂肪酸含量、肠内容物pH、实时荧光定量(RT-q PCR)及蛋白印迹的检测。结果显示:相比于LC组,HC组乙酸(P<0.01)、丙酸(P<0.01)及丁酸(P<0.05)的含量极显著或显著升高,且HC组pH值明显降低(P<0.05);HC组GPR41/43、肿瘤坏死因子-α(TNF-α)及趋化因子20(CCL20)基因表达量相比于LC组极显著升高(P<0.01),白细胞介素-1β(IL-1β)、IL-6、IL-8、IL-10及CCL5等基因表达量显著升高(P<0.05);HC组GPR41/43蛋白表达量显著高于LC组(P<0.05)。提示:高精料饲喂后,反刍动物盲肠组织中通过短链脂肪酸活化GPR41/43进而激活下游炎症信号通路诱导组织内炎症反应。
To investigate if SCFAs that are mainly included in the gastrointestinal tract are the key molecules to activate their special receptors-G protein-coupled receptor 41/43 and to affect the downstream inflammatory signaling pathways in the cecum of lactating goats during long term feeding on high concentrate diet. 12 lactating goats were divided randomly into two groups,one group fed with low-concentrate diet(concentrate: roughage= 6 ∶ 4,n= 6) as control,and the other group fed with high-concentrate diet(concentrate: roughage = 6 ∶ 4,n =6) as treatment. After 10 weeks of feeding,cecum tissues and its intestinal contents were sampled and detected by gas chromatography,Western blot,RT-q PCR and pH meter to analyze the expression of genes,proteins and SCFAs concentration,and pH value. The results showed that,acetate(P< 0. 01),propionate(P < 0. 01) and butyrate(P < 0. 05) concentrations were increased significantly,and the pH value(P<0. 05) was decreased significantly in the HC group,compared with the LC group; the expressions of GPR41/43 genes and related inflammatory cytokine genes(TNF-α) and chemokine genes(CCL20)(P<0. 01) were increased significantly,and the expressions of IL-1β,IL-6,IL-8,IL-10 and CCL5(P<0. 05) were increased significantly in HC group; the expression of key proteins(GPR41/43) was also increased significantly in HC group. It is suggested that high concentrate diet might cause inflammatory response by SCFAs activated GPR41/43,which can activate the downstream inflammatory signaling pathways in the ruminant cecum.
To investigate if SCFAs that are mainly included in the gastrointestinal tract are the key molecules to activate their special receptors-G protein-coupled receptor 41/43 and to affect the downstream inflammatory signaling pathways in the cecum of lactating goats during long term feeding on high concentrate diet. 12 lactating goats were divided randomly into two groups,one group fed with low-concentrate diet(concentrate: roughage= 6 ∶ 4,n= 6) as control,and the other group fed with high-concentrate diet(concentrate: roughage = 6 ∶ 4,n =6) as treatment. After 10 weeks of feeding,cecum tissues and its intestinal contents were sampled and detected by gas chromatography,Western blot,RT-q PCR and pH meter to analyze the expression of genes,proteins and SCFAs concentration,and pH value. The results showed that,acetate(P< 0. 01),propionate(P < 0. 01) and butyrate(P < 0. 05) concentrations were increased significantly,and the pH value(P<0. 05) was decreased significantly in the HC group,compared with the LC group; the expressions of GPR41/43 genes and related inflammatory cytokine genes(TNF-α) and chemokine genes(CCL20)(P<0. 01) were increased significantly,and the expressions of IL-1β,IL-6,IL-8,IL-10 and CCL5(P<0. 05) were increased significantly in HC group; the expression of key proteins(GPR41/43) was also increased significantly in HC group. It is suggested that high concentrate diet might cause inflammatory response by SCFAs activated GPR41/43,which can activate the downstream inflammatory signaling pathways in the ruminant cecum.
引文
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[17]Sina C,Gavrilova O,Forster M,et al.G protein-coupled receptor43 is essential for neutrophil recruitment during intestinal inflammation[J].J Immunol,2009,183(11):7514-7522.
[18]Huttenhower C,Kostic A D,Xavier R J.Inflammatory bowel disease as a model for translating the microbiome[J].Immunity,2014,40(6):843-854.
[19]Boro P,Kumaresan A,Singh A K,et al.Expression of short chain fatty acid receptors and pro-inflammatory cytokines in uteroplacental tissues is altered in cows developing retention of fetal membranes[J].Placenta,2014,35(7):455-460.
[20]郭锦锦,孙万邦.IL-10受体及其信号转导研究进展[J].临床医学工程,2012(1):135-137.
[2]Hara T,Kashihara D,Ichimura A,et al.Role of free fatty acid receptors in the regulation of energy metabolism[J].Biochim Biophys Acta,2014,1841(9):1292-1300.
[3]赵怀宝,任玉龙.短链脂肪酸在动物体内的生理特点和功能[J].饲料研究,2016(3):29-32.
[4]Gibson G R,Roberfroid M B.Colonic microbiota,nutrition and health;colonic microbiota,nutrition and health[M].Softcover of orig.ed.UK:Springer,Dordrecht,1999:101-112.
[5]Sivaprakasam S,Prasad P D,Singh N.Benefits of short-chain fatty acids and their receptors in inflammation and carcinogenesis[J].Pharmacol Ther,2016,164:144-151.
[6]Goldsmith Z G,Dhanasekaran D N.G protein regulation of MAPK networks[J].Oncogene,2007,26(22):3122-3142.
[7]Milligan G,Bolognini D,Sergeev E.Ligands at the free fatty acid receptors 2/3(GPR43/GPR41)[J].Handb Exp Pharmacol,2017,236:17-32.
[8]王永安,张春雷,王艳红,等.短链脂肪酸受体GPR41、GPR43的信号通路及生理功能[J].中国牛业科学,2013(6):49-53.
[9]Miyamoto J,Hasegawa S,Kasubuchi M,et al.Nutritional signaling via free fatty acid receptors[J].Int J Mol Sci,2016,17(4):450.
[10]Voltolini C,Battersby S,Etherington S L,et al.A novel antiinflammatory role for the short-chain fatty acids in human labor[J].Endocrinology,2012,153(1):395-403.
[11]Alvarez-Curto E,Milligan G.Metabolism meets immunity:The role of free fatty acid receptors in the immune system[J].Biochem Pharmacol,2016,114:3-13.
[12]Kim M H,Kang S G,Park J H,et al.Short-chain fatty acids activate GPR41 and GPR43 on intestinal epithelial cells to promote inflammatory responses in mice[J].Gastroenterology,2013,145(2):396-406.
[13]Miyamoto J,Kasubuchi M,Nakajima A,et al.Anti-inflammatory and insulin-sensitizing effects of free fatty acid receptors[J].Handb Exp Pharmacol,2017,236:221-231.
[14]Le Poul E,Loison C,Struyf S,et al.Functional characterization of human receptors for short chain fatty acids and their role in polymorphonuclear cell activation[J].J Biol Chem,2003,278(28):25481-25489.
[15]Yonezawa T,Kobayashi Y,Obara Y.Short-chain fatty acids induce acute phosphorylation of the p38 mitogen-activated protein kinase/heat shock protein 27 pathway via GPR43 in the MCF-7 human breast cancer cell line[J].Cell Signal,2007,19(1):185-193.
[16]Huang G,Shi L Z,Chi H.Regulation of JNK and p38 MAPK in the immune system:signal integration,propagation and termination[J].Cytokine,2009,48(3):161-169.
[17]Sina C,Gavrilova O,Forster M,et al.G protein-coupled receptor43 is essential for neutrophil recruitment during intestinal inflammation[J].J Immunol,2009,183(11):7514-7522.
[18]Huttenhower C,Kostic A D,Xavier R J.Inflammatory bowel disease as a model for translating the microbiome[J].Immunity,2014,40(6):843-854.
[19]Boro P,Kumaresan A,Singh A K,et al.Expression of short chain fatty acid receptors and pro-inflammatory cytokines in uteroplacental tissues is altered in cows developing retention of fetal membranes[J].Placenta,2014,35(7):455-460.
[20]郭锦锦,孙万邦.IL-10受体及其信号转导研究进展[J].临床医学工程,2012(1):135-137.