改构酸性成纤维细胞生长因子对小鼠胸腺细胞凋亡的影响

目的:探讨改构酸性成纤维细胞生长因子(MaFGF)对地塞米松(DEX)诱导小鼠胸腺细胞凋亡的保护作用。方法:利用地塞米松诱导小鼠胸腺细胞凋亡的模型,将实验分为空白对照组、DEX处理组、aFGF+DEX组和MaFGF+DEX组,采用DNA琼脂糖凝胶电泳和流式细胞仪两种分析方法,测定MaFGF对小鼠胸腺细胞凋亡的影响。结果:空白对照组凋亡率为168%,DEX处理后小鼠胸腺细胞出现明显的细胞凋亡,凋亡率为196%,aFGF和MaFGF处理后的细胞凋亡受到抑制,凋亡率分别下降到1595%和1293%,MaFGF效应强于aFGF,且以剂量依赖方式发挥作用。结论:MaFGF具有以剂量依赖方式的胸腺细胞保护作用。     

FGF-1改构体对小鼠脾细胞增殖与凋亡的影响

目的:主要探讨非促分裂改构人酸性成纤维细胞生长因子(MrhFGF-1)对balb／c小鼠脾细胞增殖和凋亡的影响。方法:采用3H-TdR掺入的方法检测MrhFGF-1同野生型hFGF-1相比对脾细胞增殖的影响,实验组分为(1)对照组;(2)FGF-1处理组;(3)ConA处理组;(4)FGF-1+ConA处理组。用流式细胞仪定量分析MrhFGF-1对脾细胞的凋亡保护作用,(1)对照组;(2)DEX处理组;(3)DEX+FGF-1(hFGF-1、rhFGF-1、MrhFGF-1)处理组。结果表明,利用DNA重组技术构建的一种非促分裂FGF-1突变体MrhFGF-1和野生型FGF-1相比,对脾细胞的促细胞增殖能力明显降低,但其对细胞凋亡保护作用的影响并无显著性变化,说明FGF-1的促细胞增殖能力和细胞凋亡保护信号通路并非由同一信号通路来实现的。     

The Inhibitory Effect of Staphylococcus epidermidis Slime on the Phagocytosis of Murine Peritoneal Macrophages Is Interferon-Independent

The extracellular slime produced by Staphylococcus epidermidis has been shown to interfere with several human neutrophil functions in vitro, such as chemotaxis, degranulation and phagocytosis. Slime production has been suggested as a useful marker for clinically significant infections with coagulase-negative Staphylococcus. Since the main role of macrophages in defense mechanisms is phagocytosis, the effect of slime on the phagocytic activity of macrophages was investigated. The phagocytic activity of murine peritoneal macrophages treated with slime in vitro decreased in a dose-dependent fashion. A similar decrease was also observed in macrophages isolated from mice that had previously received intraperitoneal injection of slime. To investigate whether interferon also plays a role in this process, mice were treated with interferon or an interferon inducer, polyinosinic-polycytidylic acid (poly I:C), together with slime before macrophage isolation. The slime-suppressed phagocytic activity of macrophages was partially relieved by both agents, and the recovery effect of poly I:C in slime-suppressed phagocytosis of macrophages in vivo might be attributed to the increased interferon level in peritoneal fluid and sera. However, when slime was given to poly I:C-pretreated mice, the phagocytic activity remained suppressed. Thus, it appears that slime is able to suppress the phagocytic activity of macrophages regardless of the state of macrophage activation by poly I:C. The results suggest that the inhibition of phagocytosis by S. epidermidis slime may be independent from the activation of interferon.     

成纤维细胞生长因子与其受体相互作用及其抑制剂的研究进展

成纤维细胞生长因子（FGF）有许多重要的生理功能，并与肿瘤的形成有关.为了弄清FGF与成纤维细胞生长因子受体(FGFR)相互作用的机制，人们对FGF和FGFR的各个结合结构域进行了深入、细致的研究，定位了aFGF、bFGF的肝素结合区、bFGF的受体结合区、FGF受体的肝素结合区、配体结合区和FGF受体相互结合区，提出了两个FGF与FGFR相互作用的模型，在此基础上设计了FGF的核酸类、糖类和多肽类抑制剂，为寻找新一代抗癌药物打下了理论基础.     

aFGF荧光分子探针基因的合成及其表达

通过PCR技术和体外DNA重组技术将绿色荧光蛋白cDNA和酸性成纤维细胞生长因子cDNA构建成融合基因,克隆到表达载体pET3c中,构建成表达菌株BL21(DE3)/pET3c-GAF。经IPTG诱导表达,融合蛋白的表达量占菌体总蛋白的25%;DNA测序结果表明设计合成的融合基因与预期相符。Westernblot结果表明重组蛋白具有aFGF的免疫原性。经IPTG诱导后的菌体及蛋白粗提液在荧光显微镜下可观察到强烈的绿色荧光。融合基因在大肠杆菌中实现了表达,用MTT法测得纯化融合蛋白与野生型aFGF促Blab/c3T3细胞增殖活性相当,为利用绿色荧光分子探针研究aFGF的在活体内的作用机制建立了新的方法。     

两种类型人碱性成纤维生长因子的体外稳定性研究

目的:通过野生型bFGF和改构型bFGF蛋白溶液中的聚集过程的比较,初步探讨bFGF在水溶液中发生聚集的机理。方法:在选择合适溶液体系后,采用蛋白溶解度和促有丝分裂活性能力为指标,表征野生型bFGF和突变型bFGF聚合程度,分析共价聚合和非共价聚合所起的作用。结果:在相同的溶液体系中,野生型bFGF的聚集程度高于突变型bFGF。bFGF聚合程度与浓度有依赖性。沉淀结果分析非共价聚合占主要作用。结论:野生型bFGF在溶液中共价聚和和非共价聚合同时发生,两种bFGF聚合过程中非共价聚合占较大比例。突变半胱氨酸可以减少聚合发生。     

FGF-1及其突变体的研究进展

酸性成纤维细胞生长因子(acid fibroblastgrowth factor,FGF-1)是成纤维细胞生长因子家族重要的成员之一。来源于三个胚层的细胞都能够表达FGF-1,其生物学效应非常广泛,在组织和器官发育、血管发生、血细胞生成、肿瘤发生和伤口愈合等方面发挥着重要作用。FGF-1不但可以在细胞外通过胞内信号通路而且也可以进入细胞内部发挥生物学功能。主要综述FGF-1信号转导方面的研究进展、促细胞增殖作用的可能分子机制以及将来的临床应用前景。     

Basic Fibroblast Growth Factor (bFGF) Immunoreactivity Is Present in Chromaffin Granules

Basic fibroblast growth factor (bFGF) has recently been isolated from bovine adrenal glands. Immunohistological data revealed its presence in both adrenal cortex and adrenal medulla. Using immuno-electronmicroscopy, we found that in medullary chromaffin cells bFGF-immunoreactivity is localized in the secretory granules. Immunoreactivity also was observed by electronmicroscopy in isolated granules. Western blot analysis revealed the presence of the typical 18-kDa bFGF and additional immunoreactive materials with molecular masses of approximately 24, 30, and 46 kDa in whole bovine adrenal, and in cortex and medulla. Similar results were obtained with proteins from bovine chromaffin granules, with the following two exceptions: the 46-kDa immunoreactivity was found to be highly enriched when compared with medulla or cortex, and the 18-kDa band could be detected with only an antiserum against a synthetic peptide comprising the 24 NH2-terminal amino acids of bFGF, and not with an antiserum against purified bovine pituitary bFGF. All fractions enriched for bFGF-immunoreactivity showed neurotrophic activity for chick ciliary ganglion neurons, which could be blocked by antibodies. These results demonstrate for the first time the localization and occurrence of bFGF in a cellular secretory organelle, and present further evidence for the existence of higher molecular weight immunoreactive forms of bFGF.     

FGF21类似物治疗动脉粥样硬化机制研究进展

动脉粥样硬化是冠心病、脑梗死、外周血管病的主要诱因,近年来发病率越来越高,严重威胁着人类生命健康。脂质代谢障碍是动脉粥样硬化的病理基础。成纤维细胞生长因子21(FGF21)是FGF家族的一种内分泌因子,它能够增加葡萄糖的摄取,调节脂质代谢,并使代谢活跃的器官（如肝脏和脂肪组织）中胰岛素反应敏感。FGF21水平与动脉粥样硬化的发生率和严重程度密切相关。然而,FGF21原型在血浆的半衰期短、体外易聚集,严重限制了其临床应用。近年来,对FGF21类似物的研究取得了突破性进展。综述了FGF21的生理作用,并讨论了基于FGF21类似物治疗动脉粥样硬化的主要突破和局限性,为FGF21蛋白类新药的开发提供了理论依据。     

肽类生长因子对游离细胞核中拓扑异构酶活性的影响

将表皮生长因子及神经生长因子分别作用于分离所得的鼠胚成纤维细胞及其转化细胞的细胞核。结果表明表皮生长因子可以提高游离细胞核中ＤＮＡ拓扑异构酶的活性,但神经生长因子无此作用。     

牛脑成纤维细胞生长因子的分离纯化与鉴定

新鲜牛脑组织匀浆液经两步硫酸铵沉淀、CM-Sephadex C50 离子交换层析以及肝素-Sepharose 亲和层析,可得到纯化的酸性和碱性成纤维细胞生长因子(aFGF 和bFGF),分子量分别为13.2kD 和15.2—15.8kD.两种因子均可有效促进3T3细胞的 DNA 合成,ED₅₀分别为15.8ng/ml 和 0.32ng/ml.进一步对 aFGF 的等电点及氨基酸组成做了分析.     

bFGF对热射病大鼠模型ET21, VWF, VEGF水平的影响

目的:研究碱性成纤维细胞生长因子(basic Fibroblast growth factor,bFGF)对热射病大鼠模型ET21、VWF、VEGF水平的影响。方法:选择SD大鼠36只随机分为6组,每组个6只,建立热射病大鼠模型后分为模型组、常温组、常温bFGF组、降温组、降温bFGF组,建模后0 h、2 h,比较不同级别生命征、血管功能因子。结果:建模后0 h,模型组Tr、HR、RR、MAP明显高于空白组(P0.05);建模后2 h,常温bFGF组、降温bFGF组明显低于常温组、降温组,降温组、降温bFGF组明显低于常温组、常温bFGF组(P0.05)。建模后0 h,模型组ET21、VWF的含量均明显高于空白组,VEGF含量明显低于热空白组(P0.05);建模后2 h,常温组ET21、VWF明显升高,VEGF明显降低,常温bFGF组、降温组、降温bFGF组ET21、VWF明显降低,VEGF明显升高(P0.05);降温组、降温bFGF组ET21、VWF明显低于常温组、常温bFGF组,VEGF明显高于常温组、常温bFGF组(P0.05);常温bFGF组、降温bFGF组ET21、VWF明显低于常温组与降温组,VEGF明显高于常温组与降温组(P0.05)。结论:碱性成纤维细胞生长因子有助于改善热射病大鼠生命体征,调节血管功能因子含量,提高热射病大鼠预后。     

Epidermal Growth Factor and Basic Fibroblast Growth Factor Have Independent Actions on Mesencephalic Dopamine Neurons in Culture

Abstract: Epidermal growth factor (EGF) and basic fibro-blast growth factor (bFGF) are both trophic for dopamine neurons s in cultures of dissociated embryonic rat mesen-cephaion, but the significance of this apparent overlap in neurotrophic activity is not yet known. In this study, we investigated the mechanisms of action of these two growth factors and the potential relationship between them, Using a nuclease protection assay, we determined that bFGF mRNA was expressed in the cultures. Double-label immunocytochemistry revealed that bFGF immunore-active material could be detected in tyrosine hydroxylase immunoreactive neurons and glial fibrillary acidic protein immunoreactive astrocytes. EGF treatment increased bFGF mRNA content per culture dish. As we have previously demonstrated that EGF exerts its dopaminergic neurotrophic activity via an intermediate cell type, studies were designed to address whether the pathway by which EGF acts on dopaminergic neurons is mediated by the release of bFGF. However, the trophic action of EGF on dopamine neurons, represented by high-affinity neuronal dopamine uptake, could not be blocked by immunoneutralization of bFGF, suggesting that the actions of EGF were not mediated by bFGF release. The time course of the effects of EGF and bFGF on dopamine uptake were similar, with significant increases detectable only after 5 days in culture. Both growth factors were active in the picomolar-to-nannomolar range with maximal trophic activity between 0.4 and 2.5 n M. EGF, however, was the more potent mitogen under these conditions. When cultures were simultaneously incubated with maximal concentrations of EGF and bFGF, the effect on dopamine uptake was significantly greater than with either growth factor alone and, in fact, approximated the sum of the individual effects. On the basis of these results we conclude that these growth factors have independent effects on dopamine neurons of the mesencephalon.     

Localization of Basic Fibroblast Growth Factor in Bovine Adrenal Chromaffin Cells

Abstract: We have investigated basic fibroblast growth factor (FGF-2) localization in and release from isolated bovine adrenal chromaffin cells. In contrast to previous reports, we found no evidence of fibroblast growth factor (FGF) storage in catecholamine-containing chromaffin granules. Subcellular fractionation studies did not show enrichment of FGF-2 immunoreactivity in granules, and cholinergic stimulation failed to release FGF-2 into the medium. Our results suggest that adrenal chromaffin cells resemble other FGF-2-synthesizing cell types with respect to FGF storage and secretion.     

Developmental Changes in Distribution of Acidic Fibroblast Growth Factor in Rat Brain Evaluated by a Sensitive Two-Site Enzyme Immunoassay

We developed a sensitive two-site enzyme immunoassay (EIA) system for acidic fibroblast growth factor (aFGF), using a polyclonal antibody raised in rats. This assay is based on the sandwiching of the antigen between anti-aFGF antibody immunoglobulin G (IgG) coated on plates and biotinylated anti-aFGF antibody IgG; the detection of biotinylated IgG was performed by enzyme reaction of streptavidin-conjugated beta-D-galactosidase (beta-D-galactoside hydrolase; EC 3.2.1.23). Our system was specific for aFGF, because basic fibroblast growth factor, which shares a 55% homology of amino acid sequence with aFGF, hardly cross-reacted at all. The sensitivity of this system (0.2 ng/ml) enabled us to quantify endogenous immunoreactive aFGF in the CNS. Using this two-site EIA system, we examined the levels of aFGF in various regions of rat brain and their developmental changes. At the early stage of neonatal development, i.e., 2 days after birth, all brain regions registered low aFGF levels (less than 10 ng/g tissue). However, at the young adult stage (21- to 49-day-old animals), an extremely high level of aFGF (75-90 ng/g tissue) was found in the ponsmedulla; relatively high levels (30-40 ng/g tissue) were found in the diencephalon and mesencephalon; and comparatively low aFGF levels (5-15 ng/g tissue) were found in various other brain regions such as the frontal cortex, piriform cortex, hippocampus, olfactory bulb, cerebellum, and striatum. This marked change in the regional distribution of aFGF in the rat brain during postnatal development from 2 to 21 days after birth suggests that this factor plays a significant role in the brain during this period.     

人碱性成纤维细胞生长因子突变体的高效表达

用PCR法将人碱性成纤维细胞生长因子(hbFGF)基因中编码第25、69和92位的半胱氨酸(Cys)密码子突变为丝氨酸(Ser)密码子,将突变的hbFGFcDNA片断与表达质粒pET3c连接,构建重组质粒pET3chbFGFSer25,69,92。hbFGFSer25,69,92在大肠杆菌BL21(DE3)中的表达量大于30%。通过阳离子交换和肝素亲和层析两步纯化,得到纯度大于95%的hbFGFSer25,69,92。MTT法测定纯化的产物活性表明,hbFGFSer25,69,92突变体促Balb/c细胞增殖的活性与野生型hbFGF相当,为下一步对hbFGFSer25,69,92突变体进行定点化学修饰打下了基础。     

Baicalein Relieves Ferroptosis-Mediated Phagocytosis Inhibition of Macrophages in Ovarian Endometriosis

Iron overload and oxidative stress have been reported to contribute to ferroptosis in endometriotic lesions. However, the possible roles of iron overload on macrophages in endometriosis (EMs) remain unknown. Based on recent reports by single-cell sequencing data of endometriosis, here we found significant upregulations of ferroptosis-associated genes in the macrophage of the endometriotic lesion. Additionally, there was an elevated expression of HMOX1, FTH1, and FTL in macrophages of peritoneal fluid in EMs, as well as iron accumulation in the endometriotic lesions. Notably, cyst fluid significantly up-regulated levels of intracellular iron and ferroptosis in Phorbol-12-myristate-13-acetate (PMA)-stimulated THP-1 cells. Additionally, high iron-induced ferroptosis obviously reduced PMA-stimulated THP-1 cells’ phagocytosis and increased the expression of angiogenic cytokines, such as vascular endothelial growth factor A (VEGFA) and interleukin 8 (IL8). Baicalein, a potential anti-ferroptosis compound, increased GPX4 expression, significantly inhibited ferroptosis, and restored phagocytosis of THP-1 cells in vitro. Collectively, our study reveals that ferroptosis triggered by high iron in cyst fluid promotes the development of EMs by impairing macrophage phagocytosis and producing more angiogenic cytokines (e.g., IL8 and VEGFA). Baicalein displays the potential for the treatment of EMs, especially in patients with high ferroptosis and low phagocytosis of macrophages.     

Epidermal Growth Factor and Basic Fibroblast Growth Factor Protect Dopaminergic Neurons from Glutamate Toxicity in Culture

Abstract: In this report we characterize the toxicity of the excitatory amino acid l -glutamate with respect to dopaminergic neurons cultured from embryonic rat mesencephalon. We also demonstrate that two growth factors, epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF), can protect these neurons from damage. Micromolar concentrations of l -glutamate, as well as agonists that specifically activate N -methyl- d -aspartate (NMDA) and non-NMDA receptors, are all toxic to dopamine neurons in a concentration-dependent manner, as reflected by decreases in high-affinity dopamine uptake and confirmed by decreases in numbers of tyrosine hydroxylase-immunoreactive neurons. Although the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione could attenuate the effects of quisqualate, treatment with this antagonist could not eliminate the effects of glutamate itself. Similarly, (±)-2-amino-5-phosphonopentanoic acid was effective against NMDA toxicity but could not protect cells from quisqualate toxicity. Thus, each type of receptor could mediate neurotoxicity independently of the other. The presence of EGF or bFGF in the culture medium conferred a relative resistance of dopaminergic neurons to glutamate and quisqualate neurotoxicity by increased glutamate transport. However, treatment of the cultures with l - trans -pyrrolidine-2,4-dicarboxylic acid, an inhibitor of glutamate transport, attenuated but did not eliminate the protective effects of both growth factors against glutamate toxicity. When cultures were incubated with conditioned medium from growth factor-treated cultures, neuroprotection was also achieved. These results suggest that both EGF and bFGF can protect neurons from neurotoxicity in culture by increasing the capacity of the culture for glutamate uptake as well as by the secretion of soluble factors into the medium.     

碱性成纤维细胞生长因子对大鼠晚期周围神经再生作用的实验研究

目的探讨外源性碱性成纤维细胞生长因子(bFGF)对晚期周围神经再生的作用.方法50只SD大鼠随机分治疗组、对照组各25只,切断右侧坐骨神经,12周后予以修复,修复术后每日分别给予bFGF和生理盐水,行神经电生理和组织学检查.结果治疗组和对照组修复处远段神经均有不同程度再生,4周时已可见到再生轴突,且治疗组多见.计量分析治疗组运动神经传导速度、神经肌肉动作电位幅值、髓鞘厚度、再生轴突直径和截面积明显优于对照组.治疗组与对照组相比,差异有显著性.结论bFGF能促进晚期周围神经再生.     

Tumor Necrosis Factor-α and Basic Fibroblast Growth Factor Decrease Glial Fibrillary Acidic Protein and Its Encoding mRNA in Astrocyte Cultures and Glioblastoma Cells

Abstract: Tumor necrosis factor-α is a pluripotent cytokine that is reportedly mitogenic to astrocytes. We examined expression of the astrocyte intermediate filament component glial fibrillary acidic protein in astrocyte cultures and the U373 glioblastoma cell line after treatment with tumor necrosis factor-α. Treatment with tumor necrosis factor-α for 72 h resulted in a decrease in content of glial fibrillary acidic protein and its encoding mRNA. At the same time, tumor necrosis factor-α treatment increased the expression of the cytokine interleukin-6 by astrocytes. The decrease in glial fibrillary acidic protein expression was greater when cells were subconfluent than when they were confluent. Thymidine uptake studies demonstrated that U373 cells proliferated in response to tumor necrosis factor-α, but primary neonatal astrocytes did not. However, in both U373 cells and primary astrocytes tumor necrosis factor-α induced an increase in total cellular protein content. Treatment of astrocytes and U373 cells for 72 h with the mitogenic cytokine basic fibroblast growth factor also induced a decrease in glial fibrillary acidic protein content and an increase in total protein level, demonstrating that this effect is not specific for tumor necrosis factor-α. The decrease in content of glial fibrillary acidic protein detected after tumor necrosis factor-α treatment is most likely due to dilution by other proteins that are synthesized rapidly in response to cytokine stimulation.