Dehydroepiandrosterone sulfate-binding sites in plasma membrane from human uterine cervical fibroblasts

Dehydroepiandrosterone sulfate (DHA-S) plays a critical role in cervical dilation at labor. Incubation of cervical fibroblasts with [³H]DHA-S caused a rapid and saturable increase in cellular radioactivity: an apparent equilibrium was reached by 2 min. There was no detectable conversion of DHA-S into DHA or oestradiol. When the fibroblasts loaded with [³H]DHA-S were homogenized and fractionated, the specific radioactivity in the plasma membrane fraction was enriched approximately 8- to 9-fold compared with the whole homogenate; only low amounts of radioactivity were observed in the other subcellular fractions. The binding of DHA-S to plasma membrane preparations showed saturation kinetics with an apparent equilibrium dissociation constant (K _d) of 12 nM, and the binding capacity (B _max) was calculated to be 1.25 fmol/mg protein. Neither DHA nor oestrone sulfate affected [³H]DHA-S binding to the plasma membrane. The plasma membranes of skin fibroblasts did not show specific binding sites for DHA-S. These findings demonstrate the presence of specific binding sites for DHA-S in the plasma membrane of cervical stroma cells. The fetal adrenal steroid may exert its action on cervical ripening at least in part through membrane-associated binding sites, or receptors.     

新型前交叉韧带支架材料的构建及其生物相容性的实验研究

探讨以聚羟基丁酸己酯/聚左旋乳酸(PHB/PLLA1∶1)胶原杂化支架作为前交叉韧带组织工程载体材料的可行性。制备"三明治"样结构PHB/PLLA共聚物并测量其孔隙率等指标。以I型胶原对制备的PHB/PLLA支架进行杂化,获得PHB/PLLA胶原杂化支架。扫描电镜观察其表面结构。将兔皮肤成纤维细胞(SF)接种于PHB/PLLA胶原杂化支架,共培养5d后,扫描电镜下观察其在材料上生长情况。PHB/PLLA支架杂化后胶原填充于纤维空隙,分布比较均匀。体外培养的皮肤成纤维细胞成功种植在支架材料上,在材料上粘附、生长良好。说明构建的支架材料具有良好的三维构型和生物相容性,有望为前交叉韧带损伤的修复提供了一种新型的支架材料。     

采用图像信息学方法研究三维支架中细胞的信号通路

通过慢病毒介导的短发夹RNA对成纤维细胞的转化生长因子β(TGF-β)信号通路进行干扰,采用双光子荧光显微成像技术并结合基于细胞-细胞外基质三维系统的图像信息学方法,研究了植于多孔胶原蛋白支架中成纤维细胞分化过程的TGF-β信号通路.结果表明,在介导成纤维细胞分化的过程中,生长因子TGF-β1比TGF-β3发挥的作用更大,在TGF-β1介导的成纤维细胞分化过程中,蛋白SMAD1和SMAD3通过不同方式也发挥了重要作用.所得结果与通过生化或遗传学方法的结果相吻合.     

Connective tissue growth factor induces transformation of renal fibroblasts into myofibroblasts

Connective tissue growth factor (CTGF) is thought to be a specific mediator of TGF-β profibrogenic effect, but the role of CTGF in the transformation of renal fibroblasts to myofibroblasts, which are the most important host cells in the renal chronic fibrosis, is still unknown. Aimed at observing the biological effect of CTGF, we set up a cell line overexpressing CTGF, and analyzed the amount of myofibroblast and the level of extracellular matrix Collagen Ⅲ mRNA in cultured cells. Our results show that CTGF can directly induce myofibroblasts formation, and increase the level of Collagen Ⅲ mRNA. This suggests that CTGF may be a novel pharmacotherapeutical target protein for the interference with renal fibrosis, thus providing useful theoretic possibility for clinical prevention of chronic progression of kidney disease.     

Involvement of small Ras GTPases and their effectors in chronic renal disease

The mechanisms involved in the development of renal fibrosis are poorly understood. Small Ras GTPases control cell proliferation, differentiation, cellular growth and apoptosis, with cell-specific expression in the kidney. Cytokines, high glucose medium or advanced glycation end-products activate Ras in different renal cells. Increased Ras activation has been found in experimental tubulointerstitial fibrosis. Transforming growth factor-β1 (TGF-β1) and Ras signalling pathways are close related: TGF-β1 overcomes Ras mitogenic effects, and Ras counteracts TGF-β signalling. However, Ras activation is also an intracellular signal transduction point for several molecules (e.g. TGF-β1) involved in kidney damage. Ras isoforms play different roles in regulating extracellular matrix synthesis in fibroblasts and mesangial cells. These data give evidence for a role for Ras in renal fibrosis, but no reviews are available on the role of p21 Ras in this process. Thus, our goal is to review the role of Ras activation and signalling in renal fibrosis. Received 7 June 2007; received after revision 17 September 2007; accepted 1 October 2007     

Ciliated fibroblasts and smooth muscle cells in the rat uterus

Summary Ciliation in endometrial fibroblasts and myometrial muscle cells of the rat was examined by transmission electron microscopy. Quantification of the number of ciliated cells during the estrus cycle did not show any firm relationship between cilation and ovarian hormonal activity. In the case of most cilia, there is a spatial relationship between their basal centrioles and the Golgi complex, so that a Golgi-cilium complex is created. A possible role of ciliation in uterine fibroblasts and smooth muscle cells is discussed.     

Erythrophagocytosis by cultured skin fibroblasts from patients with hereditary metabolic disorders

Phagocytosis of native allogenic red blood cells was observed in cultures of skin fibroblasts obtained from patients with neuronal ceroid-lipofuscinosis, Niemann-Pick disease type C and morbus Fabry. Occasional phagocytizing cells were observed in 9 other syndromes. Cells from three normal donors did not phagocytize.     

葡甘聚糖-壳聚糖复合膜的制备及细胞相容性评价

采用溶液共混法制备了葡甘聚糖-壳聚糖复合膜,并测定了材料的吸水率等理化性能。模拟体内生理条件用溶菌酶对复合膜进行体外酶解实验,并与壳聚糖膜作了比较;以体外培养的小鼠成纤维细胞(N IH 3T 3)为对象,利用复合膜的浸渍液培养与膜材料表面直接培养法,同时结合复合膜的溶血反应实验,研究了复合膜的细胞相容性。结果表明:溶菌酶对复合膜的生物降解有促进作用,复合膜降解速率较壳聚糖膜快。四甲基偶氮唑盐比色法(MTT法)实验表明复合膜浸渍液对细胞无毒性效应,显微镜和扫描电镜显示N IH 3T 3在复合膜上能很好地贴附,生长旺盛。复合膜溶血率小于5%。葡甘聚糖-壳聚糖复合膜具有良好的生物降解性和细胞相容性。     

培养方法及胎牛血清浓度对人胎儿成纤维细胞体外生长的影响

采用单细胞培养法和组织块培养法从原代培养人胎儿成纤维细胞,并在不同体积分数胎牛血清(FBS)(0%、4%、8%、10%、12%)的条件下观察人胎儿成纤维细胞培养的状况并采用台盼蓝染色法进行活细胞计数.结果表明,早期由组织块培养法原代培养获取的细胞形态要比单细胞培养法原代培养获取的细胞稍好些,且在细胞传代后极少出现无法贴壁的死细胞.FBS对hFFs生长影响比较明显.在实际应用中,在体积分数8%FBS下,采用组织块培养法进行人胎儿成纤维细胞体外培养,效果良好.     

前交叉韧带组织工程种子细胞筛选的初步研究

为组织工程方法构建前交叉韧带筛选一种理想的种子细胞。用体外分离培养兔前交叉韧带（ACL）、内侧副韧带（MCL）、跟腱（AT）、皮肤成纤维细胞（SK）,比较四种细胞的生长状态、增殖活性及Ⅰ、Ⅲ型胶原分泌量。在含体积分数10％胎牛血清的DMEM培养液中,四种细胞生长良好、细胞形态相似,但以SK细胞生长最快,AT细胞次之,MCL细胞和ACL细胞生长最慢。SK、MCL、ACL细胞均分泌Ⅰ、Ⅲ型胶原,AT细胞只分泌Ⅰ型胶原。SK细胞Ⅰ型胶原染色密度较MCL、ACL、AT细胞高,SK细胞Ⅲ型胶原染色密度较MCL、ACL细胞高。说明SK细胞增殖活性高,分泌Ⅰ、Ⅲ型胶原多,取材简便,是理想的前交叉韧带组织工程种子细胞。