Early growth response gene 1-mediated apoptosis is essential for transforming growth factor beta1-induced pulmonary fibrosis

Fibrosis and apoptosis are juxtaposed in pulmonary disorders such as asthma and the interstitial diseases, and transforming growth factor (TGF)-beta(1) has been implicated in the pathogenesis of these responses. However, the in vivo effector functions of TGF-beta(1) in the lung and its roles in the pathogenesis of these responses are not completely understood. In addition, the relationships between apoptosis and other TGF-beta(1)-induced responses have not been defined. To address these issues, we targeted bioactive TGF-beta(1) to the murine lung using a novel externally regulatable, triple transgenic system. TGF-beta(1) produced a transient wave of epithelial apoptosis that was followed by mononuclear-rich inflammation, tissue fibrosis, myofibroblast and myocyte hyperplasia, and septal rupture with honeycombing. Studies of these mice highlighted the reversibility of this fibrotic response. They also demonstrated that a null mutation of early growth response gene (Egr)-1 or caspase inhibition blocked TGF-beta(1)-induced apoptosis. Interestingly, both interventions markedly ameliorated TGF-beta(1)-induced fibrosis and alveolar remodeling. These studies illustrate the complex effects of TGF-beta(1) in vivo and define the critical role of Egr-1 in the TGF-beta(1) phenotype. They also demonstrate that Egr-1-mediated apoptosis is a prerequisite for TGF-beta(1)-induced fibrosis and remodeling.     

Differential regulation of mesothelial cell fibrinolysis by transforming growth factor beta 1

Management of hypertension and diabetes mellitus in primary health care requires occasional assessment of kidney function. Monitoring the urinary albumin excretion every 24?h is often used as a diagnostic gold standard but measurement of U‐Albumin concentrations in morning urine either alone or together with U‐Creatinine is a well‐established surrogate measure. We compared the ratio U‐Albumin/U‐Creatinine and U‐Albumin concentrations measured by commonly used POC (Point of care) instruments with those obtained in a central laboratory and estimated the uncertainty of the results after establishing an uncertainty budget. It is concluded that the presentation of ratios or concentrations on an ordinal scale is less satisfactory than reporting U‐Albumin concentration on a ratio scale. Moreover the latter will have the advantage of allowing the physician to adjust the diagnostic sensitivity and specificity to local needs. The present report is a methodological study and does not consider the diagnostic performance of the studied properties per se.     

Production of transforming growth factor beta by human T lymphocytes and its potential role in the regulation of T cell growth

This study examines the potential role of transforming growth factor beta (TGF-beta) in the regulation of human T lymphocyte proliferation, and proposes that TGF-beta is an important autoregulatory lymphokine that limits T lymphocyte clonal expansion, and that TGF-beta production by T lymphocytes is important in T cell interactions with other cell types. TGF-beta was shown to inhibit IL-2-dependent T cell proliferation. The addition of picograms amounts of TGF-beta to cultures of IL-2-stimulated human T lymphocytes suppressed DNA synthesis by 60-80%. A potential mechanism of this inhibition was found. TGF-beta inhibited IL-2-induced upregulation of the IL-2 and transferrin receptors. Specific high-affinity receptors for TGF-beta were found both on resting and activated T cells. Cellular activation was shown to result in a five- to sixfold increase in the number of TGF-beta receptors on a per cell basis, without a change in the affinity of the receptor. Finally, the observations that activated T cells produce TGF-beta mRNA and that TGF-beta biologic activity is present in supernatants conditioned by activated T cells is strong evidence that T cells themselves are a source of TGF-beta. Resting T cells were found to have low to undetectable levels of TGF-beta mRNA, while PHA activation resulted in a rapid increase in TGF-beta mRNA levels (within 2 h). Both T4 and T8 lymphocytes were found to make mRNA for TGF-beta upon activation. Using both a soft agar assay and a competitive binding assay, TGF-beta biologic activity was found in supernatants conditioned by T cells; T cell activation resulted in a 10-50-fold increase in TGF-beta production. Thus, TGF-beta may be an important antigen-nonspecific regulator of human T cell proliferation, and important in T cell interaction with other cell types whose cellular functions are modulated by TGF-beta.     

Disruption of T cell homeostasis in mice expressing a T cell-specific dominant negative transforming growth factor beta II receptor

The immune system, despite its complexity, is maintained at a relative steady state. Mechanisms involved in maintaining lymphocyte homeostasis are poorly understood; however, recent availability of transgenic (Tg) and knockout mouse models with altered balance of lymphocyte cell populations suggest that cytokines play a major role in maintaining lymphocyte homeostasis. We show here that transforming growth factor (TGF)-beta plays a critical role in maintaining CD8(+) T cell homeostasis in a Tg mouse model that specifically overexpresses a dominant negative TGF-beta II receptor (DNRII) on T cells. DNRII T cell Tg mice develop a CD8(+) T cell lymphoproliferative disorder resulting in the massive expansion of the lymphoid organs. These CD8(+) T cells are phenotypically "naive" except for the upregulation of the cell surface molecule CD44, a molecule usually associated with memory T cells. Despite their dominance in the peripheral lymphoid organs, CD8(+) T cells appear to develop normally in the thymus, suggesting that TGF-beta exerts its homeostatic control in the peripheral immune system.     

Dog mastocytoma cells produce transforming growth factor beta 1.

Transforming growth factor-beta (TGF beta) promotes deposition of extracellular matrix and is associated with fibrotic conditions both in experimental animals and in humans. Although a role for mast cells has been suspected in the pathogenesis of fibrosis, no potent mediator capable of stimulating fibroblast growth or extracellular matrix deposition has been identified in mast cell supernatants. We report here the constitutive production of TGF beta 1 by four dog mastocytoma cell lines. TGF beta 1 was identified by characteristic biologic activity, blockade of biologic effect by specific neutralizing antibody, and by recognition of a band with the appropriate migration by western blot. TGF beta 1 mRNA, but not TGF beta 2 or TGF beta 3 mRNA, was also produced constitutively by all four cell lines. Quantitation by bioassay revealed baseline TGF beta secretion of approximately 1 ng/10(6) cells over 48 h. Stimulation of mastocytoma cells with phorbol ester increased the rate of release of TGF beta 1, most markedly in the first 30 min after stimulation, without increasing TGF beta 1 mRNA. Dog mastocytoma cells produced TGF beta 1 primarily in a latent form, inactive until treated with acid. Both pure TGF beta 1 and TGF beta-containing mastocytoma cell-conditioned media inhibited mitogenesis and proliferation in dog mastocytoma cell lines, suggesting that mast cell tumor lines would not grow preferentially based on their ability to produce TGF beta. These studies may make possible further investigation of the mechanism by which mast cells contribute to the induction of fibrosis.     

转化生长因子β1在兔房水中的浓度变化

背景:转化生长因子β1可参与角膜损伤后的修复。目的:观察转化生长因子β1滴眼液滴眼后房水中的浓度变化规律。方法:将新西兰大白兔随机分为5组,分别给予PBS和质量浓度为0.5,1.0,2.0,4.0mg/L的转化生长因子β1滴眼液滴右眼。结果与结论:通过裂隙灯观察兔角膜和结膜结构,各组兔眼均无结膜分泌物、球结膜充血、角膜水肿增厚、角膜后沉着物、前房炎性反应及晶状体混浊改变。ELISA检测结果显示,与PBS组比较,质量浓度2.0和4.0mg/L转化生长因子β1滴眼液能有效提高兔眼房水中转化生长因子β1的质量浓度(P<0.01),角膜穿透性良好,在房水中可以达到有效的治疗浓度。     

转化生长因子β1在大鼠房水中的药物浓度

背景:研究证实,转化生长因子β1滴眼液具有促进角膜损伤愈合的作用.目的:观察转化生长因子β1滴眼液表面应用后在房水中的药物浓度,验证其在预防和治疗角膜移植排斥反应中的作用.设计、时间及地点:随机对照动物实验,于2008-04/07在珠江医院中心实验室完成.材料:成年雌性SPF级SD大鼠32只,体质量200-250 g.干粉状转化生长因子β1为美国SANTA公司产品.方法:将32只大鼠随机分为4组,空白对照组,转化生长因子β1 0.5,1,2mg/L组,每组各8只.分别给予生理盐水、0.5,1,2mg/L转化生长因子β1滴眼液,3次/d,用药1周.主要观察指标:①观察眼部刺激症状.②应用酶联免疫吸附法定量测定房水中的转化生长因子β1质量浓度.结果:大体与裂隙灯显微镜观察,实验前后和实验过程中,各组大鼠眼均无结膜分泌物、球结膜充血、角膜水肿、角膜后沉着物、前房炎性反应及晶状体混浊改变.空白组和3种不同浓度的转化生长因子β1滴眼液滴眼后房水中药物质量浓度分别是(0.429±0.187),(0.964±0.284),(2.127±0.673),(2.282±0.745)μg/L.结论:1 mg/L和2mg/L转化生长因子β1滴眼液能有效提高大鼠眼房水中转化生长因子β1的质量浓度(P<0.05),具有良好的角膜穿透性,在房水中可以达到有效的治疗浓度.转化生长因子β1滴眼液能够在局部发挥药物的作用,避免药物全身应用的不良反应.     

转化生长因子β1在兔房水中的浓度变化

背景：转化生长因子β1可参与角膜损伤后的修复。目的：观察转化生长因子β1滴眼液滴眼后房水中的浓度变化规律。方法：将新西兰大白兔随机分为5组,分别给予PBS和质量浓度为0.5,1.0,2.0,4.0mg/L的转化生长因子β1滴眼液滴右眼。结果与结论：通过裂隙灯观察兔角膜和结膜结构,各组兔眼均无结膜分泌物、球结膜充血、角膜水肿增厚、角膜后沉着物、前房炎性反应及晶状体混浊改变。ELISA检测结果显示,与PBS组比较,质量浓度2.0和4.0mg/L转化生长因子β1滴眼液能有效提高兔眼房水中转化生长因子β1的质量浓度（P〈0.01）,角膜穿透性良好,在房水中可以达到有效的治疗浓度。     

Bacterial cell wall-induced immunosuppression. Role of transforming growth factor beta

Group A streptococcal cell wall (SCW)-injected rats exhibit a profound immunosuppression that persists for months after the initial intraperitoneal injection of SCW. The goal of this study was to determine the mechanisms for the suppressed T lymphocyte proliferative responses in this experimental model of chronic inflammation. When spleen cell preparations were depleted of adherent cells, restoration of T cell proliferative responses to Con A and PHA occurred, implicating adherent macrophages in the regulation of immunosuppression. Furthermore, macrophages from SCW-treated animals, when cocultured with normal spleen cells in the presence of Con A or PHA, effectively inhibited the proliferative response. Supernatants from suppressed spleen cell cultures were found to inhibit normal T cell mitogenesis. Taken together, these results implicated a soluble macrophage-derived suppressor factor in the down regulation of T cell proliferation after exposure to SCW in vivo. Subsequent in vitro studies to identify this suppressor molecule(s) revealed the activity to be indistinguishable from the polypeptide transforming growth factor beta (TGF-beta). Furthermore, TGF-beta was identified by immunolocalization within the spleens of SCW-injected animals. The cells within the spleen that stained positively for TGF-beta were phagocytic cells that had ingested, and were presumably activated by, the SCW. These studies document that TGF-beta, previously shown to be a potent immunosuppressive agent in vitro, also effectively inhibits immune function in chronic inflammatory lesions in vivo.     

Inhibition of cytotoxic T cell development by transforming growth factor beta and reversal by recombinant tumor necrosis factor alpha

The immunoregulatory effects of transforming growth factor beta (TGF-beta) and recombinant murine tumor necrosis factor alpha (rMuTNF-alpha) on CTL generation and activity were examined. The results demonstrate that TGF-beta, in a dose-dependent manner, inhibited CTL generation but not CTL activity. The inhibitory effects were detected only when TGF-beta was added within the first 48 h of the MLC. Little activity was seen when it was added thereafter, including the addition of TGF-beta to the cytotoxicity assay. The production of TNF-alpha, which occurs during early phases of the MLC and which is inhibited in the presence of TGF-beta, appears to have an important regulatory role, as altering the levels of TNF-alpha in an MLC can significantly influence CTL development. The inhibitory effects of TGF-beta on the MLC can be significantly reversed by the addition of rMuTNF-alpha to the cultures. These results demonstrate that TGF-beta can inhibit MLC and subsequent CTL generation at early stages of the reaction, and such inhibition may involve the suppression of TNF-alpha production.     

深低温冻存同种异体皮肤移植中转化生长因子β的表达

背景：皮肤移植和创面覆盖是创伤、烧伤治疗的重要措施。目前,同种异体皮仍然是可利用的最佳创面覆盖物。目的：通过检测转化生长因子β在深低温冻存后的大鼠同种异体皮肤移植后的表达,评估深低温贮存同种异体皮在创面上的应用。方法：取大鼠皮肤保存于低温-20℃（低温保存组）及80℃深低温（深低温保存组）,冻存1周、1,2个月后,皮片分组移植于同种大鼠背部进行实验,并以自体移植大鼠作为对照组。结果与结论：同低温保存组相比,深低温保存组转化生长因子β的表达被抑制,移植皮肤排斥反应出现时间延迟及存活时间延长,排斥反应评分也较低。同低温冻存相比,深低温冷冻保存能使同种异体移植物的转化生长因子β抗原表达降低。提示深低温冻存的异体皮作为一种创面临时覆盖物,在存在皮肤组织缺失或皮源不足的情况下具有广阔的应用前景。     

转化生长因子β1与椎间盘退变的相关研究

学术背景:转化生长因子β1是蛋白多糖及胶原代谢等椎间盘细胞外基质的重要调节物质之一,有可能在腰椎间盘退变的病理过程及基因治疗中发挥作用.目的:分析转化生长因子β1的基因结构特点及其对椎间盘细胞的作用,认识转化生长因子β1在椎间盘退变的病理过程及基因治疗方面的作用.检索策略:应用计算机检索维普中文科技期刊数据库(VIP)、清华同方中文系列等数据库1995-01/2007-10的相关文章,并限定文章语言种类为中文和"English",检索词"间盘退变,转化生长因子β1,中医药"同时检索Medline数据库1995-01/2007-10期间的相关文章,检索词"ntervertebral disc degeneration, TGF-β1, Traditional Chinise Medicine"纳入标准:① 转化生长因子β1基因结构等特点文章.② 椎间盘退变、转化生长因子β1与中医药三者关联研究.排除标准:综述文献、重复研究、Meta分析类文章.文献评价:初检得到398篇文献,纳入37篇符合标准的文献.主要介绍转化生长因子β1基因的特点、转化生长因子β1对椎间盘细胞的影响、转化生长因子β1与椎间盘退变的生物治疗及中医药对转化生长因子β1表达的影响.资料综合:目前认为转化生长因子β1可通过改善退变的椎间盘细胞活性,刺激胶原基因表达来延缓椎间盘组织的退变,对退变早期的椎间盘尤其是髓核具有修复功能,可逆转椎间盘的退变.部分中药可促进椎间盘细胞转化生长因子β1的表达但机制未明.运用转基因疗法发挥转化生长因子β1对椎间盘退变性疾病的治疗在理论上可行,但现有技术还不能把转化生长因子β1安全、有效地注入人的椎间盘.结论:转化生长因子β1的结构及基因定位已明确,转化生长因子β1是转基因疗法治疗椎间盘疾病比较理想的目的基因,但真正应用于临床仍有待组织工程学等相关领域的深入研究.在中医药领域寻找促进转化生长因子β1表达的特异性效应物,有可能为椎间盘退变性疾病的治疗提供新的途径.     

Role of transforming growth factor beta in rat bladder smooth muscle cell proliferation

Conditions associated with hypertrophy of the urinary bladder have repeatedly been associated with an increased urinary excretion of transforming growth factor (TGF) beta in both rats and patients. Because TGFbeta can have both growth-promoting and -inhibiting effects, we have studied its effects on cell growth and death in primary cultures of rat bladder smooth muscle cells. TGFbeta1, TGFbeta2, or TGFbeta3 did not cause apoptosis, but all three isoforms inhibited DNA synthesis with similar potency (EC(50) of approximately 0.1 ng/ml) and efficacy. Such inhibition was antagonized by a specific TGFbeta receptor antagonist and independent of the presence of serum. Mitogen-activated protein kinases (MAPKs) are involved in the control of cell growth, and all three TGFbeta isoforms inhibited activation of the extracellular signal-regulated kinase, c-Jun NH(2)-terminal kinase, and p38 MAPK subfamilies. Nevertheless, the inhibitory effects of the TGFbeta isoforms on DNA synthesis were not affected by presence of inhibitors of the three MAPK pathways. TGFbeta did not alter cell size as measured by flow cytometry or mitochondrial activity, an integrated measure of cell size and number. We conclude that our data do not support the hypothesis that TGFbeta is a mediator of rat bladder hypertrophy.     

Recombinant latent transforming growth factor beta 1 has a longer plasma half-life in rats than active transforming growth factor beta 1, and a different tissue distribution.

Transforming growth factor beta 1 (TGF-beta 1) is a key regulator of cell growth and differentiation. Under normal physiological conditions, it is made as a biologically latent complex whose significance is unknown. Previous work has indicated that active TGF-beta 1 has a very short plasma half-life in rats (Coffey, R. J., L. J. Kost, R. M. Lyons, H. L. Moses, and N. F. La-Russo. 1987. J. Clin. Invest. 80:750-757). We have investigated the possibility that latent complex formation may extend the plasma half-life of TGF-beta 1 and alter its organ distribution. Radiolabeled latent TGF-beta 1 was formed by noncovalent association of 125I-TGF-beta 1 with the TGF-beta 1 precursor "pro" region from recombinant sources. TGF-beta 1 in this latent complex had a greatly extended plasma half-life (greater than 100 min) in rats compared with active TGF-beta 1 (2-3 min). Whereas active TGF-beta 1 was rapidly taken up by the liver, kidneys, lungs, and spleen and degraded, TGF-beta 1 in the latent complex was largely confined to the circulation, and was less than 5% degraded after 90 min. The pharmacokinetics of TGF-beta 1 in the latent complex were shown to be critically dependent on the degree of sialylation of the complex. The results suggest that formation of latent complexes may switch endogenous TGF-beta 1 from an autocrine/paracrine mode of action to a more endocrine mode involving target organs distant from the site of synthesis.     

Role of transforming growth factor beta in conjunctival scarring

Glaucoma is the major cause of irreversible blindness throughout the world. Of all of the treatments that are available at present, the most effective appears to be surgery; however, excessive conjunctival scarring can lead to surgical failure. In the last decade, the introduction of the anti-metabolites mitomycin-C and 5-fluorouracil as anti-scarring treatments have greatly improved the results of glaucoma surgery, but these agents are associated with complications that can potentially result in blindness. A possible target for a more physiological approach to anti-scarring is transforming growth factor beta. This review examines the role of transforming growth factor beta in conjunctival scarring and discusses promising new ways of modifying its activity.     

大鼠转化生长因子β1基因转染成骨细胞后的表达

背景：将生物材料复合细胞因子基因，或复合转入细胞因子基因的细胞植入骨缺损处可以促进骨修复。目的：观察将大鼠转化生长因子β1基因转染成骨细胞后进行骨缺损基因治疗的可行性。设计：对照观察实验。单位：华中科技大学同济医学院附属协和医院骨科。对象：新生SD大鼠5只，雌雄不限。方法：实验于2000—02／09在华中科技大学同济医学院附属协和医院骨科实验室完成。通过脂质体介导将转化生长因子β1基因导人大鼠成骨细胞，并以质粒pcDNA，转染细胞作为对照。转染24h后通过链霉亲和素-生物素化过氧化物酶复合物法和原位杂交法检测目的基因瞬时表达的情况。采用G418筛选转染细胞2周，获得阳性细胞克隆，用链霉亲和素-生物素化过氧化物酶复合物法检测转染细胞稳定表达转化生长因子β1的情况。主要观察指标：链霉亲和素-生物素化过氧化物酶复合物法和原位杂交法检测转染细胞基因表达情况。结果：①pcDNA3-TGF-β1转染成骨细胞瞬时表达组化检测和原位杂交检测结果：24h转染成骨细胞胞浆中充满染色的棕黄色颗粒，对照组空载体转染细胞腧浆中刚没有棕黄倘．颗粒．说明转基因细胞中转化毕长因子β1 mRNA明显增高。②G418筛选转基因细胞组化检测：G418筛选2周后的转染细胞仍然有较高的转化生长因子β表达。结论：利用基因转染技术可使成骨细胞瞬时、高效表达细胞因子，转染后瞬时和筛选2周后，均呈现转化生长因子β1基因转染成骨细胞后稳定的高表达，说明采用细胞因子基因转染成骨细胞进行骨缺损的基因治疗具有可行性。     

大鼠转化生长因子β1基因转染成骨细胞后的表达

背景将生物材料复合细胞因子基因,或复合转入细胞因子基因的细胞植入骨缺损处可以促进骨修复.目的观察将大鼠转化生长因子β1基因转染成骨细胞后进行骨缺损基因治疗的可行性.设计对照观察实验.单位华中科技大学同济医学院附属协和医院骨科.对象新生SD大鼠5只,雌雄不限.方法实验于2000-02/09在华中科技大学同济医学院附属协和医院骨科实验室完成.通过脂质体介导将转化生长因子β1基因导入大鼠成骨细胞,并以质粒pcDNA3转染细胞作为对照.转染24 h后通过链霉亲和素-生物素化过氧化物酶复合物法和原位杂交法检测目的基因瞬时表达的情况.采用G418筛选转染细胞2周,获得阳性细胞克隆,用链霉亲和素-生物素化过氧化物酶复合物法检测转染细胞稳定表达转化生长因子β1的情况.主要观察指标链霉亲和素-生物素化过氧化物酶复合物法和原位杂交法检测转染细胞基因表达情况.结果①pcDNA3-TGF-β1转染成骨细胞瞬时表达组化检测和原位杂交检测结果24 h转染成骨细胞胞浆中充满染色的棕黄色颗粒,对照组空载体转染细胞胞浆中则没有棕黄色颗粒,说明转基因细胞中转化生长因子β1 mRNA明显增高.②G418筛选转基因细胞组化检测G418筛选2周后的转染细胞仍然有较高的转化生长因子β1表达.结论利用基因转染技术可使成骨细胞瞬时、高效表达细胞因子,转染后瞬时和筛选2周后,均呈现转化生长因子β1基因转染成骨细胞后稳定的高表达,说明采用细胞因子基因转染成骨细胞进行骨缺损的基因治疗具有可行性.