BACKGROUND Colorectal cancer(CRC)is a worldwide problem,which has been associated with changes in diet and lifestyle pattern.As a result of colonic fermentation of dietary fibres,short chain free fatty acids are gener...BACKGROUND Colorectal cancer(CRC)is a worldwide problem,which has been associated with changes in diet and lifestyle pattern.As a result of colonic fermentation of dietary fibres,short chain free fatty acids are generated which activate free fatty acid receptors(FFAR)2 and 3.FFAR2 and FFAR3 genes are abundantly expressed in colonic epithelium and play an important role in the metabolic homeostasis of colonic epithelial cells.Earlier studies point to the involvement of FFAR2 in colorectal carcinogenesis.AIM To understand the role of short chain FFARs in CRC.METHODS Transcriptome analysis console software was used to analyse microarray data from CRC patients and cell lines.We employed short-hairpin RNA mediated down regulation of FFAR2 and FFAR3 genes,which was validated using quantitative real time polymerase chain reaction.Assays for glucose uptake and cyclic adenosine monophosphate(cAMP)generation was done along with immunofluorescence studies to study the effects of FFAR2/FFAR3 knockdown.For measuring cell proliferation,we employed real time electrical impedancebased assay available from xCELLigence.RESULTS Microarray data analysis of CRC patient samples showed a significant down regulation of FFAR2 gene expression.This prompted us to study the FFAR2 in CRC.Since,FFAR3 shares significant structural and functional homology with FFAR2,we knocked down both these receptors in CRC cell line HCT 116.These modified cell lines exhibited higher proliferation rate and were found to have increased glucose uptake as well as increased level of glucose transporter 1.Since,FFAR2 and FFAR3 signal through G protein subunit(Gαi),knockdown of these receptors was associated with increased cAMP.Inhibition of protein kinase A(PKA)did not alter the growth and proliferation of these cells indicating a mechanism independent of cAMP/PKA pathway.CONCLUSION Our results suggest role of FFAR2/FFAR3 genes in increased proliferation of colon cancer cells via enhanced glucose uptake and exclude the role of PKA mediated cAMP signalling.Alternate pathways could be involved that would ultimately result in increased cell proliferation as a result of down regulated FFAR2/FFAR3 genes.This study paves the way to understand the mechanism of action of short chain FFARs in CRC.展开更多
Free fatty acids are essential dietary components and recognized as important molecules in the maintenance of cellular homeostasis.In the last decade,the molecular pathways for free fatty acid sensing in the gastroint...Free fatty acids are essential dietary components and recognized as important molecules in the maintenance of cellular homeostasis.In the last decade,the molecular pathways for free fatty acid sensing in the gastrointestinal tract have been further elucidated by molecular identification and functional characterization of fatty acid binding receptors.These sensing molecules belong to the family of G proteincoupled receptors.In the intestine,four important receptors have been described so far.They differ in molecular structure,ligand specificity,expression pattern,and functional properties.In this review,an overview of intestinal fatty acid binding receptors and their role in intestinal physiology and pathophysiology is given.展开更多
Inflammatory bowel diseases(IBDs) are characterized by inflammation in the gastrointestinal tract and include Ulcerative Colitis and Crohn’s Disease.These diseases are costly to health services,substantially reduce p...Inflammatory bowel diseases(IBDs) are characterized by inflammation in the gastrointestinal tract and include Ulcerative Colitis and Crohn’s Disease.These diseases are costly to health services,substantially reduce patients’ quality of life,and can lead to complications such as cancer and even death.Symptoms include abdominal pain,stool bleeding,diarrhea,and weight loss.The treatment of these diseases is symptomatic,seeking disease remission.The intestine is colonized by several microorganisms,such as fungi,viruses,and bacteria,which constitute the intestinal microbiota(IM).IM bacteria promotes dietary fibers fermentation and produces short-chain fatty acids(SCFAs) that exert several beneficial effects on intestinal health.SCFAs can bind to G protein-coupled receptors,such as GPR41 and GPR43,promoting improvements in the intestinal barrier,anti-inflammatory,and antioxidant effects.Thus,SCFAs could be a therapeutic tool for IBDs.However,the mechanisms involved in these beneficial effects of SCFAs remain poorly understood.Therefore,this paper aims to provide a review addressing the main aspects of IBDs,and a more detailed sight of SCFAs,focusing on the main effects on different aspects of the intestine with an emphasis on IBDs.展开更多
目的:观察长链游离脂肪酸对体外培养的角质形成细胞的作用,探讨其对TLR及NF-ΚB信号转导通路的影响。方法:①体外培养Hacat细胞,实验分为N、PA、SA、LPS四组,N组为正常对照组,软脂酸(Palmitic Acid,PA)750μmol/L、硬脂酸(Stearic Acid,...目的:观察长链游离脂肪酸对体外培养的角质形成细胞的作用,探讨其对TLR及NF-ΚB信号转导通路的影响。方法:①体外培养Hacat细胞,实验分为N、PA、SA、LPS四组,N组为正常对照组,软脂酸(Palmitic Acid,PA)750μmol/L、硬脂酸(Stearic Acid,SA)750μmol/L、脂多糖(lipopolysaccharide,LPS)500 ng/L和Hacat细胞共培养72h组分别为N组、PA组、SA组、LPS组;②LPS、SA与角质形成细胞共培养72h检测LPS、SA对Hacat细胞NF-κB、p-IκBα、IκBα表达的影响。通过免疫印迹法(Weston blot)检测SA、LPS对Hacat细胞TLR2、TLR4、NF-ΚB、p-IκBα、IκBα表达的影响;Real Ti me PCR方法检测对Hacat细胞内NF-κB、TNF-αmRNA表达的影响。结果:SA、LPS激活细胞表面TLR后可以明显促进细胞表面TLR2、TLR4表达及细胞内NF-κB、TNF-α等促炎症因子的表达。PA促进Hacat细胞内NF-ΚB、p-IκBα、IκBα表达无明显影响(P>0.05)结论:SA可促进角质形成细胞分泌NF-κB、TNF-α等因子表达促进炎症反应;其机制可能为:SA可能通过TLR及NF-κB信号传导通路促进细胞释放NF-κB、TNF-α等促炎因子。展开更多
文摘BACKGROUND Colorectal cancer(CRC)is a worldwide problem,which has been associated with changes in diet and lifestyle pattern.As a result of colonic fermentation of dietary fibres,short chain free fatty acids are generated which activate free fatty acid receptors(FFAR)2 and 3.FFAR2 and FFAR3 genes are abundantly expressed in colonic epithelium and play an important role in the metabolic homeostasis of colonic epithelial cells.Earlier studies point to the involvement of FFAR2 in colorectal carcinogenesis.AIM To understand the role of short chain FFARs in CRC.METHODS Transcriptome analysis console software was used to analyse microarray data from CRC patients and cell lines.We employed short-hairpin RNA mediated down regulation of FFAR2 and FFAR3 genes,which was validated using quantitative real time polymerase chain reaction.Assays for glucose uptake and cyclic adenosine monophosphate(cAMP)generation was done along with immunofluorescence studies to study the effects of FFAR2/FFAR3 knockdown.For measuring cell proliferation,we employed real time electrical impedancebased assay available from xCELLigence.RESULTS Microarray data analysis of CRC patient samples showed a significant down regulation of FFAR2 gene expression.This prompted us to study the FFAR2 in CRC.Since,FFAR3 shares significant structural and functional homology with FFAR2,we knocked down both these receptors in CRC cell line HCT 116.These modified cell lines exhibited higher proliferation rate and were found to have increased glucose uptake as well as increased level of glucose transporter 1.Since,FFAR2 and FFAR3 signal through G protein subunit(Gαi),knockdown of these receptors was associated with increased cAMP.Inhibition of protein kinase A(PKA)did not alter the growth and proliferation of these cells indicating a mechanism independent of cAMP/PKA pathway.CONCLUSION Our results suggest role of FFAR2/FFAR3 genes in increased proliferation of colon cancer cells via enhanced glucose uptake and exclude the role of PKA mediated cAMP signalling.Alternate pathways could be involved that would ultimately result in increased cell proliferation as a result of down regulated FFAR2/FFAR3 genes.This study paves the way to understand the mechanism of action of short chain FFARs in CRC.
基金Supported by Deutsche Forschungs gemeinschaft (DFG GA 785/5-1) and Deutsche Krebshilfe (DKH 109313)
文摘Free fatty acids are essential dietary components and recognized as important molecules in the maintenance of cellular homeostasis.In the last decade,the molecular pathways for free fatty acid sensing in the gastrointestinal tract have been further elucidated by molecular identification and functional characterization of fatty acid binding receptors.These sensing molecules belong to the family of G proteincoupled receptors.In the intestine,four important receptors have been described so far.They differ in molecular structure,ligand specificity,expression pattern,and functional properties.In this review,an overview of intestinal fatty acid binding receptors and their role in intestinal physiology and pathophysiology is given.
基金Supported by the Research Support Foundation of the State of S?o PauloFAPESP+1 种基金Brazil,No.n2014/25927-2 and No.2018/07862-1Higher Education Personnel Improvement Coordination (CAPES,Brazil),No.n88887.506345/2020-00。
文摘Inflammatory bowel diseases(IBDs) are characterized by inflammation in the gastrointestinal tract and include Ulcerative Colitis and Crohn’s Disease.These diseases are costly to health services,substantially reduce patients’ quality of life,and can lead to complications such as cancer and even death.Symptoms include abdominal pain,stool bleeding,diarrhea,and weight loss.The treatment of these diseases is symptomatic,seeking disease remission.The intestine is colonized by several microorganisms,such as fungi,viruses,and bacteria,which constitute the intestinal microbiota(IM).IM bacteria promotes dietary fibers fermentation and produces short-chain fatty acids(SCFAs) that exert several beneficial effects on intestinal health.SCFAs can bind to G protein-coupled receptors,such as GPR41 and GPR43,promoting improvements in the intestinal barrier,anti-inflammatory,and antioxidant effects.Thus,SCFAs could be a therapeutic tool for IBDs.However,the mechanisms involved in these beneficial effects of SCFAs remain poorly understood.Therefore,this paper aims to provide a review addressing the main aspects of IBDs,and a more detailed sight of SCFAs,focusing on the main effects on different aspects of the intestine with an emphasis on IBDs.
文摘目的:观察长链游离脂肪酸对体外培养的角质形成细胞的作用,探讨其对TLR及NF-ΚB信号转导通路的影响。方法:①体外培养Hacat细胞,实验分为N、PA、SA、LPS四组,N组为正常对照组,软脂酸(Palmitic Acid,PA)750μmol/L、硬脂酸(Stearic Acid,SA)750μmol/L、脂多糖(lipopolysaccharide,LPS)500 ng/L和Hacat细胞共培养72h组分别为N组、PA组、SA组、LPS组;②LPS、SA与角质形成细胞共培养72h检测LPS、SA对Hacat细胞NF-κB、p-IκBα、IκBα表达的影响。通过免疫印迹法(Weston blot)检测SA、LPS对Hacat细胞TLR2、TLR4、NF-ΚB、p-IκBα、IκBα表达的影响;Real Ti me PCR方法检测对Hacat细胞内NF-κB、TNF-αmRNA表达的影响。结果:SA、LPS激活细胞表面TLR后可以明显促进细胞表面TLR2、TLR4表达及细胞内NF-κB、TNF-α等促炎症因子的表达。PA促进Hacat细胞内NF-ΚB、p-IκBα、IκBα表达无明显影响(P>0.05)结论:SA可促进角质形成细胞分泌NF-κB、TNF-α等因子表达促进炎症反应;其机制可能为:SA可能通过TLR及NF-κB信号传导通路促进细胞释放NF-κB、TNF-α等促炎因子。
