目的:研究stomatin样蛋白2(stomatin-like protein 2,SLP-2)对人子宫内膜基质细胞系T-HESC的增殖和分化的调控作用。方法:培养子宫内膜基质细胞系T-HESC,体外诱导蜕膜化,RT-q PCR、Western blot和免疫荧光方法检测SLP-2在体外诱导蜕膜...目的:研究stomatin样蛋白2(stomatin-like protein 2,SLP-2)对人子宫内膜基质细胞系T-HESC的增殖和分化的调控作用。方法:培养子宫内膜基质细胞系T-HESC,体外诱导蜕膜化,RT-q PCR、Western blot和免疫荧光方法检测SLP-2在体外诱导蜕膜化模型中的表达变化以及蜕膜化标志分子催乳素(prolactin,PRL)、胰岛素样生长因子结合蛋白1(insulin like growth factor binding protein 1,IGFBP1)和Desmin的表达。转染特异性SLP-2干扰小RNA(si RNA),敲低SLP-2的表达,分析敲低SLP-2表达对基质细胞增殖和分化(蜕膜化)的影响。结果:体外诱导蜕膜化条件下,诱导蜕膜化标志物IGFBP1和PRL的表达明显增加;Desmin显著表达于在体外诱导蜕膜化的细胞中;SLP-2表达明显升高。在蜕膜化细胞中转染SLP-2 si RNA敲低SLP-2表达,蜕膜化行为受到显著抑制;细胞增殖标志分子CCND3表达明显下降,细胞增殖降低。结论:SLP-2的表达可影响人子宫内膜基质细胞细胞系T-HESC的增殖和分化。展开更多
AP-2β是转录因子AP-2家族中的重要一员.AP-2是一个与哺乳动物的发育、细胞生长、分化、凋亡和肿瘤发生都有密切联系的重要的转录因子家族.最近的研究表明,AP-2能通过与其他蛋白的相互作用来调控AP-2下游基因的转录活性.采用免疫共沉淀...AP-2β是转录因子AP-2家族中的重要一员.AP-2是一个与哺乳动物的发育、细胞生长、分化、凋亡和肿瘤发生都有密切联系的重要的转录因子家族.最近的研究表明,AP-2能通过与其他蛋白的相互作用来调控AP-2下游基因的转录活性.采用免疫共沉淀结合质谱鉴定的方法筛选与AP-2β相互作用的蛋白.结果筛选到Stomatin like protein2(SLP-2)蛋白可能与AP-2β相互作用.免疫共沉淀实验证实外源过表达和内源的SLP-2蛋白都能与AP-2β蛋白相互作用.而且,细胞免疫荧光实验发现外源表达的Myc-AP-2β蛋白与内源的SLP-2蛋白在MCF-7细胞的胞质中有共定位.此外,免疫印记实验结果表明过表达SLP-2能上调AP-2β的蛋白水平,这就表明SLP-2与AP-2β相互作用后,SLP-2可能使AP-2β蛋白更稳定.这些研究结果为进一步研究这两个基因的功能提供了新的切入点.展开更多
Myelination of Schwann cells in the peripheral nervous system is an intricate process involving myelin protein trafficking. Recently, the role and mechanism of the endosomal/lysosomal system in myelin formation were e...Myelination of Schwann cells in the peripheral nervous system is an intricate process involving myelin protein trafficking. Recently, the role and mechanism of the endosomal/lysosomal system in myelin formation were emphasized. Our previous results demonstrated that a small GTPase Rab27a regulates lysosomal exocytosis and myelin protein trafficking in Schwann cells. In this present study, we established a dorsal root ganglion （DRG） neuron and Schwann cell co-culture model to identify the signals associated with Rab27a during myelination. First, Slp2-a, as the Rab27a effector, was endogenously expressed in Schwann cells. Second, Rab27a expression significantly increased during Schwann cell myelination. Finally, Rab27a and Slp2-a silencing in Schwann cells not only reduced myelin protein expression, but also impaired formation of myelin-like membranes in DRG neuron and Schwann cell co-cultures. Our findings suggest that the Rab27a/ Slp2-a complex affects Schwann cell myelination in vitro.展开更多
Granulysin is a cytotoxic granular protein that was identified from human T cells by using the gene subtraction method in 1987. Based on its amino acid homology, granulysin belongs to the saposin-like protein family. ...Granulysin is a cytotoxic granular protein that was identified from human T cells by using the gene subtraction method in 1987. Based on its amino acid homology, granulysin belongs to the saposin-like protein family. The bioactive 9-k Da form of granulysin is processed from the 15-k Da pro-product in the cytoplasmic granules. It is expressed in CD8-positive αβT cells 5 d after mitogenic stimulation and constitutively in natural killer(NK) cells and γδT cells, although regulation of its expression has not yet been precisely determined. The 9-k Da granulysin form has anti-microbial activity against microorganisms such as bacteria, fungi, mycobacteria and parasites, as well as tumoricidal activity against some tumors at 1-10 μmol/L concentrations. Granulysin is secreted in both Ca-dependent and-inde-pendent manners. In sera, only the 15-k Da form is detectable and is expected to be a biomarker for immune potency, acute viral infection, anti-tumor immune reaction, acute graft vs host disease, and NK cell associated neoplasm.展开更多
文摘目的:研究stomatin样蛋白2(stomatin-like protein 2,SLP-2)对人子宫内膜基质细胞系T-HESC的增殖和分化的调控作用。方法:培养子宫内膜基质细胞系T-HESC,体外诱导蜕膜化,RT-q PCR、Western blot和免疫荧光方法检测SLP-2在体外诱导蜕膜化模型中的表达变化以及蜕膜化标志分子催乳素(prolactin,PRL)、胰岛素样生长因子结合蛋白1(insulin like growth factor binding protein 1,IGFBP1)和Desmin的表达。转染特异性SLP-2干扰小RNA(si RNA),敲低SLP-2的表达,分析敲低SLP-2表达对基质细胞增殖和分化(蜕膜化)的影响。结果:体外诱导蜕膜化条件下,诱导蜕膜化标志物IGFBP1和PRL的表达明显增加;Desmin显著表达于在体外诱导蜕膜化的细胞中;SLP-2表达明显升高。在蜕膜化细胞中转染SLP-2 si RNA敲低SLP-2表达,蜕膜化行为受到显著抑制;细胞增殖标志分子CCND3表达明显下降,细胞增殖降低。结论:SLP-2的表达可影响人子宫内膜基质细胞细胞系T-HESC的增殖和分化。
文摘AP-2β是转录因子AP-2家族中的重要一员.AP-2是一个与哺乳动物的发育、细胞生长、分化、凋亡和肿瘤发生都有密切联系的重要的转录因子家族.最近的研究表明,AP-2能通过与其他蛋白的相互作用来调控AP-2下游基因的转录活性.采用免疫共沉淀结合质谱鉴定的方法筛选与AP-2β相互作用的蛋白.结果筛选到Stomatin like protein2(SLP-2)蛋白可能与AP-2β相互作用.免疫共沉淀实验证实外源过表达和内源的SLP-2蛋白都能与AP-2β蛋白相互作用.而且,细胞免疫荧光实验发现外源表达的Myc-AP-2β蛋白与内源的SLP-2蛋白在MCF-7细胞的胞质中有共定位.此外,免疫印记实验结果表明过表达SLP-2能上调AP-2β的蛋白水平,这就表明SLP-2与AP-2β相互作用后,SLP-2可能使AP-2β蛋白更稳定.这些研究结果为进一步研究这两个基因的功能提供了新的切入点.
基金supported by the National Natural Science Foundation of China,No.31071251,81471255,and 81471259a grant from the Ministry of Science and Technology of China(973 Program),No.2014CB542202+2 种基金a grant from the Basic Research Program of Education Department of Jiangsu Province,China,No.14KJA310004a grant from the Natural Science Research Project of Nantong Science and Technology Bureau,China,No.HS2013014a grant from the Natural Science Research Project of Nantong University,China,No.13Z008
文摘Myelination of Schwann cells in the peripheral nervous system is an intricate process involving myelin protein trafficking. Recently, the role and mechanism of the endosomal/lysosomal system in myelin formation were emphasized. Our previous results demonstrated that a small GTPase Rab27a regulates lysosomal exocytosis and myelin protein trafficking in Schwann cells. In this present study, we established a dorsal root ganglion （DRG） neuron and Schwann cell co-culture model to identify the signals associated with Rab27a during myelination. First, Slp2-a, as the Rab27a effector, was endogenously expressed in Schwann cells. Second, Rab27a expression significantly increased during Schwann cell myelination. Finally, Rab27a and Slp2-a silencing in Schwann cells not only reduced myelin protein expression, but also impaired formation of myelin-like membranes in DRG neuron and Schwann cell co-cultures. Our findings suggest that the Rab27a/ Slp2-a complex affects Schwann cell myelination in vitro.
基金Supported by The Grant-in-Aid for Scientific Research from Ministry of Education,Science and Culture Japan,24591541 to Nagasawa M
文摘Granulysin is a cytotoxic granular protein that was identified from human T cells by using the gene subtraction method in 1987. Based on its amino acid homology, granulysin belongs to the saposin-like protein family. The bioactive 9-k Da form of granulysin is processed from the 15-k Da pro-product in the cytoplasmic granules. It is expressed in CD8-positive αβT cells 5 d after mitogenic stimulation and constitutively in natural killer(NK) cells and γδT cells, although regulation of its expression has not yet been precisely determined. The 9-k Da granulysin form has anti-microbial activity against microorganisms such as bacteria, fungi, mycobacteria and parasites, as well as tumoricidal activity against some tumors at 1-10 μmol/L concentrations. Granulysin is secreted in both Ca-dependent and-inde-pendent manners. In sera, only the 15-k Da form is detectable and is expected to be a biomarker for immune potency, acute viral infection, anti-tumor immune reaction, acute graft vs host disease, and NK cell associated neoplasm.
