不同生长因子对人胚胎脑海马区神经干细胞体外生长的影响

本研究比较了不同生长因子对人胚胎脑海马区神经干细胞(NSCs)体外生长的影响。取胎龄8 ~12周的人胚胎脑海马,按基础培养液中生长因子的不同将培养细胞分为:h EGF组、h -bFGF组、h- EGF+h -bFGF组、h -EGF+h LIF组、h- bFGF+h LIF组、h -EGF+h bFGF+h- LIF组,将机械分离的单细胞悬液以2×106 个细胞/ml接种到上述组别,观察各生长因子的作用。结果发现,原代细胞培养6h时,各组细胞情形区别不大;随培养时间的延长,h -EGF+h- bFGF组和h- -EGF+h -bFGF+h- LIF组先形成许多小细胞簇, 7d后,这两组的细胞球数量进一步增加;而h- bFGF+h -LIF组和h- EGF+h -LIF组有较少的细胞球,h bFGF组偶见小细胞球,h- EGF组细胞几乎全部死亡。将培养的h -EGF+h -bFGF+h -LIF组的神经干细胞用1%FBS诱导分化后,行免疫细胞化学鉴定。结果表明,诱导分化12h时,细胞呈RNA 结合蛋白(Musashi1)阳性, 7d时,细胞大部分呈βⅢ管蛋白(βⅢ- tu- bulin)和胶质原纤维酸性蛋白(GFAP)阳性,极少数为半乳糖脑苷脂(Galc)阳性。以上结果提示,人胚胎脑海马区NSCs的体外存活增殖必须依赖于h -EGF和h- bFGF的共同作用,而h- LIF对早期培养的人NSCs的影响不明显。     

人胚胎脑海马区神经前体细胞移植到损伤的成年大鼠脑内存活及迁移的研究

本文从人胚胎脑海马区分离、培养、鉴定了神经前体细胞(neuralprecursorcells,NPCs),并初步观察了大鼠纹状体海人酸(kainicacid,KA)损伤后,人神经前体细胞(hNPCs)移植到成年大鼠侧脑室和纹状体后存活和迁移的情况。取胎龄8~12周的人胚胎脑海马区细胞,用含人表皮生长因子(humanepidermalgrowthfactor,h-EGF)、人碱性成纤维细胞生长因子(humanbasicfi-broblastgrowthfactor,h-bFGF)以及人白细胞抑制因子(humanleukemiainhibitorygrowthfactor,h-LIF)的DMEM/F12培养液离体培养,以含1%胎牛血清(fetalbovineserum,FBS)的DMEM/F12诱导分化,巢蛋白(nestin)免疫荧光染色鉴定NPCs的特征。向成年大鼠右侧纹状体内立体定位注射KA,造成大鼠纹状体局部损伤。用溴脱氧尿嘧啶核苷(bromodeoxyuridine,BrdU)体外标记第2代hNPCs,48h后分别移植到正常成年大鼠和损伤大鼠手术侧的侧脑室和纹状体。6周后用抗BrdU抗体检测HNPCs的存活和迁移。结果显示:体外培养呈悬浮球状生长的细胞是nestin阳性的hNPCs。hNPCs移植6周后,在大鼠损伤侧的纹状体和侧脑室内,均检测到了BrdU阳性细胞,并有一部分细胞迁移到了周围纹状体实质和胼胝体。结果提示:体外分离培养的hNPCs移植到成年大鼠脑损伤区周围可以存活和迁移,损伤区域可能对移植细胞的迁移有一定的诱向作用。     

Enhanced half-life of genetically engineered human IgG1 antibodies in a humanized FcRn mouse model: potential application in humorally mediated autoimmune disease

The MHC class I-like Fc receptor FcRn plays an essential role in extending the half-life (t(1/2)) of IgG antibodies and IgG-Fc-based therapeutics in the circulation. The goal of this study was to analyze the effect of human IgG1 (hIgG1) antibodies with enhanced in vitro binding to FcRn on their in vivo t(1/2) in mice expressing human FcRn (hFcRn). Mutants of the humanized monoclonal Herceptin antibody (Hu4D5-IgG1), directed against human epidermal growth factor receptor 2 (p185 (HER2)), show altered pH-dependent binding to hFcRn in vitro. Two engineered IgG1 mutants (N434A and T307A/E380A/N434A) showed a considerably extended t(1/2) in vivo compared with wild-type antibody in mice expressing an hFcRn transgene (Tg) but not in mice expressing the endogenous mouse FcRn. The efficiency of hFcRn-mediated protection was dependent on hFcRn Tg copy number. Moreover, when injected into FcRn-humanized mice at a concentration sufficient to partially saturate hFcRn, the engineered IgG1 mutants with an extended serum t(1/2) were most effective in reducing the t(1/2) of a tracer hIgG1 antibody. Finally, administration of mutant with high binding to hFcRn ameliorated arthritis induced by passive transfer with human pathogenic plasma. These results indicate that Fc regions modified for high binding affinity to hFcRn increases serum persistence of therapeutic antibodies, that the same approach can be exploited as an anti-autoimmune therapy to promote the clearance of endogenous pathogenic IgG and that FcRn-humanized mice are a promising surrogate for hIgG therapeutic development.     

Immunohistochemical localization of the epidermal growth factor receptor in normal human tissues

A monoclonal antibody recognizing an epitope of the external domain of the human epidermal growth factor (EGF) receptor was used to localize this protein in selected normal human tissues. Two patterns of reactivity were recognized: strong linear or granular cell surface staining, and granular cytoplasmic staining. In one tissue, the endometrium, a change in the reaction pattern associated with changes in hormonal stimulation was observed. In some tissues such as epididymis and skin, the antibody showed surface reactivity with cells considered to represent part of the proliferating cell compartment, whereas in liver, pancreas, and prostate, all cells were reactive with the antibody, though the predominant reactivity was localized in the cytoplasm. The differential distribution of the epidermal growth factor receptor to specific cell types and cellular compartments may signify adaptations that permit growth factor responsiveness in a milieu of available ligand.     

Interleukin-13 induces proliferation of human airway epithelial cells in vitro via a mechanism mediated by transforming growth factor-alpha.

Remodeling of the airways, as occurs in asthmatic patients, is associated with the continual presence of inflammatory mediators and Th2 cytokines, especially interleukin (IL)-13, during cycles of epithelial injury and repair. In this study, we examined the effect of IL-13 on well-differentiated normal human bronchial epithelial (NHBE) cells maintained in air-liquid interface culture. IL-13 induced proliferation of NHBE cells after 24 h exposure, as reflected by [(3)H]thymidine uptake and cell counts. The effects of IL-13 were mediated through the epidermal growth factor receptor (EGFR), as proliferation was attenuated by AG1478, an EGFR tyrosine kinase inhibitor. Proliferation appeared to be mediated by transforming growth factor (TGF)-alpha, a potent ligand for EGFR, which was released rapidly from NHBE cells in response to IL-13. Neutralizing antibody to TGF-alpha, but not antibodies against other potentially important growth factors (EGF, heparin binding epidermal growth factor-like growth factor [HB-EGF], platelet-derived growth factor [PDGF]), inhibited the mitogenic response to IL-13. This study provides the first experimental evidence that IL-13 can initiate a proliferative response of human airway epithelium in the absence of inflammatory cells or other cell types. The results are consistent with a mechanism whereby IL-13 induces release of TGF-alpha from the epithelial cells, which in turn binds via an autocrine/paracrine-type action to the EGFR, initiating proliferation. IL-13-induced airway remodeling in vivo may involve this epithelium-driven response.     

Affinity alteration of insulin receptor induced by a phorbol ester

The effect of a phorbol ester tumor promoter, 12-O-tetradecanoyl phorbol-13-acetate (TPA) on 125I-insulin binding to human cells was examined. TPA markedly inhibits insulin binding to cultured human lymphocytes and macrophages but has a minimal effect on human fibroblasts. This inhibition is temperature, time, and concentration dependent. The inhibition of insulin binding to the cells at 37 degrees C occurs within minutes and diminishes by 6 h of incubation. Insulin binding is decreased by TPA whether the phorbol ester is added before, after, or simultaneously with 125I-insulin to the cell suspension. Scatchard analysis of binding to IM-9 lymphocytes indicates that TPA affects the affinity rather than the number of insulin receptors. The phorbol ester has only a small effect on 125I-human growth hormone binding in cultured human lymphocytes. TPA perturbs the insulin receptor of cultured human lymphocytes in a fashion similar to its effect on the epidermal growth factor receptor of several other cell types. The specific mechanism of TPA action that affects the receptor of these two potent growth factors (i.e., insulin and epidermal growth factor), however, is unknown.     

Human kappa light chain targeted Pseudomonas exotoxin A--identifying human antibodies and Fab fragments with favorable characteristics for antibody-drug conjugate development

Antibody-drug conjugates (ADC) represent promising agents for targeted cancer therapy. To allow rational selection of human antibodies with favorable characteristics for ADC development a screening tool was designed obviating the need of preparing individual covalently linked conjugates. Therefore, α-kappa-ETA' was designed as a fusion protein consisting of a human kappa light chain binding antibody fragment and a truncated version of Pseudomonas exotoxin A. α-kappa-ETA' specifically bound to human kappa light chains of human or human-mouse chimeric antibodies and Fab fragments. Antibody-redirected α-kappa-ETA' specifically inhibited proliferation of antigen-expressing cell lines at low toxin and antibody concentrations. Selected antibodies that efficiently delivered α-kappa-ETA' in the novel assay system were used to generate scFv-based covalently linked immunotoxins. These molecules efficiently triggered apoptosis of target cells, indicating that antibodies identified in our assay system can be converted to functional immunoconjugates. Finally, a panel of human epidermal growth factor receptor (EGFR) antibodies was screened--demonstrating favorable characteristics with antibody 2F8. These data suggest that antibodies with potential for Pseudomonas exotoxin A-based ADC development can be identified using the novel α-kappa-ETA' conjugate.     

Epidermal ridge formation in the human fetus: A correlation to the appearance of basal cell heterogeneity and the expression of epidermal growth factor receptor and cytokeratin polypeptides in the epidermis

This paper aims to clarify an expression of epidermal growth factor receptor (EGFR) and cytokeratin 10 and/or 11 in relation to primary and secondary epidermal ridge formation of the human fetus. Firstly, scanning electron microscopy revealed heterogeneity in basal cell morphology during epidermal ridge formation. Basal cells had a uniform, smooth, and polygonal dermal surface until formation of the primary epidermal ridges. Thereafter, the dermal surface became ruffled and elliptic except at the primary epidermal ridges. Secondly, EGFR was detected by monoclonal antibody and autoradiography using ¹²⁵I-EGF. The antibody reacted with primary epidermal ridge, stratum basale, stratum intermedium, and outer layer of sweat duct. The reactivity became stronger at the primary epidermal ridge than at the secondary one. The binding of ¹²⁵I-EGF was concentrated in the primary epidermal ridge and sweat duct. Thirdly, cytokeratin 10 and/or 11, a maturation marker of keratinocytes, was detected by monoclonal antibody. The antibody reacted only with the stratum intermedium before secondary epidermal ridge formation. Afterward, it also reacted with the stratum basale of the secondary epidermal ridge but never reacted with that of primary epidermal ridge. The results indicate that basal cells of the secondary epidermal ridge enter the maturation process and suggest a localization of epidermal stem cells on the primary epidermal ridges. Concerning epidermal ridge formation, we suppose that the formation of the primary epidermal ridge causes the segregation of the epidermal stem cells, and that the increased density of the basal cells between the two primary epidermal ridges brings about the change in their dermal surface shape and the formation of the secondary epidermal ridge.     

Development of an attenuated interleukin-2 fusion protein that can be activated by tumour-expressed proteases

The ability to alter the cytokine microenvironment has the potential to shape immune responses in many physiological settings, including the immunotherapy of tumours. We set out to develop a general approach in which cytokines could be functionally attenuated until activated. We report the development and initial characterization of fusion proteins in which human or mouse interleukin-2 (IL-2), a potent growth factor for immune cells, is joined to a specific IL-2 inhibitory binding component separated by a protease site. The rationale is that upon cleavage by a protease the cytokine is free to dissociate from the inhibitory component and becomes biologically more available. We describe the successful development of two attenuation strategies using specific binding: the first uses the mouse IL-2 receptor alpha chain as the inhibitory binding component whereas the second employs a human antibody fragment (scFv) reactive with human IL-2. We demonstrated that the fusion proteins containing a prostate-specific antigen or a matrix metalloproteinase (MMP) protease cleavage site are markedly attenuated in the intact fusion protein but had enhanced bioactivity of IL-2 in vitro when cleaved. Further, we showed that a fusion protein composed of the IL-2/IL-2 receptor alpha chain with an MMP cleavage site reduced tumour growth in vivo in a peritoneal mouse tumour model. This general strategy should be applicable to other proteases and immune modulators allowing site-specific activation of immunomodulators while reducing unwanted side-effects.     

Two distinct monoclonal antibodies raised against mouse beta nerve growth factor. Generation of bi-specific anti-nerve growth factor anti-horseradish peroxidase antibodies for use in a homogeneous enzyme immunoassay

Two hybridomas producing monoclonal antibodies against mouse beta nerve growth factor (NGF) were obtained from the fusion of hyperimmune splenocytes from rats immunized with polymerized beta-NGF and Sp2/0.Ag mouse myeloma cells. The monoclonal antibodies coded IgG 24 and 30 produced and secreted by the hybrid cells are both of the IgG2a subclass. Both monoclonal antibodies are capable of recognizing native NGF coated on microassay plates as well as the denatured factor on Western blots. However, only IgG 30 has been found to block NGF-induced process outgrowth from the rat pheochromocytoma cell line (PC12) as well as NGF-induced increase in choline acetyltransferase activity in rat primary septal cell cultures. In addition, only IgG 30 was able to detect immunocytochemically NGF-immunoreactive sites in fixed tissue. And, finally, IgG 24 could not compete for IgG 30 binding to immobilized native NGF. Consequently, it appears that these antibodies are recognizing different epitopes on the NGF molecule. Neither monoclonal antibody displayed any crossreactivity with serum albumin, aprotinin, epidermal growth factor or insulin. A hybrid-hybridoma producing bi-specific anti-NGF anti-horseradish peroxidase (HRP) monoclonal antibodies was generated from the fusion of an azaguanine resistant anti-HRP hybridoma, coded RAP2.Ag and the anti-NGF IgG 30 hybridoma treated with emetine. The potential merits of using these bi-specific antibodies in combination with their mono-specific anti-NGF parent in a homogeneous sandwich immunoassay for the quantitation of NGF are discussed.     

表皮生长因子单抗对人牙龈上皮细胞增殖的影响

本文采用体外人牙龈上皮细胞 (HGEC)培养技术和四唑盐 (MTT)比色法 ,探讨表皮生长因子 (EGF)对HGEC的作用 ,并评价抗EGF受体 (EGFR)单克隆抗体对它的影响。结果发现 ,从第 2天开始 ,1μg/L的EGF对HGEC有明显的促增殖作用(P <0 0 5 ) ,第 4天起 ,促增殖非常显著 (P <0 0 1) ,并持续到第 8天 ;EGFR抗体浓度在 1∶10 0 0～ 1∶10时 ,能显著地抑制EGF对HGEC的促增殖作用 (P <0 0 1) ,1∶10 0浓度抑制作用最大 ,并持续作用至第 8天 (P <0 0 1)。从而表明 ,EGF对HGEC有显著的促增殖作用 ,这种作用能被EGFR抗体所阻断。     

Presence of growth factors in human pituitary.

Recent reports indicate that fibroblast growth factors known to be present in the pituitary in high levels regulate the action of growth hormone and prolactin. New data also suggest a regulatory role in the pituitary for other growth factors, such as epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha). Since in most systems cooperation of several growth factors is required for their optimal function, we sought to demonstrate the presence of certain growth factors in the pituitary. Acid ethanol extracts from approximately 50 autopsy-derived human pituitaries were subjected to molecular sieve chromatography and were tested for growth factors. Low molecular weight protein (10 micrograms) eluted from the molecular sieve column contained 10-20 ng material binding to the EGF/TGF-alpha receptor as determined by the EGF/TGF alpha radioreceptor binding assay which represents 11 ng EGF/TGF alpha per pituitary. By Western blotting we found EGF but could not document the presence of TGF-alpha in this material. Radioimmunoassay for insulin-like growth factor I detected 0.4-0.8 ng insulin-like growth factor-I/100 micrograms extract. TGF-beta eluted between 14,000 and 20,000 M(r) at levels of 3-4 ng/pituitary. Its ability to inhibit growth of CC164 mink lung cells was abolished by antibody to TGF-beta 1 but not by antibody raised against TGF-beta 2. The detection of platelet derived growth factor was equivocal and not fully reproducible. We have partially purified TGFe from the pituitary; it stimulated soft agar growth of carcinoma SW-13 cells, and it followed an elution pattern identical to bovine kidney TGFe on molecular sieve column and high pressure liquid and high performance electrophoretic chromatography. Our data show that in addition to fibroblast growth factors, the human pituitary contains other growth factors, such as EGF/TGF-alpha, TGF-beta, insulin-like growth factor I, and TGFe.     

Interleukin-6 and interleukin-6 soluble receptor regulate proliferation of normal, human papillomavirus-immortalized, and carcinoma-derived cervical cells in vitro.

A variety of sexually transmitted diseases frequently accompany infection with human papillomavirus and stimulate inflammation of the cervical mucosa. Inflammation and cell injury cause release of proinflammatory cytokines, which in turn might regulate growth of human papillomavirus-infected cells. This study compared the interaction of the proinflammatory cytokine, interleukin-6 (IL-6), and its soluble receptor with normal ecto- and endocervical cells, human papillomavirus-immortalized ectocervical cells, and squamous carcinoma-derived cell lines. Proliferation of normal cervical cells was enhanced by IL-6 but inhibited by its soluble receptor. However, both IL-6 and its soluble receptor significantly stimulated growth of the three immortal and four cervical carcinoma-derived cell lines analyzed. Stimulation by IL-6 was dose dependent and was blocked by an antibody that neutralized IL-6 activity. IL-6-mediated proliferation was accompanied by increased expression of RNAs encoding transforming growth factor-alpha and amphiregulin, two epidermal growth factor receptor ligands. Furthermore, growth stimulation by IL-6 was significantly inhibited by antibodies that either blocked signal transduction by the epidermal growth factor receptor or that neutralized transforming growth factor-alpha or amphiregulin activity. Thus, IL-6 stimulates proliferation of human papillomavirus-immortalized cervical cells via an epidermal growth factor receptor-dependent pathway involving autocrine stimulation by transforming growth factor-alpha and amphiregulin.     

Separation of transforming growth factors TGF alpha and TGF beta during chromatographic purification of extracts from mouse C-234 tumors

Polypeptide transforming growth factors: TGF alpha and TGF beta were isolated and separated from acidic ethanol extracts of mouse C-243 tumors. The purification of the acid-soluble extract was achieved by Bio-Gel P-60 filtration chromatography, followed by CM-Sepharose CL-6B ion exchange, Bio-Gel P-10 filtration, and dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). At the Bio-Gel P-10 purification step, TGF alpha was separated from TGF beta. TGF alpha stimulated mouse Balb/c-3T3, rat NRK-49F and human A549 cells to form colonies in soft agar, and competed with 125I-labeled epidermal growth factor (EGF) for binding to human placenta membrane receptors. Over 40,000-fold SDS-PAGE-purified TGF beta had an Mr of 25,000. Unlike TGF alpha, TGF beta stimulated the clonal growth of NRK fibroblasts only in the presence of the suboptimal amounts of EGF (0.5 ng/ml). TGF beta significantly inhibited the anchorage-independent growth of malignant human lung carcinoma A549 cells, and in the radioreceptor assay with 125I-EGF it had no affinity to EGF receptors.     

Conditioned medium from mouse sarcoma 180 cells contains vascular endothelial growth factor

Medium conditioned by mouse sarcoma 180 cells stimulates the growth of capillary endothelial cells. The growth factor produced by mouse sarcoma 180 cells is heparin-binding, dithiothreitol-sensitive, endothelial cell specific, and secreted into the medium. The characteristics of this mouse sarcoma-derived growth factor are very similar to those of vascular endothelial growth factor (VEGF) first described by Ferrara and Henzel (1989). The N-terminal amino acid sequences of the two growth factors are similar. Since the amino acid sequence of vascular permeability factor (VPF) is essentially identical to that of VEGF, a Western blot of mouse sarcoma 180-derived endothelial growth factor was probed with a polyclonal antibody raised against human VPF. This antibody reacted with several proteins of approximately 23 kDa, suggesting the presence of multiple forms of a VEGF-like protein. A full length cDNA probe for bovine VEGF reacted strongly with RNA isolated from mouse sarcoma 180 cells. We conclude that an endothelial growth factor found in conditioned medium from mouse sarcoma 180 cells is VEGF.     

Aberrant expression of the human epidermal growth factor receptor 2 oncogene is not a common feature in osteosarcoma

Human epidermal growth factor receptor 2 expression in osteosarcoma and its relationship to prognosis have been the subject of several conflicting reports, most of them relying on immunohistochemical studies. Because the urgent need of prognostic markers and effective new treatment options for osteosarcoma patients, we evaluated the role of human epidermal growth factor receptor 2 in 2 well-characterized sets of pretherapeutic osteosarcoma samples (46 paraffin-embedded and 46 fresh-frozen biopsy samples) using immunohistochemistry with 2 different antibodies [DAKO A0485 (Glostrup, Denmark) and Novocastra CB11 (Newcastle, UK)] as well as fluorescence in situ hybridization, real-time polymerase chain reaction, and SNP array analyses and correlated our findings with clinicopathological parameters. However, our study failed to detect unequivocal evidence of human epidermal growth factor receptor 2 gene amplification or overexpression of human epidermal growth factor receptor 2 messenger RNA or protein in any of the investigated tumors. Only in a small subset of samples, a moderate increase in messenger RNA levels (13.6%) or focal membranous immunoreactivity (8.7%; A0485) was detected but did not correlate with survival or response to chemotherapy. Cytoplasmic staining was identified more frequently (63%; CB11) but again did not show any association with clinicopathological parameters. In conclusion, our study does not support a role for human epidermal growth factor receptor 2 as a prognostic marker in osteosarcoma.     

细胞因子抗体芯片筛选脓毒症患者血清蛋白标记物

目的:应用细胞因子抗体芯片(CAC)分析脓毒症患者血清中炎性细胞因子表达的变化.方法:采用CAC技术平台,测定9例脓毒症患者(脓毒症组)和4例正常健康人(对照组)血清中79种细胞因子.以两组样本中每组至少有4个样品的杂交信号值超过400为有表达的蛋白,筛选出有表达的蛋白38个.应用芯片差异显著性分析(SAM)软件对38个有表达的蛋白进行差异表达分析.结果:筛选到胰岛素样生长因子结合蛋白-1( IGFBP-1)、肝细胞生长因子(HGF)、骨桥蛋白(Osteopotin)、胰岛素样生长因子结合蛋白-4(IGFBP-4)、干扰素诱导蛋白-10( IP-10)和B淋巴细胞趋化因子(BLC)等6个在脓毒症患者血清中高表达的细胞因子,以及血小板衍生生长因子-BB(PDGF-BB)、脑源性神经营养因子(BDNF)、巨噬细胞炎性蛋白(MIP-11β)、白细胞介素-8( IL-8)、表皮生长因子(EGF)和白介素-1β(IL-1β)等6个在脓毒症患者血清中低表达的细胞因子.聚类分析表明,上述这12个差异表达的细胞因子能够区分脓毒症和正常健康对照.结论:CAC是一种高效、快速的蛋白质组学技术,利用CAC从血清中筛选脓毒症相关蛋白分子标记物是可行的.     

Growth suppression of hybrids between transformed cells and normal fibroblasts in serum-free medium: Correlation with retention of human chromosomes

Somatic cell hybrids formed by crossing PG19 mouse melanoma cells with mouse embryo fibroblasts have a reduced ability to proliferate in growth factor-unsupplemented serum-free medium relative to the parental melanoma cells. The suppression of growth of the hybrid cells in serum-free medium is attributable to a strict requirement of these cells for polypeptide growth factors (insulin plus platelet-derived growth factor, fibroblast growth factor, or epidermal growth factor). In contrast, the parental melanoma cells are able to grow without exogenously added growth factors. Fifteen hybrids derived from crosses between mouse L cells and normal human skin fibroblasts also have been tested for ability to grow in growth factor-unsupplemented serum-free medium. Depending on which human chromosomes are retained, growth of these hybrids in serum-free medium is also suppressed relative to growth of the L cell parent. There appear to be several genes on different chromosomes that are involved in suppression of serum-free growth of the fibroblast × L cell hybrids. One weak suppressor gene appears to be on the human X chromosome.     

抗大鼠视网膜神经节细胞诱向因子单克隆抗体的产生及其鉴定

视网膜神经节细胞诱向因子(RGNTF,Retinal ganglion neuronotrop-bic factor)是新发现的一神经生长因子,它能特异性地促进体外培养的视网膜神经节细胞的生长活性。作者用Phast System电泳实验证实,RGNTF的分子量为30kD蛋白质。并用之免疫动物,制备出3株抗RGNTF单克隆抗体,分别命名为A1、D3、E8。经鉴定杂交瘤细胞系染色体大多为端着丝点染色体,并有1～2条中部着丝点染色体。抗体的亚型鉴定A1、D3是IgG3型、E8是IgM型。用ELISA方法测定RGNTF-McAb与相应抗原结合的效价为10~(-4)～10~(-5)结果发现随抗体浓度的增强,抗体与抗原结合率也增加,以E8株结合率最高。本单抗与其它生长因子MGF、EGF无交叉反应,表明其相应抗原是一种新的生长因子。     

Collagenous matrices as release carriers of exogenous growth factors

We have investigated the use of natural and synthetic collagenous matrices as carriers of exogenous growth factors. A bladder acellular matrix (BAM) was processed from rat bladder and compared with sponge matrix of porcine type 1 collagen. The lyophilized matrices were rehydrated by the aqueous solutions of basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), platelet derived growth factor-BB (PDGF-BB), vascular endothelial growth factor (VEGF), insulin like growth factor-1 (IGF-1) and heparin binding epidermal growth factor-like growth factor (HB-EGF), to obtain the matrix incorporating each growth factor. The rehydration method enabled the growth factor protein to distribute into the matrix homogeneously. In vivo release test in the mouse subcutis revealed that, the property of BAM for growth factor release was similar to that of collagen sponge. Among the growth factors examined, bFGF release was the most sustained, followed by HGF and PDGF-BB. bFGF released from the two matrices showed similar in vivo angiogenic activity at the mouse subcutis in a dose-dependent manner. These findings demonstrate that the collagenous matrices function as release carriers of growth factors. This feature is promising to create a scaffold, which has a nature to control the tissue regeneration actively.