Objective:To isolate and characterize RNA aptamers that are specific to human CD36 protein using systematic evolution of ligands by exponential enrichment(SELEX)technology to identify candidates for adjunct therapy to...Objective:To isolate and characterize RNA aptamers that are specific to human CD36 protein using systematic evolution of ligands by exponential enrichment(SELEX)technology to identify candidates for adjunct therapy to reverse the binding of Plasmodiuminfected erythrocytes.Methods:RNA aptamers were isolated using nitrocellulose membrane-based SELEX and binding analysis was screened using an electrophoretic mobility shift assay and enzyme-linked oligonucleotide assay.Results:Thirteen cycles of nitrocellulose membrane-based SELEX yielded three aptamers(RC60,RC25,RC04)exhibiting high binding against CD36 protein as shown on electrophoretic mobility shift assay.The sequence analysis revealed a G-quadruplex sequence within all the isolated aptamers that might contribute to aptamer binding and thermodynamic stability.The specificity assay further showed that RC60 and RC25 were highly specific to CD36.The competitive inhibition assay demonstrated that RC60 and RC25 shared a similar binding epitope recognized by m Ab FA6-152,a specific monoclonal antibody against CD36.Conclusions:RC60 and RC25 are promising candidates as anticytoadherence for severe malaria adjunct therapy.展开更多
目的:观察人参皂苷Rd对氧化低密度脂蛋白(OX-LDL)诱导肝星状细胞(HSCs)活化与分泌胶原作用的影响及其机制。方法:10μg/ml OX-LDL孵育诱导肝星状细胞活化建立体外肝纤维化模型,分别给予人参皂苷Rd(18.8mg/L或37.6mg/L)处理肝星状细胞48...目的:观察人参皂苷Rd对氧化低密度脂蛋白(OX-LDL)诱导肝星状细胞(HSCs)活化与分泌胶原作用的影响及其机制。方法:10μg/ml OX-LDL孵育诱导肝星状细胞活化建立体外肝纤维化模型,分别给予人参皂苷Rd(18.8mg/L或37.6mg/L)处理肝星状细胞48小时。MTT法检测肝星状细胞增殖,荧光定量PCR检测转化生长因子(TGF)-β1和CD36 m RNA表达;Western blot检测I型胶原及CD36、Smad2、Smad4表达。结果:与正常对照组相比,模型处理组的I型胶原蛋白表达与TGF-β1 m RNA表达均明显增加;与模型处理组相比,人参皂苷Rd(38.7mg/L)明显抑制肝星状细胞增殖,降低CD36、I型胶原和Smad4蛋白表达,同时,人参皂苷Rd对Smad2蛋白表达没有影响。结论:人参皂苷Rd可能通过降低CD36及Smad4蛋白表达减少细胞外胶原蛋白的合成,进而发挥抗纤维化作用。展开更多
基金supported by an Exploratory Research Grant Scheme(ERGS203/CIPPM/6730106)+2 种基金Higher Institution Centre of Excellent(HICo E311/CIPPM/4401005)from the Malaysian,Ministry of Higher EducationUniversiti Sains Malaysia Fellowship Scheme.
文摘Objective:To isolate and characterize RNA aptamers that are specific to human CD36 protein using systematic evolution of ligands by exponential enrichment(SELEX)technology to identify candidates for adjunct therapy to reverse the binding of Plasmodiuminfected erythrocytes.Methods:RNA aptamers were isolated using nitrocellulose membrane-based SELEX and binding analysis was screened using an electrophoretic mobility shift assay and enzyme-linked oligonucleotide assay.Results:Thirteen cycles of nitrocellulose membrane-based SELEX yielded three aptamers(RC60,RC25,RC04)exhibiting high binding against CD36 protein as shown on electrophoretic mobility shift assay.The sequence analysis revealed a G-quadruplex sequence within all the isolated aptamers that might contribute to aptamer binding and thermodynamic stability.The specificity assay further showed that RC60 and RC25 were highly specific to CD36.The competitive inhibition assay demonstrated that RC60 and RC25 shared a similar binding epitope recognized by m Ab FA6-152,a specific monoclonal antibody against CD36.Conclusions:RC60 and RC25 are promising candidates as anticytoadherence for severe malaria adjunct therapy.
文摘目的:观察人参皂苷Rd对氧化低密度脂蛋白(OX-LDL)诱导肝星状细胞(HSCs)活化与分泌胶原作用的影响及其机制。方法:10μg/ml OX-LDL孵育诱导肝星状细胞活化建立体外肝纤维化模型,分别给予人参皂苷Rd(18.8mg/L或37.6mg/L)处理肝星状细胞48小时。MTT法检测肝星状细胞增殖,荧光定量PCR检测转化生长因子(TGF)-β1和CD36 m RNA表达;Western blot检测I型胶原及CD36、Smad2、Smad4表达。结果:与正常对照组相比,模型处理组的I型胶原蛋白表达与TGF-β1 m RNA表达均明显增加;与模型处理组相比,人参皂苷Rd(38.7mg/L)明显抑制肝星状细胞增殖,降低CD36、I型胶原和Smad4蛋白表达,同时,人参皂苷Rd对Smad2蛋白表达没有影响。结论:人参皂苷Rd可能通过降低CD36及Smad4蛋白表达减少细胞外胶原蛋白的合成,进而发挥抗纤维化作用。
