TNF recepter-55 is the main mediator of TNF-induced apoptosis. TNF receptor-75-dependent induction or enhancement of cytotoxicity has been explained by intracellular signaling, "ligand passing" , or inductio...TNF recepter-55 is the main mediator of TNF-induced apoptosis. TNF receptor-75-dependent induction or enhancement of cytotoxicity has been explained by intracellular signaling, "ligand passing" , or induction of endogenous TNF. To study the function of human TNF receptor-75 (hTR75) and the interaction between human TNF receptor-55 (hTR55) and hTR75 in hTNFa-induced cytotoxicity, HEp-2 cells were transfected with bicistronic expression vector of hTR75 and its deletion mutants genes. hTNFa-induced cytotoxicity was determined by crystal violet colorimetric method. The expression of hTR75 and its deletion mutants in HEp-2 cells was demonstrated by RT-PCR and indirect ELISA. We found that the overexpressed hTR75 could significantly increase the susceptibility of HEp-2 cells to hTNFa which especially required TRAF2 binding site. hTR75 could not only mediate partial hTNFa-induced cytotoxicity independently but also fulfill an accessory role in enhancing or synergizing hTR55-mediated展开更多
Human tumor necrosis factor α (hTNFα), a pleiotropic cytokine withactivities ranging from host defense mechanisms in infection and injury to severe toxicity in septicshock or other related diseases, is a promising t...Human tumor necrosis factor α (hTNFα), a pleiotropic cytokine withactivities ranging from host defense mechanisms in infection and injury to severe toxicity in septicshock or other related diseases, is a promising target for drug screening. Using the SELEX(systematic evolution of ligands by exponential enrichment) process, we isolated oligonucleotideligands (aptamers) with high affinities for hTNFα. Aptamers were selected from a starting pool of40 randomized sequences composed of about 10^(15) RNA molecules. Representative aptamers weretruncated to the minimal length with high affinity for hTNFα and were further modified byreplacement of 2'-OH with 2'-F and 2'-NH_2 at all ribopurine positions. These modified RNA aptamerswere resistant to nuclease. The specificity of these aptamers for hTNFα was confirmed, and theiractivity to inhibit the cytotoxicity of hTNFα on mouse L929 cells was determined. Resultsdemonstrated that four 2'-NH_2-modified aptamers bound to hTNFα with high affinity and blocked thebinding of hTNFα to its receptor, thus protecting the L929 cells from the cytotoxicity of hTNFα.Oligonucleotide aptamers described here are potential therapeutics and diagnostics forhTNFα-related diseases.展开更多
文摘目的探讨软骨细胞在肿瘤坏死因子(TNFα)作用下对其分泌基质蛋白及相关基因mRNA表达的影响。方法体外培养的小鼠软骨细胞用25,50,100 ng/ml的h TNFα干预,不加入TNFα作为对照,通过RT-q PCR检测h TNFα对软骨细胞分泌基质蛋白及相关基因mRNA表达的影响。结果 h TNFα能降低软骨细胞分泌的Ⅱ型胶原及刺激促Ⅱ型胶原分泌的SOX9基因的表达;对基质可提高基质酶MMP-9,MMP-13,ADAMTS-4和ADAMTS-5基因的表达,提高炎性因子IL-1α,TGFβ,并增加凋亡基因BAX,caspase3的表达。结论 h TNFα通过不同途径影响软骨细胞及基质相关基因的表达,从而促进了关节软骨的降解。
文摘TNF recepter-55 is the main mediator of TNF-induced apoptosis. TNF receptor-75-dependent induction or enhancement of cytotoxicity has been explained by intracellular signaling, "ligand passing" , or induction of endogenous TNF. To study the function of human TNF receptor-75 (hTR75) and the interaction between human TNF receptor-55 (hTR55) and hTR75 in hTNFa-induced cytotoxicity, HEp-2 cells were transfected with bicistronic expression vector of hTR75 and its deletion mutants genes. hTNFa-induced cytotoxicity was determined by crystal violet colorimetric method. The expression of hTR75 and its deletion mutants in HEp-2 cells was demonstrated by RT-PCR and indirect ELISA. We found that the overexpressed hTR75 could significantly increase the susceptibility of HEp-2 cells to hTNFa which especially required TRAF2 binding site. hTR75 could not only mediate partial hTNFa-induced cytotoxicity independently but also fulfill an accessory role in enhancing or synergizing hTR55-mediated
文摘Human tumor necrosis factor α (hTNFα), a pleiotropic cytokine withactivities ranging from host defense mechanisms in infection and injury to severe toxicity in septicshock or other related diseases, is a promising target for drug screening. Using the SELEX(systematic evolution of ligands by exponential enrichment) process, we isolated oligonucleotideligands (aptamers) with high affinities for hTNFα. Aptamers were selected from a starting pool of40 randomized sequences composed of about 10^(15) RNA molecules. Representative aptamers weretruncated to the minimal length with high affinity for hTNFα and were further modified byreplacement of 2'-OH with 2'-F and 2'-NH_2 at all ribopurine positions. These modified RNA aptamerswere resistant to nuclease. The specificity of these aptamers for hTNFα was confirmed, and theiractivity to inhibit the cytotoxicity of hTNFα on mouse L929 cells was determined. Resultsdemonstrated that four 2'-NH_2-modified aptamers bound to hTNFα with high affinity and blocked thebinding of hTNFα to its receptor, thus protecting the L929 cells from the cytotoxicity of hTNFα.Oligonucleotide aptamers described here are potential therapeutics and diagnostics forhTNFα-related diseases.
