目的 :探讨Klf10沉默对人牙周膜细胞(human periodontal ligament cells,HPDLCs)受力后骨向分化的影响。方法:分离培养HPDLCs,RNA干扰沉默Klf10的表达,用离心加力法对细胞加载机械力,并予hedgehog信号通路特异性激动剂purmorphamine进...目的 :探讨Klf10沉默对人牙周膜细胞(human periodontal ligament cells,HPDLCs)受力后骨向分化的影响。方法:分离培养HPDLCs,RNA干扰沉默Klf10的表达,用离心加力法对细胞加载机械力,并予hedgehog信号通路特异性激动剂purmorphamine进行干预。采用ELISA法检测碱性磷酸酶(alkaline phosphatase,ALP)活性,采用RT-PCR和Western印迹法分别检测各组细胞Klf10 m RNA和蛋白表达水平,并检测成骨标志基因Runt相关转录因子2(runt-related transcription factor 2,Runx2)、骨桥蛋白(osteopontin,OPN)和骨钙素(osteocalcin,OCN)m RNA和蛋白的表达情况;利用Western免疫印迹法检测hedgehog信号转导通路成员胶质瘤相关癌基因同源物1(glioma-associated oncogene homolog 1,GLI1)和Ptch1(patched-1)的蛋白表达。采用SPSS 20.0软件包对数据进行统计学分析。结果 :机械力诱导6 h后,Klf10、Runx2、OPN和OCN m RNA和蛋白的表达显著升高,ALP活性升高,GLI1和Ptch1蛋白表达升高(P<0.05)。与空白组相比,转染Klf10 si RNA组Klf10 m RNA和蛋白水平显著降低(P<0.05)。同时,Klf10 si RNA显著抑制ALP活性,下调Runx2、OPN和OCN m RNA和蛋白的表达,抑制GLI1和Ptch1蛋白表达(P<0.05)。Purmorphamine显著抑制Klf10 si RNA沉默介导的成骨分化效应(P<0.05)。结论 :Klf10沉默可以抑制机械力作用下HPDLCs的骨向分化,可能与调节hedgehog信号转导通路有关。展开更多
Recent studies have demonstrated a new role for Klf10, a Krüppel-like transcription factor, in skeletal muscle, specifically relating to mitochondrial function. Thus, it was of interest to analyze additional tiss...Recent studies have demonstrated a new role for Klf10, a Krüppel-like transcription factor, in skeletal muscle, specifically relating to mitochondrial function. Thus, it was of interest to analyze additional tissues that are highly reliant on optimal mitochondrial function such as the cerebellum and to decipher the role of Klf10 in the functional and structural properties of this brain region. In vivo (magnetic resonance imaging and localized spectroscopy, behavior analysis) and in vitro (histology, spectroscopy analysis, enzymatic activity) techniques were applied to comprehensively assess the cerebellum of wild type (WT) and Klf10 knockout (KO) mice. Histology analysis and assessment of locomotion revealed no significant difference in Klf10 KO mice. Diffusion and texture results obtained using MRI revealed structural changes in KO mice characterized as defects in the organization of axons. These modifications may be explained by differences in the levels of specific metabolites (myo-inositol, lactate) within the KO cerebellum. Loss of Klf10 expression also led to changes in mitochondrial activity as reflected by a significant increase in the activity of citrate synthase, complexes I and IV. In summary, this study has provided evidence that Klf10 plays an important role in energy production and mitochondrial function in the cerebellum.展开更多
文摘目的 :探讨Klf10沉默对人牙周膜细胞(human periodontal ligament cells,HPDLCs)受力后骨向分化的影响。方法:分离培养HPDLCs,RNA干扰沉默Klf10的表达,用离心加力法对细胞加载机械力,并予hedgehog信号通路特异性激动剂purmorphamine进行干预。采用ELISA法检测碱性磷酸酶(alkaline phosphatase,ALP)活性,采用RT-PCR和Western印迹法分别检测各组细胞Klf10 m RNA和蛋白表达水平,并检测成骨标志基因Runt相关转录因子2(runt-related transcription factor 2,Runx2)、骨桥蛋白(osteopontin,OPN)和骨钙素(osteocalcin,OCN)m RNA和蛋白的表达情况;利用Western免疫印迹法检测hedgehog信号转导通路成员胶质瘤相关癌基因同源物1(glioma-associated oncogene homolog 1,GLI1)和Ptch1(patched-1)的蛋白表达。采用SPSS 20.0软件包对数据进行统计学分析。结果 :机械力诱导6 h后,Klf10、Runx2、OPN和OCN m RNA和蛋白的表达显著升高,ALP活性升高,GLI1和Ptch1蛋白表达升高(P<0.05)。与空白组相比,转染Klf10 si RNA组Klf10 m RNA和蛋白水平显著降低(P<0.05)。同时,Klf10 si RNA显著抑制ALP活性,下调Runx2、OPN和OCN m RNA和蛋白的表达,抑制GLI1和Ptch1蛋白表达(P<0.05)。Purmorphamine显著抑制Klf10 si RNA沉默介导的成骨分化效应(P<0.05)。结论 :Klf10沉默可以抑制机械力作用下HPDLCs的骨向分化,可能与调节hedgehog信号转导通路有关。
文摘Recent studies have demonstrated a new role for Klf10, a Krüppel-like transcription factor, in skeletal muscle, specifically relating to mitochondrial function. Thus, it was of interest to analyze additional tissues that are highly reliant on optimal mitochondrial function such as the cerebellum and to decipher the role of Klf10 in the functional and structural properties of this brain region. In vivo (magnetic resonance imaging and localized spectroscopy, behavior analysis) and in vitro (histology, spectroscopy analysis, enzymatic activity) techniques were applied to comprehensively assess the cerebellum of wild type (WT) and Klf10 knockout (KO) mice. Histology analysis and assessment of locomotion revealed no significant difference in Klf10 KO mice. Diffusion and texture results obtained using MRI revealed structural changes in KO mice characterized as defects in the organization of axons. These modifications may be explained by differences in the levels of specific metabolites (myo-inositol, lactate) within the KO cerebellum. Loss of Klf10 expression also led to changes in mitochondrial activity as reflected by a significant increase in the activity of citrate synthase, complexes I and IV. In summary, this study has provided evidence that Klf10 plays an important role in energy production and mitochondrial function in the cerebellum.
