Localisation of Placenta Growth Factor (PlGF) in Human Term Placenta

Abstract

Placenta growth factor (PlGF) is a growth factor which belongs to the vascular endothelial growth factor (VEGF) family and is known to bind to the fms-like tyrosine kinase receptor (flt-1). Using Western blot analysis a 50 kDa band was identified in placental protein extract which corresponded to PlGF homodimer. Immunoreactive PlGF was localised to the vasculosyncytial membrane and in the media of large blood vessels of the placental villi, while staining within the mesenchyme was weak and diffuse. There was moderate staining for PlGF in discrete cells in the chorion and no staining in the epithelial layer of the amnion. The maternal decidual cells showed strong staining for PlGF immunoreactive protein. PlGF mRNA was predominantly expressed by the vasculosyncytial membrane of villous trophoblast, whilst there was no apparent expression of PlGF mRNA within the villous mesenchyme. These results suggest that PlGF may be an important paracrine factor for vascular endothelial cells in placental angiogenesis and an autocrine mediator of trophoblast function.     

Ontogeny of Hepatocyte Growth Factor (HGF) and Its Receptor (c-met) in Human Placenta : Reduced HGF Expression in Intrauterine Growth Restriction

Severe intrauterine growth restriction (IUGR) is characterized by abnormal placentation. Mouse gene knockout studies show that an absence of either hepatocyte growth factor (HGF) or its receptor, c-met, leads to intrauterine death secondary to severe IUGR with deficient placentation. In this study, immunocytochemistry localized HGF protein throughout placental villi across gestation, whereas c-met protein was localized only to the perivillous trophoblast and vascular endothelium. Within the IUGR placentae, a reduction in HGF immunostaining within the villous stroma was observed. HGF mRNA was strongly expressed in the perivascular tissue around the stem villous arteries throughout gestation, with weaker expression within the villous stroma and the terminal villi. c-met mRNA expression was limited to the perivillous trophoblast, particularly in the first trimester, with only a faint hybridization signal from the villous stroma. Placental mRNA expression was examined quantitatively using a ribonuclease protection assay: HGF and c-met mRNA expression increased from the first to the second trimester, reaching a zenith before decreasing again through the third trimester to term. HGF mRNA levels were significantly reduced in the IUGR placentae (P = 0.036), whereas c-met mRNA expression was within the normal range for gestation. These findings suggest that HGF derived from the perivascular tissue of stem villous arteries may play an important role in controlling normal villous development. Whereas reduced expression of HGF within IUGR placentae does not prove a causative link with abnormal villous development, the association lends support to this possibility.     

子痫前期患者白细胞介素-8和胎盘生长因子的表达及其意义

目的通过检测子痫前期患者羊水中白细胞介素-8（IL-8）和胎盘组织中胎盘生长因子（PLGF）的水平．以探讨IL-8和PLGF在子痫前期发病中的作用。方法采用酶联免疫吸附法和免疫组化法分别测定20例轻度子痫前期患者,22例重度子痫前期患者和30例正常妊娠妇女（对照组）的羊水中IL-8和胎盘组织中的PLGF的表达。结果子痫前期轻度组及重度组羊水IL-8均高于对照组,重度组羊水IL-8显著高于轻度组,差异均有统计学意义（P〈0．01）。子痫前期轻度组及重度组胎盘组织中PLGF表达量均低于对照组（P〈0．01）,而重度组低于轻度组（P〈0．01）。子痫前期轻、重度组及对照组胎盘组织中PLGF表达量与羊水IL-8水平均呈高度负相关（P〈0．01）。结论子痫前期患者羊水中IL-8增高,而胎盘组织中PLGF表达下降,可能与子痫前期的发生、发展有关。     

Localization of Hepatocyte Growth Factor and Its Receptor met in Endocrine Cells and Related Tumors of the Gut and Pancreas: An Immunohistochemical Study

Hepatocyte growth factor (HGF), a stimulator of angiogenesis and cell migration, regulates the growth of a wide variety of cells by binding to its high-affinity receptor met and is involved in the growth and aggressiveness of several tumors. In this study we investigated the expression of HGF and met in normal endocrine cells and related neoplasms of the gut and pancreas to verify their possible role in tumor pathogenesis, growth, and aggressiveness. Normal tissues and 60 different endocrine tumors were immunostained using specific antibodies directed against HGF, met, and various hormones. HGF immunoreactivity (IR) was found in antroduodenal G cells, rectal enterochromaffin (EC) cells, and pancreatic A and B cells, whereas met IR was detected in antral EC and G cells, and in pancreatic B cells; 46 of 60 tumors examined were positive for HGF, and they were mainly represented by ECL-, EC-, and L-cell neoplasms. met IR was identified in 50/60 tumors of various phenotypes. HGF and met coexpression was found in 42/60 cases, most of which were represented by EC-cell tumors. HGF/met coexpression was significantly more frequent in ileolonic EC-cell tumors, which in the majority of cases were malignant, than in appendiceal EC-cell tumors, which were all benign. Our results demonstrated, for the first time, that HGF and met are specifically distributed in normal gut and pancreatic endocrine cells and, in addition, suggest that HGF and met may be implicated as autocrine/paracrine factors regulating the growth of gastroenteropancreatic endocrine tumors, mainly of ileocolonic EC-cell carcinoids.     

肝细胞生长因子激活因子抑制因子-1的表达与功能

肝细胞生长因子激活因子抑制因子-1(hepatocyte growth factor activator inhibitor type1,HAI-1)是一种Kunitz型丝氨酸蛋白酶抑制因子,定位于细胞的基底侧,具有膜型和分泌型两种形式,能有效抑制肝细胞生长因子激活因子HGFA和丝氨酸蛋白酶Matriptase的活性,参与HGF/c-Met信号传导途径调节。HAI-1在包括妊娠、再生及肿瘤等各种正常生理及病理状态下均有不同水平的表达,其表达水平的变化以及其与靶蛋白酶表达比例的变化直接影响到靶蛋白酶的活性,从而在调控个体发育、血管生成、组织损伤修复以及抑制肿瘤的侵袭性生长等生理和病理过程中发挥重要作用。     

促肝细胞生长素诱导人肝癌细胞（BEL－7402）凋亡

本文从细胞学、ＤＮＡ凝胶电泳、流式细胞术三方面研究促肝细胞生长素（ｐＨＧＦ）在体外对肝癌细胞增殖活性的影响。结果表明：ｐＨＧＦ对肝癌ＢＥＬ－７４０２细胞增殖有抑制作用，并存在剂量和时间相关性。其４８ｈ的半数抑制浓度（ＩＤ５０）为０．３７ｍｇ／ｍｌ±０．０４ｍｇ／ｍｌ。而３７℃灭活的ｐＨＧＦ对ＢＥＬ－７４０２细胞增殖在１５ｈ无抑制作用，在２４和４８ｈ抑制作用很弱（ＩＤ５０＞１．５ｍｇ／ｍｌ）。ＤＮＡ凝胶电泳结果表明，ｐＨＧＦ可诱导ＢＥＬ７４０２细胞产生细胞凋亡（Ａｐｏｐｔｏｓｉｓ）。流式细胞术（ＦＣＭ）结果显示：ｐＨＧＦ抑制ＢＥＬ－７４０２细胞增殖过程是先使细胞停留在Ｇ０／Ｇ１期，继而诱导细胞产生凋亡。后两项结果均显示ｐＨＧＦ对人肝癌细胞凋亡的诱导里时间和剂量相关性。     

促肝细胞生长素诱导人肝癌细胞（BEL－7402）凋亡

本文从细胞学、ＤＮＡ凝胶电泳、流式细胞术三方面研究促肝细胞生长素（ｐＨＧＦ）在体外对肝癌细胞增殖活性的影响。结果表明：ｐＨＧＦ对肝癌ＢＥＬ－７４０２细胞增殖有抑制作用，并存在剂量和时间相关性。其４８ｈ的半数抑制浓度（ＩＤ５０）为０．３７ｍｇ／ｍｌ±０．０４ｍｇ／ｍｌ。而３７℃灭活的ｐＨＧＦ对ＢＥＬ－７４０２细胞增殖在１５ｈ无抑制作用，在２４和４８ｈ抑制作用很弱（ＩＤ５０＞１．５ｍｇ／ｍｌ）。ＤＮＡ凝胶电泳结果表明，ｐＨＧＦ可诱导ＢＥＬ７４０２细胞产生细胞凋亡（Ａｐｏｐｔｏｓｉｓ）。流式细胞术（ＦＣＭ）结果显示：ｐＨＧＦ抑制ＢＥＬ－７４０２细胞增殖过程是先使细胞停留在Ｇ０／Ｇ１期，继而诱导细胞产生凋亡。后两项结果均显示ｐＨＧＦ对人肝癌细胞凋亡的诱导里时间和剂量相关性。     

Liver Anti-Fibrosis Therapy with Mesenchymal Stem Cells Secreting Hepatocyte Growth Factor

Abstract

The objective of this study is to investigate the anti-fibrotic effect of combined mesencymal stem cells (MSCs) and gene therapy on liver fibrosis. When transfected by the complex with a plasmid DNA of hepatocyte growth factor (HGF) and the spermine-introduced pullulan of gene carrier, MSCs secreted HGF protein over 1 week. The HGF secreted from transfected MSC had the biological activity to promote the albumin production of hepatocytes. After intravenous injection, the HGF-secreting MSCs (HGF-MSC) accumulated in the liver. The injection of HGF-MSC decreased the fibrosis area in a rat model of liver fibrosis to a significantly great extent compared with that of original MSC. In the in vitro experiment, the higher number of HGF-transfected MSCs was migrated by stromal cell-derived factor (SDF)-1α more strongly than the original MSC. Considering the promotion of SDF-1α secretion in the liver fibrosis, it is possible that, when transplanted, genetically-engineered MSCs are accumulated in the liver due to their higher response to SDF-1α. It is concluded that the intravenous injection of genetically-engineered MSCs is a promising therapy for liver fibrosis.     

表皮生长因子与生长抑素对人离体胎盘分泌hCG的影响

本工作作利用灌流技术，观察了不同浓度的表皮生长因子（２５，５０，１００，５００ｎｇ／ｍｌ）和生长抑素对妊娠早期人工流产新鲜胎盘绒毛分泌人绒毛膜促性腺激素的影响。结果表明，ＧＥＦ对人早期胎盘ｈＣＧ的分泌有双向作用，５０ｎｇ对ｈＣＧ分泌刺激作用最大，５００ｎｇ则抑制ｈＣＧ的分泌。生长抑素单独使用不影响ｈＣＧ分泌，但可抑制ＥＧＦ诱导的ｈＣＧ分泌。提示ＥＧＦ，ＳＲＩＦ可能参与ｈＣＧ的分泌调节。     

子宫肌瘤组织EGFR受体与IGF-1R的定位及定量分析

目的 :探讨表皮生长因子受体与胰岛素样生长因子受体在子宫肌瘤发病机理中的作用。方法 :应用流式细胞术定量分析法及免疫组化S -P法分析比较 4 0例子宫肌瘤患者子宫肌层和子宫肌瘤组织表皮生长因子受体 (EGFR)和胰岛素样生长因子受体 (IGF - 1R)的含量及表达水平 ,月经周期根据月经史及子宫内膜组织来判断。结果 :子宫肌瘤组织EGFR ,IGF - 1R的表达及含量显著高于同一子宫邻近正常肌层组织 (p <0 0 5 ,p <0 0 1 ) ;子宫肌瘤组织与其邻近肌层组织相比较 ,EGFR的含量及表达在分泌期无显著性差异 (p >0 0 5 ) ,而在增生期含量明显增高 (p <0 0 5 ) ;而IGF - 1R的表达与月经周期无关。 结论 :EGFR ,IGF - 1R在子宫肌瘤组织中高表达与其发生、发展有关     

Expression of Hepatocyte Growth Factor (HGF) and its Receptor (MET) in Medullary Carcinoma of the Thyroid

The tyrosine kinase receptor encoded by the MET oncogene has the unusual property of mediating the invasive growth of epithelial cells upon binding with the hepatocyte growth factor (HGF). The MET/HGF receptor is known to be overexpressed in thyroid carcinomas originated from follicular cells, but has not been reported in C cell tumors. To investigate the role of HGF and of its receptor (encoded by MET oncogene) in medullary carcinoma of the thyroid (MCT), we studied the expression of HGF and MET in 20 cases by means of different techniques. By RT-PCR, HGF mRNA was found in 10/20 cases, while MET mRNA presence was demonstrated in 8/20, of which 7 also expressed HGF. Northern blot analysis and in situ hybridization were performed in selected cases and confirmed RT-PCR data in some cases, although the lower sensitivity of these procedures did not allow the identification of all RT-PCR positive cases. By immunohistochemistry (using specific monoclonal antibodies) HGF was demonstrated in 8/9 RT-PCR positive cases ald a monoclonal to MET immunostained 5/6 RT-PCR positive cases. All receptor positive cases also expressed the ligand in the same tumor cell population. These findings demonstrate MET and HGF co-expression in a subset of MCT, in which autocrine/paracrine circuits may be active. No correlation was found between HGF/MET expression and clinico-pathological parameters, except for the more common multifocality of HGF/MET positive MCT. Whether the potential activation of MET in MCT is responsible for local invasion and malignant evolution is to be further investigated, especially in multifocal and aggressive tumors.     

Hepatocyte growth factor in human semen and its association with semen parameters

Hepatocyte growth factor (HGF) is a structurally unique growth factor with potent motogenic (motility inducing) effects. Studies in the murine male genital tract have suggested important associations between HGF and the acquisition of sperm motility during epididymal maturation. The aim of this study was, therefore, to determine the concentration of HGF in human semen and assess its correlation, if any, with sperm motility and other semen parameters. Semen samples were collected by masturbation and analysed using standard procedures. HGF concentrations were measured in duplicate using an enzyme-linked immunoassay technique. Total protein estimations were also made in a subset of samples. The 95 subjects were divided into three groups for analysis: normozoospermic, subnormal semen and azoospermic. HGF was detected in all samples (median 0.456, 25th centile 0.388, 75th centile 0.556 ng/ml). No significant correlations were found between semen HGF concentrations and sperm concentration, motility, total sperm count or total motile count. There were no significant differences in mean HGF concentrations between the three subgroups. In conclusion HGF is present in human semen in significant quantities. The data do not suggest HGF concentrations are correlated with parameters of sperm motility.     

Placenta growth factor (PlGF) induces vascular endothelial growth factor (VEGF) secretion from mononuclear cells and is co-expressed with VEGF in synovial fluid

The aims of this study were (i) to determine whether PlGF, VEGF and PlGF/VEGF heterodimers are detected in synovial fluid (SF) and plasma samples from patients with a range of arthropathies; (ii) to describe whether any correlation exists between SF PlGF, VEGF and PlGF/VEGF heterodimer levels and the total and differential SF leucocyte counts; and (iii) to investigate the regulation of peripheral blood mononuclear cell (PBMC) VEGF secretion by stimuli relevant to inflammatory joints. PlGF, VEGF and PlGF/VEGF heterodimer levels were measured in the SF and plasma of patients with a range of arthropathies and normal controls by ELISA. Western blotting for PlGF was performed on SF from three patients with rheumatoid arthritis (RA) and primary inflammatory arthropathies. VEGF was quantified in cell culture supernatants after stimulation with lipopolysaccharide (LPS), PlGF or cobalt ions of PBMC isolated from RA patients and controls. PlGF and VEGF were detected in all SF samples. PlGF/VEGF heterodimers were detected in 10.2% of SF samples, most frequently in RA samples. Western blotting confirmed the presence of PlGF in RA SF. PlGF was detected in 52% of RA and 31% of control plasma samples, and VEGF was detected in 38% of RA and 38% of control plasma samples. PlGF/VEGF heterodimers were detected in 21% of RA samples and none of the control samples. In primary inflammatory arthropathy patients, SF PlGF and VEGF levels correlated significantly with the SF total leucocyte count and the neutrophil count. PlGF was the most potent inducer of PBMC VEGF production in both RA and control subjects. This is the first report of the detection of PlGF and PlGF/VEGF heterodimers in the SF of patients with inflammatory arthropathies, and we have shown for the first time that PlGF up-regulates PBMC VEGF production. PlGF may therefore play a key role in the production of VEGF in the inflammatory joint.     

表皮生长因子受体与他莫西芬耐药机制

雌激素受体及其信号通路在乳腺癌的发生发展中发挥着关键作用.到目前为止,抑制或阻断雌激素信号通路的内分泌治疗尤其是他莫西芬,仍是对雌激素受体阳性乳腺癌患者最有效的治疗手段之一.然而,他莫西芬的耐药问题直接影响了乳腺癌患者的治疗及预后.最近多项研究表明雌激素受体与表皮生长因子受体家族尤其是HER2介导的信号传导通路在多个点上相互交叉,彼此影响,与他莫西芬的耐药密切相关     

Recombinant Human Mullerian Inhibiting Substance Inhibits Epidermal Growth Factor Receptor Tyrosine Kinase

Abstract

Autophosphorylation of the epidermal growth factor (EGF) receptor in A-431 cells and plasma membrane fractions was inhibited by partially purified recombinant human Mullerian Inhibiting Substance (MIS). Immunoprecipitation of the EFG receptor using anti-EGF receptor or anti-phosphotyrosine antibodies, and phosphoamino acid analysis of this receptor, demonstrated that MIS specifically inhibited EGF-induced tyrosine phosphorylation. Inhibition of EGF receptor autophosphorylation by MIS in membrane preparations was not affected by increasing concentrations of EGF, manganese or [γ-(32)P] ATP. Thus, it is unlikely that MIS competes for EGF binding sites or sequesters substrate. Immunoabsorption of MIS with anti-human MIS antibody blocked the MIS inhibition of EGF receptor autophosphorylation, indicating that the inhibition was due to MIS. Our data suggest that MIS regulates the activity of the EGF receptor tyrosine kinase in A-431 cells.     

ImmunohistochemicaI and UItrastructuraI LocaIization of Epidermal Growth Factor Receptor in Human Liver and Hepatocellular Carcinoma Tissues

Expression of epidermal growth factor receptor (EGF-R) in human hepatocellular carcinoma (HCC), hepatoblastoma and non-cancerous liver tissues was investigated immuno-histochemically in order to evaluate the possible role of EGF-R expression in neoplastic transformation of he-patocytes. lmmunoreactive EGF-R molecules were identified on frozen sections by means of the avidin-biotin im-munoperoxidase complex technique using a monoclonal antibody recognizing an epitope of the external domain of human EGF-R. Linear positive staining was present on the surface of carcinoma cells in one hepatoblastoma and in 9 of 11HCCs. In addition, an enhanced level of surface EGF-R expression was observed on the tumor cells in 9 of 12 cases in comparison with that on hepatocytes in surrounding non-cancerous liver tissue, which in most cases showed chronic inflammation, hepatocyte injury or regeneration. No positive staining in the form of coalescent cytoplasmic granules was present in HCC or hepatoblastoma cells, nor in the cytoplasm of hepatocytes in normal or non-cancerous diseased liver tissue. Little or no reactivity was present on the surface membrane of hepatocytes in the normal liver tissues of 8 control cases. Furthermore, immunoelectron microscopy revealed the localization of this immunoreactive EGF-R molecule on the plasma membrane. Considering that the functional form of EGF-R could be localized on the plasma membrane, the enhanced expression of immunoreactive EGF-R on the tumor cell surface demonstrated here may suggest a possible role of EGF-R in the development or progression of human HCC as well as in hepatocyte regeneration. Acta Pathol Jpn 40: 22–29, 1990.