Expression of beta-nerve growth factor receptor mRNA in Sertoli cells downregulated by testosterone

Nerve growth factor (NGF) is synthesized in male germ cells. The NGF receptor (NGFR) mRNA was found in the Sertoli cells of rat testis. Hypophysectomy increased both NGFR mRNA in testis and the number of NGFR hybridizing cells in seminiferous tubules. This was suppressed by treatment with chorionic gonadotropin or testosterone, but not with follicle-stimulating hormone. The NGFR mRNA also increased after destruction of Leydig cells or blocking of the androgen receptor. This suggests that NGF produced by male germ cells regulates testicular function in an androgen-modulated fashion by mediating an interaction germ and Sertoli cells.     

Molecular cloning of a gene sequence regulated by nerve growth factor

Nerve growth factor (NGF) is essential for the development and differentiation of sympathetic or sensory neurons. A complementary DNA was cloned that corresponds to a gene sequence induced more than 50-fold in a cultured target cell line of pheochromocytoma cells (PC12 cells) 5 hours after the addition of NGF. The induced messenger RNA encodes a 90,000-dalton polypeptide that may represent one of the primary events in NGF-induced differentiation of neurons.     

Bone cells: a serum-free medium supports proliferation in primary culture

Bone-cells isolated from embryonic rat calvaria increase in number two-to threefold when cultured at high, but not at low, population densities in a serum-free medium that contains albumin. Cultured cells respond to parathyroid hormone and exhibit a marked rise in alkaline phosphatase activity during proliferation, which suggests the progressive differentiation or preferential growth of osteoblast-like cells.     

Monocyte-derived human B-cell growth factor identified as interferon-beta 2 (BSF-2, IL-6)

Soluble products of either Epstein-Barr virus (EBV)-infected B cells or activated monocytes promote the proliferation of EBV-infected B cells and permit their growth at low cell densities. This suggests that growth factors are important for B-cell immortalization by EBV. In this study, a monocyte-derived factor that promotes the growth of EBV-infected b cells was purified and identified as interferon-beta 2 (IFN-beta 2), which is also known as 26-kilodalton protein, B-cell differentiation factor (BSF-2), and interleukin-6 (IL-6). The purified protein has a specific activity of approximately 4 X 10(7) units per milligram of protein in assays of B-cell growth. Thus, IFN-beta 2/BSF-2 is a B-cell growth factor that promotes the proliferation of human B cells infected with EBV.     

Nonidentity of fetuin and protein growth (flattening) factor

Fetuin, a fetal calf serum glycoprotein, appeared to possess activity with cultured mammalian cells similar to that of a protein growth factor partially purified from adult bovine and human sera. Column chromatography, however, yielded highly purified but inactive fetuin. These results leave open questions regarding the role of this interesting and readily purified protein.     

兔脂肪间充质干细胞分离培养及鉴定

为了建立一种体外培养兔脂肪间充质干细胞(ADSCs)的方法,在无菌条件下取家兔双侧腹股沟脂肪垫,Ⅰ型胶原酶消化后,用含10%胎牛血清的DMEM 培养液培养.细胞连续传代后,MTT法检测第3、5、10代细胞的增殖情况.采用油红染色法和Von Kossa染色法观察第3代ADSCs向成脂和成骨方向分化情况,并采用流式细胞仪分析细胞表面标志.结果显示,第3、5、10代的ADSCs生长曲线并无显著差异,细胞增殖较快;ADSCs成脂诱导后油红染色阳性,成骨诱导后Von kossa染色形成典型的黑色钙化结节.ADSCs 表面标志检测结果为:CD29 和CD90阳性,CD45和CD80阴性.成功分离和培养了兔的ADSCs,其具有成脂和成骨潜能,可作为组织工程的种子细胞.     

大鼠胎儿神经干细胞的分离、培养及诱导分化

为了探讨神经干细胞(Neural stem cells,NSCs)在体外培养中的生物学特性,研究不同细胞因子对NSCs分化的影响,试验对大鼠胎儿神经干细胞进行了分离培养,并研究了10,50和100 ng/mL的神经生长因子(nerve growth factor,NGF)、脑源性神经营养因子(brain derived neruotrophic factor,BDNF)和胶质源性神经生长因子(glial cell line-derived neurotrophic factor,GDNF)对NSCs的定向诱导分化作用。结果表明,大鼠胎儿NSCs可在体外增殖并形成典型的神经球,指数增长期在传代后的第4~5天;10和50 ng/mL的NGF、BDNF和GDNF对NSCs的诱导分化作用不明显,100 ng/mL的NGF、BDNF和GDNF可诱导NSCs分化为神经元或胶质细胞。说明NGF和BDNF可诱导NSCs向神经元方向分化,GDNF诱导其向胶质细胞分化。     

Nerve growth factor gene expression in the developing rat brain

The regulation of nerve growth factor (NGF) protein and NGF messenger RNA (mRNA) in the developing rat brain has been studied to assess the hypothesis that NGF supports the differentiation of cholinergic neurons in the basal forebrain. In the adult, the major targets of these neurons, the hippocampus and neocortex, contain the highest concentrations of NGF mRNA, but comparatively low ratios of NGF protein to its mRNA. In contrast, a high concentration of NGF protein and a low concentration of NGF mRNA were seen in the basal forebrain, consistent with retrograde transport of NGF protein into this region from the neocortex and hippocampus. In these two target regions NGF and NGF mRNA were barely detectable at birth, their concentrations increased to a peak at day 21, and then NGF mRNA, but not NGF protein, declined threefold by day 35. NGF accumulation in the basal forebrain paralleled that in the target regions and preceded an increase in choline acetyltransferase, suggesting that the differentiation of cholinergic projection neurons is indeed regulated by retrogradely transported NGF. In addition, high ratios of NGF protein to NGF mRNA, comparable to that in the basal forebrain, were seen in the olfactory bulb and cerebellum, suggesting that NGF may be transported into these regions by unidentified neurons.     

Vasoconstriction: a new activity for platelet-derived growth factor

Platelet-derived growth factor (PDGF) is a potent mitogen for vascular smooth muscle cells that has been implicated in the pathogenesis of atherosclerosis. The potential role of PDGF in the altered vasoreactivity of atherosclerotic vessels has been studied through an examination of its effects on contractility in the rat aorta. PDGF caused a concentration-dependent contraction of aortic strips and was significantly more potent on a molar basis than the classic vasoconstrictor peptide angiotensin II. Furthermore, PDGF increased the cytosolic free calcium concentration in cultured rat aortic smooth muscle cells. These observations suggest a new biological activity for PDGF that may contribute to the enhanced vasoreactivity of certain atherosclerotic vessels.     

Superinduction of c-fos by nerve growth factor in the presence of peripherally active benzodiazepines

Alterations in proto-oncogene expression after stimulation of rat pheochromocytoma (PC12) cells by nerve growth factor (NGF) have been investigated. A specific stimulation of c-fos messenger RNA and protein was detected 30 minutes after treatment. This induction was enhanced more than 100-fold in the presence of peripherally active benzodiazepines. The effect was specific as very little change was observed in the levels of c-rasHa, c-rasKi, c-myc, and N-myc messenger RNA's. Under the conditions used here, NGF treatment ultimately results in neurite outgrowth, with a reduction or cessation of cell division. Thus, stimulation of the c-fos gene in this system appeared to be associated with differentiation and not with cellular proliferation. The effect of benzodiazepines was stereospecific and represents a novel action of these compounds at the level of gene expression.     

Simian sarcoma virus--transformed cells secrete a mitogen identical to platelet-derived growth factor

Normal rat kidney (NRK) cells transformed by simian sarcoma virus (SSV) release into the culture medium a biologically active mitogen with properties identical to those of human platelet-derived growth factor (PDGF). Like PDGF, the growth factor derived from SSV-NRK cells was shown to be stable to heat and sensitive to reducing agents. It was capable of inhibiting binding of labeled PDGF to the receptor on human fibroblasts. It also stimulated the phosphorylation of the same membrane protein (185 kilodaltons) in isolated plasma membranes from human fibroblasts. Immunoprecipitation of metabolically labeled proteins released by SSV-NRK cells showed that a 34-kilodalton protein was specifically precipitated by antiserum to PDGF. Upon reduction, this protein had a molecular size of 17 kilodaltons. PDGF has been shown to consist of two 14- to 18-kilodalton proteins linked by disulfide bonds.     

波形纤维蛋白在前体脂肪细胞分化中的表达

采用免疫组化 (HIGH- SABC)法对前体脂肪细胞分化中波形纤维蛋白的表达进行了定性和定量检测 ,并结合计算机图像分析系统对细胞形态参数值 (细胞总面积、胞核面积、细胞周长、等效直径及胞核质比 )进行了分析。结果表明 :(1)波形纤维蛋白参与前体脂肪细胞分化的全过程 ,而且其表达具有明显的时间依赖性——随分化的进行呈下降的趋势 ,染色模式为细胞质 ;(2 )在前体脂肪细胞积聚脂滴向脂肪细胞分化的过程中 ,细胞总面积、胞核面积、细胞周长和等效直径总的呈上升趋势 ,胞核质比呈下降倾向。     

内在调节蛋白及生长因子对精原干细胞增殖与分化的调控

文章阐述了在转录水平上精原干细胞在增殖与分化过程中的内在调节蛋白Plzf、Sohlh及其对精原干细胞的调节机制。介绍了胶质细胞源性神经营养因子、干细胞因子、Ets相关分子、骨形态发生蛋白、肝细胞生长因子、斯里兰卡肉桂碱受体等由支持细胞分泌的对精原细胞增殖与分化有重要作用的生长因子。     

Immunoprecipitation of insulin receptors from cultured human lymphocytes (IM-9 cells) by antibodies to pp60src

The family of tyrosine-specific protein kinases includes proteins encoded by retroviral oncogenes as well as receptors for insulin and several growth factors. Antibodies to pp60src, the protein encoded by the src oncogene of Rous sarcoma virus (RSV), can specifically immunoprecipitate affinity-labeled insulin receptors from cultured human lymphocytes (IM-9 cells). This precipitation is specifically inhibited by the src gene product purified from RSV-transformed rat cells. These observations provide evidence that there is structural homology between the insulin receptors and pp60src.     

猪肌肉干细胞在三维水凝胶中的分化研究

【目的】 探究猪肌肉干细胞在三维水凝胶中的分化效果,为体外诱导肌肉干细胞分化成为肌肉组织提供方法指导。【方法】 将一定数目的猪肌肉干细胞分别在二维和三维条件下诱导分化（二维条件指在培养皿中培养细胞,三维条件指在水凝胶中培养细胞）,分别收取增殖阶段、预分化阶段、分化初期、分化成熟、分化末期的二维培养细胞的RNA和蛋白样品,以及分化7和14 d的三维培养细胞的RNA和蛋白样品。利用RT-qPCR技术检测细胞在两种条件下分化至不同阶段时成肌相关基因MYOG、CAV-3、MyHC-slow、MyHC-2a的表达水平;利用Western Blot技术检测细胞在两种条件下分化至不同阶段时MYOG、MyHC蛋白的表达水平;利用免疫荧光染色技术观察猪肌肉干细胞在二维和三维条件下融合形成的肌管;使用氨基酸自动分析仪检测分化14 d培养肌肉组织的氨基酸含量及组成。【结果】 二维培养的猪肌肉干细胞在分化第3天时开始发生肌融合,在分化第7天形成成熟肌管,随后进入分化末期,肌管开始脱落。三维培养的猪肌肉干细胞在分化第7天时还未完全伸展,细胞的MYOG和CAV-3表达水平低;分化第14天时水凝胶内已形成多核肌管,MYOG和CAV-3表达达到二维分化水平。三维分化有利于终末分化基因MyHC-slow和MyHC-2a的表达,分化14 d时MyHC-slow的表达量是二维分化7 d的12倍,MyHC-2a的表达量是二维分化7 d的4倍,但是MyHC蛋白的表达量仅为二维分化7 d时的1/6。氨基酸分析结果表明体外培养肌肉组织中17种水解氨基酸含量均低于猪肉,且必需氨基酸在总氨基酸的占比也低于猪肉,但是呈味氨基酸的占比相较猪肉更高。【结论】 猪肌肉干细胞可以在三维胶原水凝胶中分化形成肌管,且三维条件有利于成肌分化相关基因表达,但要实现MyHC蛋白的高表达还需进一步研究,按此方法体外培养的肌肉组织有较高的呈味氨基酸含量,可能会有较好的风味。     

Coexistence of guanylate cyclase and atrial natriuretic factor receptor in a 180-kD protein

Atrial natriuretic factor (ANF) is a peptide hormone that is released from atria and regulates a number of physiological processes, including steroidogenesis in adrenal cortex and testes. The parallel stimulation of membrane guanylate cyclase and corticosterone production in isolated fasciculata cells of rat adrenal cortex has supported the hypothesis of a mediatory role for cyclic guanosine monophosphate (cyclic GMP) in signal transduction. A novel particulate guanylate cyclase tightly coupled with ANF receptor was purified approximately 273,000-fold by two-step affinity chromatography. The enzyme had a molecular size of 180 kilodaltons and was acidic in nature with a pI of 4.7. Its specific activity was 1800 nanomoles of cyclic GMP formed per minute per milligram of protein. The purified enzyme bound ANF with a specific binding activity of 4.01 nanomoles per milligram of protein, a value that is close to the theoretical binding activity of 5.55 nanomoles per milligram of protein for 1 mole of the ligand binding 1 mole of the receptor protein. These results indicate that the guanylate cyclase-coupled ANF receptor exists in a 180-kilodalton protein of rat adrenocortical carcinoma and represent a step toward the elucidation of the basic mechanism of cyclic GMP-mediated transmembrane signal transduction in response to a hormone.     

Growth factors modulate gonadotropin receptor induction in granulosa cell cultures

Epidermal growth factor and fibroblast growth factor inhibited follicle-stimulating hormone-dependent induction of luteinizing hormone receptor in cultured ovarian granulosa cells of the rat. In contrast, platelet-derived growth factor potentiated the induction of luteinizing hormone receptor by follicle-stimulating hormone. These results indicate that growth factors, well known for their effects on mitosis and DNA synthesis in cultured mammalian cells, are also able to modulate hormone-dependent differentiation in an endocrine target cell.     

Tumor cell rejection through terminal cell differentiation

Leukemic cells cultured in the presence of various conditioned media differentiate into macrophages. This finding suggested that the maintenance of undifferentiated state and self-renewal in vivo may be related to the inability of the host to generate an appropriate level of differentiation factor (DF). Evidence for this hypothesis was derived from experiments in vitro and in vivo with myeloid leukemia of rat. The following results were obtained: (i) in vitro, the percentage of cell differentiation at a fixed concentration of DF was inversely related to the concentration of cells; (ii) leukemic cell inoculates that were lethal to 7-day-old rats were rejected by 21-day-old rats; (iii) leukemic cells in diffusion chambers underwent differentiation in 21-day-old rats but not in 7-day-old rats; (iv) organs from 21-day-old rats contained more DF activity than those of 7-day-old rats; (v) treatment of rats with DF in diffusion chambers resulted in leukemic cell differentiation inside the chamber; and (vi) the development of leukemia in 7-day-old rats was aborted by treatment with DF. These results show that the differentiation of rat leukemia cells requires the appropriate level of DF. The proliferation of transplanted leukemia cells in 7-day-old rats goes unchecked because of inadequate generation of DF. Conversely, in the 21-day-old rats, rejection is accomplished by differentiation of the transplanted cells.     

The trk proto-oncogene product: a signal transducing receptor for nerve growth factor.

The trk proto-oncogene encodes a 140-kilodalton, membrane-spanning protein tyrosine kinase (p140prototrk) that is expressed only in neural tissues. Nerve growth factor (NGF) stimulates phosphorylation of p140prototrk in neural cell lines and in embryonic dorsal root ganglia. Affinity cross-linking and equilibrium binding experiments with 125I-labeled NGF indicate that p140prototrk binds NGF specifically in cultured cells with a dissociation constant of 10(-9) molar. The identification of p140prototrk as an NGF receptor indicates that this protein participates in the primary signal transduction mechanism of NGF.     

Transforming growth factor-beta: biological function and chemical structure

Transforming growth factor-beta (TGF-beta) is a multifunctional peptide that controls proliferation, differentiation, and other functions in many cell types. Many cells synthesize TGF-beta and essentially all of them have specific receptors for this peptide. TGF-beta regulates the actions of many other peptide growth factors and determines a positive or negative direction of their effects. Its marked ability to enhance formation of connective tissue in vivo suggests several therapeutic applications.