Background:Type 2 diabetes(T2D)has already become a global pandemic.As its simple,rapid,economical,and relatively non-invasive,metabolic markers have become a method for T2D diagnosis.However,region,race,and diet all ...Background:Type 2 diabetes(T2D)has already become a global pandemic.As its simple,rapid,economical,and relatively non-invasive,metabolic markers have become a method for T2D diagnosis.However,region,race,and diet all affect the metabolism of the body.The purpose of current study is to explore the differences of metabolites in T2D patients from regions.Methods:We recruited 103 T2D patients in two clinical centers,including 52 T2D patients from Beijing(T2D_(B))and 51 T2D patients from Kaifeng(T2D_(K)).The serum samples from T2D patients were analyzed using high-resolution mass spectrometer.After screened using univariate and multivariate analysis,the differential metabolites were identified.Moreover,to reveal biological information,we performed pathway analysis with the differential metabolites.Results:Thirty-six differential metabolites were identified,including 16 metabolites were higher concentrations while 20 metabolites were lower concentrations in the serum of T2D_(B) patients than T2D_(K) patients.There were higher serum concentrations of L-phenylalanine,4-methyl-2-oxovaleric acid,L-carnitine,decanoylcarnitine,9-decenoylcarnitine and sphinganine in T2D_(B) patients,in which decanoylcarnitine in T2D_(B) patients was up to 35-fold higher than T2D_(K) patients.While there were lower concentrations of L-valine,L-isoleucine,arachidonic acid,oleic acid,16-hydroxyhexadecanoic acid,lysophosphatidylcholine(18:0)and 1-Phenylethylamine in T2D_(B) patients,in which 1-phenylethylamine in T2D_(B) patients was decreased to 0.45-fold lower than T2D_(K) patients.The reason for the differences might be that phosphatidylethanolamine biosynthesis,phosphatidylcholine biosynthesis,valine,leucine and isoleucine degradation,and beta-oxidation of very long chain fatty acids were different in T2D_(B) patients and in T2D_(K) patients.Conclusion:Metabolites from different pathways are independently related to regions,providing valuable insight and potential for the diagnosis and treatment of T2D.展开更多
AIM： To investigate the transactivating effect of pre-S2 protein of hepatitis B virus （HBV） and construct a subtractive cDNA library of genes transactivated by pre-S2 protein with suppression subtractive hybridizat...AIM： To investigate the transactivating effect of pre-S2 protein of hepatitis B virus （HBV） and construct a subtractive cDNA library of genes transactivated by pre-S2 protein with suppression subtractive hybridization （SSH） technique, and to pave the way for elucidating the pathogenesis of HBV infection. METHODS： pcDNA3.1（-）-pre-S2 containing pre-S2 region of HBV genome was constructed by routine molecular methods. HepG2 cells were cotransfected with pcDNA3.1 （-）-pre-S21pSV-lacZ and empty pcDNA3.1（-）/pSV-lacZ. After 48 h, cells were collected and detected for the expression of β-galactosidase （β-gal）. SSH and bioinformatics techniques were used, the mRNA of HepG2 cells transfected with pcDNA3.1（-）-pre-S2 and pcDNA3.1（-） empty vector was isolated, respectively, cDNA was synthesized. After digestion with restriction enzyme RsaI, cDNA fragments were obtained. Tester cDNA was then divided into two groups and ligated to the specific adaptor 1 and adaptor 2, respectively. After tester cDNA was hybridized with driver cDNA twice and underwent two times of nested PCR, amplified cDNA fragments were subcloned into pGEM-Teasy vectors to set up the subtractive library. Amplification of the library was carried out with E.coli strain DH5α The cDNA was sequenced and analyzed in GenBank with Blast search after PCR. RESULTS： The pre-S2 mRNA could be detected in HepG2 cells transfected with pcDNA3.1（-）-pre-S2 plasmid. The activity of β-gal in HepG2 cells transfected with pcDNA3.1 （-）-pre-S2/pSV-lacZ was 7.0 times higher than that of control plasmid （P〈0.01）. The subtractive library of genes transactivated by HBV pre-S2 protein was constructed successfully. The amplified library contains 96 positiveclones. Colony PCR showed that 86 clones contained 200-1 000 bp inserts. Sequence analysis was performed in 50 clones randomly, and the full length sequences were obtained with bioinformatics method and searched for homologous DNA sequence from GenBank, altogether 25 coding sequences were obtained, these cDNA sequences might be the target genes transactivated by pre-S2 protein. CONCLUSION： The pre-S2 protein of HBV has transactivating effect on SV40 early promoter. The obtained sequences may be target genes transactivated by pre-S2 protein among which some genes coding proteins involved in cell cycle regulation, metabolism, immunity, signal transcluction and cell apoptosis.This finding brings some new clues for studying the biological functions of pre-S2 protein and further understanding of HBV hepatocarcinogesis.展开更多
基金supported by the National Natural Science Foundation of China(82130112)Capital’s Funds for Health Improvement and Research(CFH2024-2-1193)the Youth Beijing Scholar(2022-051).
文摘Background:Type 2 diabetes(T2D)has already become a global pandemic.As its simple,rapid,economical,and relatively non-invasive,metabolic markers have become a method for T2D diagnosis.However,region,race,and diet all affect the metabolism of the body.The purpose of current study is to explore the differences of metabolites in T2D patients from regions.Methods:We recruited 103 T2D patients in two clinical centers,including 52 T2D patients from Beijing(T2D_(B))and 51 T2D patients from Kaifeng(T2D_(K)).The serum samples from T2D patients were analyzed using high-resolution mass spectrometer.After screened using univariate and multivariate analysis,the differential metabolites were identified.Moreover,to reveal biological information,we performed pathway analysis with the differential metabolites.Results:Thirty-six differential metabolites were identified,including 16 metabolites were higher concentrations while 20 metabolites were lower concentrations in the serum of T2D_(B) patients than T2D_(K) patients.There were higher serum concentrations of L-phenylalanine,4-methyl-2-oxovaleric acid,L-carnitine,decanoylcarnitine,9-decenoylcarnitine and sphinganine in T2D_(B) patients,in which decanoylcarnitine in T2D_(B) patients was up to 35-fold higher than T2D_(K) patients.While there were lower concentrations of L-valine,L-isoleucine,arachidonic acid,oleic acid,16-hydroxyhexadecanoic acid,lysophosphatidylcholine(18:0)and 1-Phenylethylamine in T2D_(B) patients,in which 1-phenylethylamine in T2D_(B) patients was decreased to 0.45-fold lower than T2D_(K) patients.The reason for the differences might be that phosphatidylethanolamine biosynthesis,phosphatidylcholine biosynthesis,valine,leucine and isoleucine degradation,and beta-oxidation of very long chain fatty acids were different in T2D_(B) patients and in T2D_(K) patients.Conclusion:Metabolites from different pathways are independently related to regions,providing valuable insight and potential for the diagnosis and treatment of T2D.
文摘AIM： To investigate the transactivating effect of pre-S2 protein of hepatitis B virus （HBV） and construct a subtractive cDNA library of genes transactivated by pre-S2 protein with suppression subtractive hybridization （SSH） technique, and to pave the way for elucidating the pathogenesis of HBV infection. METHODS： pcDNA3.1（-）-pre-S2 containing pre-S2 region of HBV genome was constructed by routine molecular methods. HepG2 cells were cotransfected with pcDNA3.1 （-）-pre-S21pSV-lacZ and empty pcDNA3.1（-）/pSV-lacZ. After 48 h, cells were collected and detected for the expression of β-galactosidase （β-gal）. SSH and bioinformatics techniques were used, the mRNA of HepG2 cells transfected with pcDNA3.1（-）-pre-S2 and pcDNA3.1（-） empty vector was isolated, respectively, cDNA was synthesized. After digestion with restriction enzyme RsaI, cDNA fragments were obtained. Tester cDNA was then divided into two groups and ligated to the specific adaptor 1 and adaptor 2, respectively. After tester cDNA was hybridized with driver cDNA twice and underwent two times of nested PCR, amplified cDNA fragments were subcloned into pGEM-Teasy vectors to set up the subtractive library. Amplification of the library was carried out with E.coli strain DH5α The cDNA was sequenced and analyzed in GenBank with Blast search after PCR. RESULTS： The pre-S2 mRNA could be detected in HepG2 cells transfected with pcDNA3.1（-）-pre-S2 plasmid. The activity of β-gal in HepG2 cells transfected with pcDNA3.1 （-）-pre-S2/pSV-lacZ was 7.0 times higher than that of control plasmid （P〈0.01）. The subtractive library of genes transactivated by HBV pre-S2 protein was constructed successfully. The amplified library contains 96 positiveclones. Colony PCR showed that 86 clones contained 200-1 000 bp inserts. Sequence analysis was performed in 50 clones randomly, and the full length sequences were obtained with bioinformatics method and searched for homologous DNA sequence from GenBank, altogether 25 coding sequences were obtained, these cDNA sequences might be the target genes transactivated by pre-S2 protein. CONCLUSION： The pre-S2 protein of HBV has transactivating effect on SV40 early promoter. The obtained sequences may be target genes transactivated by pre-S2 protein among which some genes coding proteins involved in cell cycle regulation, metabolism, immunity, signal transcluction and cell apoptosis.This finding brings some new clues for studying the biological functions of pre-S2 protein and further understanding of HBV hepatocarcinogesis.
