Coexpression of functionally active receptors for thyrotropin and platelet-derived growth factor in human thyroid carcinoma cells.

In the present study we show the simultaneous expression of functionally active receptors for TSH and platelet-derived growth factor (PDGF) in a newly established human anaplastic thyroid carcinoma cell line, HTh 74. In Northern blot analysis of RNA extracted from HTh 74 cells a low expression of both TSH and PDGF receptor messenger RNA was found. These observations in conjunction with the fact that the cells contain cytokeratin clearly demonstrate that the cells are bona fide epithelial thyroid cells. Stimulation of HTh 74 cells with TSH led to a concentration-dependent increase in cAMP formation, showing a functional activity of the TSH receptors. Northern blot analysis, immunoprecipitation, immunofluorescence staining, and binding experiments showed the presence of both alpha- and beta-type PDGF receptors in the HTh 74 cells. The functional activity of the PDGF receptors was demonstrated by ligand-induced internalization of the receptors and PDGF-induced growth of the HTh 74 cells. The significance of the expression of PDGF receptors on thyroid epithelial cells is not clear. However, it might reflect the gain of a new growth stimulatory pathway participating in the transformation of the epithelial thyroid cells. Alternatively, the PDGF receptors may be remnants from an immature progenitor cell from which the undifferentiated carcinoma has evolved.     

Aberrant production of gliostatin/platelet-derived endothelial cell growth factor in rheumatoid synovium

Objective. To purify a protein inhibitor from rheumatoid arthritis (RA) synovial fluids which suppresses the apparent incorporation of ³H-thymidine into fibroblasts and synovial cells, and to define its biochemical features that have clinical relevance to the pathogenesis of RA. Methods. Several standard chromatographic techniques were employed for the purification of the protein. Immunochemical methods with monoclonal antibody were used to quantify and visualize the protein in sera, synovial fluids, and tissues from RA patients. Results. The chemical properties of purified inhibitor from RA synovial fluids confirmed its identity as gliostatin/platelet-derived endothelial cell growth factor (PD-ECGF), a potent angiogenic factor. The gliostatin/ PD-ECGF level in synovial fluid and serum was higher in RA patients than in osteoarthritis controls. Conclusion. These findings strongly suggest that gliostatin/PD-ECGF might play an important role in the aberrant neovascularization of rheumatoid synovium.     

Immunohistochemical expression of thymidine phosphorylase/platelet-derived endothelial cell growth factor in squamous cell carcinoma of the esophagus.

BACKGROUND/AIMS: Thymidine phosphorylase (dThdPase) is identical to platelet-derived endothelial cell growth factor which is an angiogenic factor. We attempted to clarify the significance of dThdPase expression in esophageal squamous cell carcinoma (SCC). METHODOLOGY: Tissues samples were taken from 50 patients with esophageal SCC after curative surgery. The expression of dThdPase was immunohistochemically examined using a monoclonal antibody to dThdPase (clone 654-1). Microvessels in SCC stained for Factor VIII-related antigen were counted. Vascular endothelial growth factor (VEGF) was immunostained with R11. Ki-67 antigen was immunostained with MIB-1, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick end labeling was performed, and Ki-67 labeling index (LI) and apoptotic index were calculated. RESULTS: The expression of dThdPase was observed in 30 patients (60%). Between the SCC with and without dThdPase expression, significant differences were found in microvessel count (p < 0.001) and VEGF expression (p < 0.01), but not in Ki-67 LI and apoptotic index. With regard to the clinicopathologic factors, significant differences were observed in histologic venous invasion (p < 0.01) and lymph node metastasis (p < 0.05). Survival rate after surgery was better in the patients with dThdPase-negative SCC (p < 0.05), and distant organ metastasis after surgery was frequently observed in the patients with dThdPase-positive SCC (p < 0.05). CONCLUSIONS: These results suggest that dThdPase expression may be associated with angiogenic promotion and may be one of the prognostic factors in esophageal SCC.     

PD-ECGF、Survivin在胰腺癌组织中的表达及其与细胞凋亡的关系

目的 探讨血小板衍化内皮细胞生长因子(PD-ECGF)和存活素(Survivin)在胰腺癌组织中的表达及其与细胞凋亡的关系.方法 应用免疫组化技术检测42例胰腺癌患者癌组织的PD-ECGF和Survivin表达,流式细胞术检测胰腺癌凋亡指数(AI).结果 PD-ECGF的表达与胰腺癌淋巴结转移显著相关,Survivin的表达与胰腺癌组织分化程度显著相关(P均<0.05);两者与AI值均呈显著负相关,两者共同表达阳性者AI值显著低于共同表达阴性者(P均<0.01).结论 胰腺癌组织中PD-ECGF和Survivin可抑制胰腺癌细胞凋亡,促进其增殖与转移.     

血小板源性生长因子及其受体在心血管疾病中的作用

血小板源性生成因子（PDGFs）及其受体（PDGFRs）在心血管疾病中发挥着重要作用,二者不仅参与了动脉粥样硬化（AS）的形成,而且在血管生成、心肌纤维化过程中也扮演了重要角色。PDGFRs被其配体PDGFs激活后,在血管平滑肌细胞、纤维母细胞、内皮细胞等中的表达上调,通过细胞内信号转导通路,促使该类靶细胞增殖分化,从而参与AS、血管生成和心肌纤维化的发生。本文就PDGFs/PDGFRs在心血管疾病中的研究进展做一综述。     

Human microvascular endothelial cells express receptors for platelet-derived growth factor.

Endothelial cells have been widely thought to be unresponsive to platelet-derived growth factor (PDGF, a major growth factor released from stimulated platelets at the sites of vascular insults) and devoid of PDGF receptors. Nevertheless, in examining the growth-factor responses of microvascular endothelial cells isolated from human omental adipose tissue, we were surprised to detect PDGF-induced tyrosine phosphorylation of a 180-kDa glycoprotein, subsequently identified as the cellular receptor for PDGF by specific immunoprecipitation. Scatchard analysis of 125I-labeled PDGF binding to human microvascular endothelial cells revealed 30,000 PDGF receptors per cell with a Kd of 0.14 nM. PDGF stimulated tyrosine phosphorylation of PDGF receptors and other cellular proteins in a dose- and time-dependent manner, with half-maximal receptor phosphorylation occurring at 0.3 nM recombinant human PDGF (B chain) and a less than or equal to 1-min exposure to PDGF. Normal cellular consequences of receptor activation were also observed, including tyrosine phosphorylation of a 42-kDa protein and serine phosphorylation of ribosomal protein S6. Furthermore, PDGF was mitogenic for these cells. Microvascular endothelial cells play a central role in neovascularization required for wound healing and solid tumor growth. Thus, the discovery of functional PDGF receptors on human microvascular endothelial cells suggests a direct role for PDGF in this process.     

Clodronate alleviates cachexia and prolongs survival in nude mice xenografted with an anaplastic thyroid carcinoma cell line

Cancer cachexia is one of the most common manifestations of advanced malignant disease and is frequently associated with decreased survival. Previously, we reported the establishment of a new anaplastic thyroid carcinoma cell line, Thena, and its mouse xenograft, Thena-Nu, which induced cachexia in athymic nude mice. Subsequent studies showed that the addition of clodronate to Thena-Nu cultures reduced cell proliferation as well as cytokine production in a dose- and time-dependent manner. Weekly administration of clodronate induced tumor cytostasis, attenuation of cachexia, as well as prolongation of survival in Thena-Nu-bearing mice. Reduced serum interleukin 6, tumor necrosis factor-alpha, and granulocyte colony stimulating factor levels were detected, whereas, serum leukemia inhibitory factor levels were not reduced. Liver necrosis, observed in tumor-bearing mice, was also improved following clodronate treatment. Discontinuation of clodronate treatment, however, resulted in progressive tumor growth and weight loss. Our results demonstrated that clodronate could exert therapeutic efficacy on amelioration of cancer cachexia in the hosts. Nevertheless, this study also points out that a longer period of treatment is required to maintain these effects.     

Fibroblast growth factor receptors as molecular targets in thyroid carcinoma

Several molecular abnormalities of potential therapeutic target value have been described in thyroid neoplastic transition. We report the expression of the fibroblast growth factor receptor family (FGFR-1-4) in normal thyroid tissues, human thyroid cancers of various types and behaviors, and cell lines representative of the spectrum of differentiation of tumors derived from follicular epithelial cells. FGFR-2 was the only receptor consistently detected in normal human thyroid tissue, and its expression diminished in all thyroid cancers and carcinoma cell lines, suggesting that it may have a protective role. FGFR-1 and FGFR-3 were expressed in most well-differentiated tumor types. FGFR-4, however, was expressed predominantly in aggressive tumor types and the most rapidly proliferative cell lines, indicating that it may promote the progression of these tumors. To specifically determine the function of FGFR-4 in thyroid carcinoma, gain- or loss-of-function studies were performed in cell lines representative of the spectrum of thyroid cancer behavior. Introduction of FGFR-4 resulted in enhanced cell proliferation, an effect that was more pronounced in cell lines derived from aggressive tumors than in those derived from more indolent neoplasms. Moreover, transduction of a dominant-negative FGFR attenuated cell proliferation in the aggressive poorly differentiated cell lines with no appreciable effect in well-differentiated cells. Pharmacologic FGFR-4 tyrosine kinase inhibition resulted in significant proliferation arrest in an aggressive cell line endogenously expressing the receptor. Furthermore, systemic administration of the FGFR tyrosine kinase inhibitor PD173074 resulted in significant inhibition of follicular thyroid carcinoma-derived cell growth in xenografted severe combined immunodeficient mice. These data indicate a role for FGFR-4 in human thyroid cancer cell progression and provide a rationale for FGFR manipulation as a potentially novel therapeutic approach.     

大肠癌的血管生成与血小板源生长因子的表达研究

目的探讨血管生成与血小板源生长因子的表达在大肠癌的发生、发展中的作用.方法采用免疫组化SABC法,对45例大肠癌患者和10例正常大肠组织标本进行微血管密度和PDGF表达的检测,微血管的染色结果用图象分析系统进行处理,并结合大肠癌的临床病理进行分析.结果PDGF染色阳性29例,占64.4%,其中强阳性10例,占22.2%,而在正常的大肠组织中没有1例PDGF的表达阳性.血管生成与大肠癌的局部浸润程度(0.32±0.09 vs 0.29±0.09)、Dukes分期(0.36±0.06 vs 0.25±0.05)、器官转移情况相关(0.38±0.06 vs 0.27±0.08).PDGF表达阳性组的血管生成较PDGF表达阴性组明显增高(0.31±0.08 vs 0.25±0.06).结论PDGF是肿瘤血管生成的正性调节因子,血管生成在大肠癌的发展、演进中起重要的作用.     

Epidermal growth factor induces the functional expression of dopamine receptors in the GH3 cell line.

GH3 cells are a clonal strain from a rat pituitary tumor that synthesizes and secretes both PRL and GH. The peculiarity of these cells is that they do not express receptors for dopamine; thus the hormone release is insensitive to the inhibitory effect of dopamine and D2 receptor agonists. Exposure of GH3 cells to epidermal growth factor for 4 consecutive days markedly altered the cell morphology, from a spherical appearance to an elongated flattened shape, and increased the cell size. These morphological changes were accompanied by the functional expression of D2 dopamine receptors as shown by the presence of a specific, saturable, and stereoselective high affinity binding for [3H]spiroperidol in epidermal growth factor-treated cells and by the fact that the selective D2 agonist quinpirole recovered the property to inhibit PRL secretion in the cell cultures exposed to the neurotrophic factor. The effect of EGF on the functional expression of D2 receptors was dose dependent (EC50 = 8 pM) and reversible. These data suggest that EGF elicits major effects on the expression of specific genes leading to the differentiation of GH3 cells into lactotroph-like cells endowed with dopamine D2 receptors.     

Butyrate alters the expression and activity of cell cycle components in anaplastic thyroid carcinoma cells.

Anaplastic thyroid carcinoma (ATC) is the most malignant and aggressive form of thyroid cancer. Most patients die within months of diagnosis, primarily due to the absence of effective chemotherapeutic strategies. Identifying alternative therapies is necessary to increase long-term survival. Butyrate elicits a number of responses from cancer cells both in vitro and in vivo including growth repression, cell cycle arrest, differentiation, and apoptosis. Even though many types of cancer cells have been studied, little is known of the response of ATC cells to this drug. In this study, we report that butyrate induces differential cell cycle arrest (arrest in G1 and G2/M phases) in an ATC cell line that correlates with changes in the expression, phosphorylation, and activity of key components of the cell cycle machinery. Exposure to butyrate increases the expression of the cyclin-dependent kinase inhibitors, p21/Cip1 and p27/Kip1, decreases the expression of cyclin A and cyclin B, inhibits the phosphorylation of the retinoblastoma protein (pRb), and decreases the activity of cdk1 and cdk2-associated kinases. These results suggest that butyrate may be useful in the clinical treatment of ATC.     

Two platelet-derived growth factor receptors in vascular smooth muscle cells.

Distinct genes encode alpha and beta PDGF receptors which differ in their abilities to be triggered by three dimeric forms of the PDGF molecules. By use of a strategy involving introduction of expression vectors for alpha and beta PDGF receptor cDNA into the cells originally lacking these receptors, we demonstrated that each receptor was able to couple independently with mitogenic signal transduction pathways inherently present in these cells. Moreover, both receptors were capable of inducing a readily detectable chemotactic response. The vascular smooth muscle cells which express both types of PDGF receptors are mitogenic and chemotactic for PDGFs. Moreover, the alpha receptor is the preferred receptor for platelet PDGF-AB as well as the PDGF-AA isoform which is ubiquitously produced in many cells forming atherosclerotic plaques including macrophages, endothelial cells and even arterial smooth muscle cells. Our results indicated that the availability of specific PDGF isoforms and the relative expression of each receptor gene product appear to be major determinants of the PDGF response. An understanding of the mechanisms by which the expression of PDGF and their receptors on vascular smooth muscle cells are regulated will give greater insights as to how these gene products are involved in atherosclerosis.     

Production of platelet-derived growth factor-like molecules and reduced expression of platelet-derived growth factor receptors accompany transformation by a wide spectrum of agents.

A series of nontransformed human and murine cells and derivative cell lines transformed by methylcholanthrene; by simian virus 40, Kirsten and Moloney murine sarcoma viruses, simian sarcoma virus, and adenovirus; and by a "spontaneous" event in culture were examined for the expression of receptors for the platelet-derived growth factor (PDGF) and for production of substances able to compete with 125I-labeled PDGF for binding to the cell-surface PDGF receptor. In each case, transformation resulted in a 50-100% decrease in available PDGF receptors. All transformed cells except the methylcholanthrene-transformed mouse cells produce a PDGF competitor into the conditioned medium. Levels of PDGF competitor in conditioned medium at the end of a 48-hr collection were as high as 2 ng/ml--high enough to be measured by radioreceptor assay diluted 1:30 and to maximally stimulate [3H]thymidine incorporation by human fibroblasts. The PDGF competitor activity detected in a radioreceptor assay does not reflect irreversible (e.g., proteolytic) damage to the receptor of test cells since its effects are reversed by acetic acid dissociation. Antiserum against human PDGF neutralizes 20-80% of the PDGF competitor found in conditioned medium from different transformed human cells and 100% of the activity from normal human endothelial cells. The possibility that induction of expression of the cellular PDGF gene may be involved in the mechanism of transformation of PDGF-responsive mesenchymal cells is discussed.     

Rat Leydig cells bind platelet-derived growth factor through specific receptors and produce platelet-derived growth factor-like molecules.

The platelet-derived growth factor (PDGF) is a major mitogen for cells of mesenchymal origin. Because Leydig cells arise from mesenchymal precursors, we tested the hypothesis that these cells might be a target for PDGF. We also investigated a possible production of a PDGF-like substance by Leydig cells in culture and the distribution of PDGF-like material in the rat testis using immunohistochemistry. PDGF was found to bind specifically to high affinity receptors on the surface of purified adult rat Leydig cells. Conditioned medium from cultured Leydig cells competed with 125I-labeled PDGF for binding to the Leydig cells. The secretion of PDGF receptor-competing activity was stimulated in a dose-dependent manner by the trophic hormone hCG. Immunohistochemical studies revealed specific staining for PDGF in the Leydig cells of adult rat testis. Taken together these observations suggest that PDGF may play a role in the local control mechanisms of testicular function.     

胃癌组织血小板衍化内皮细胞生长因子的表达与血管生成和凋亡的关系

目的探讨血小板衍化内皮细胞生长因子（PD．ECGF）在胃癌组织中的表达及与血管生成和凋亡的关系。方法应用免疫组化技术对67例胃癌组织进行PD－ECGF表达、肿瘤组织微血管密度（MVD）检测，流式细胞技术检测胃癌组织细胞凋亡指数（川，凋亡百分率八结果＊r＊＊＊F的表达与胃癌淋巴结转移（P＜0．oj）。分化程度（P＜0．05）及组织分型（P＜0．05）显著相关，与＊V则尸＜001）和胃癌细胞凋亡指数（P＜001）显著相关。＊＊D和＊1与淋巳结转移（P＜001）显著相关。结论胃癌组织中P＊。 ECGF可促进血管生成，抑制胃癌细胞凋亡，促进胃癌的增殖与转移。     

Autocrine expression of platelet-derived growth factor B in B cell chronic lymphocytic leukemia

Platelet-derived growth factor (PDGF) regulates clonal proliferation of malignant pre-B cell lines, but little is known about its role in normal B lymphocyte differentiation and malignant transformation. To understand the expression of PDGF-A, PDGF-B and the beta-receptor (PDGF-Rbeta) in B cell lymphoproliferative disorders, we used an immunohistochemical method to stain formalin-fixed, paraffin-embedded tissues in 5 patients with reactive lymphoid hyperplasia, 15 with non-Hodgkin's lymphoma and 23 with B cell chronic lymphocytic leukemia (B-CLL). Abundant PDGF-A, rather than PDGF-B, was expressed in normal B cell differentiation. There was no difference in the expression of PDGF-A and PDGF-B between patients with reactive lymphoid hyperplasia and patients with malignant lymphoproliferative disorders. Among the patients with B-CLL, the expression of PDGF-B was much stronger than the expression of PDGF-A, and 18 of the patients had coexpression of PDGF-B and PDGF-Rbeta. A larger proportion of patients with B-CLL than with non-Hodgkin's lymphoma had expression of PDGF-B and PDGF-Rbeta. In conclusion, PDGF-A expression in all stages of B lymphocyte differentiation suggests that it is important in B cell differentiation and proliferation. Expression of PDGF-B and PDGF-Rbeta suggests that autocrine signaling of PDGF may be important in malignant transformation of B-CLL. However, further studies are necessary to confirm these conclusions.     

Recycling of epidermal growth factor in a human pancreatic carcinoma cell line.

PANC-1 human pancreatic carcinoma cells readily bound and internalized 125I-labeled epidermal growth factor (EGF). Bound 125I-labeled EGF was then partially processed to a number of high molecular weight acidic species. Percoll gradient centrifugation of cell homogenates indicated that the majority of 125I activity localized to several intracellular vesicular compartments. Both intact EGF and its processed species were subsequently released into the incubation medium. A major portion of the released radioactivity was capable of rebinding to the cell. Only a small amount of bound 125I-labeled EGF was degraded to low molecular weight products, and this degradation was completely blocked by methylamine. This lysosomotropic compound did not arrest either the generation or the extrusion of the major high molecular weight species of processed EGF (pI 4.2). These findings suggest that in PANC-1 cells, bound EGF undergoes only limited processing. Both intact EGF and its major processed species bypass the cellular degradative pathways, are slowly released from the cell, and then rebind to the cell.     

Dual control of cell growth by somatomedins and platelet-derived growth factor.

Quiescent BALB/c 3T3 cells exposed briefly to a platelet-derived growth factor (PDGF) become "competent" to replicate their DNA but do not "progress" into S phase unless incubated with growth factors contained in platelet-poor plasma. Plasma from hypophysectomized rats is deficient in progression activity; it does not stimulate PDGF-treated competent cells to synthesize DNA, demonstrating that somatomedin C is required for progression. Various growth factors were tested for progression activity and competence activity by using BALB/c 3T3 tissue culture assays. Multiplication stimulating activity and other members of the somatomedin family of growth factors are (like somatomedin C) potent mediators of progression. Other mitogenic agents, such as fibroblast growth factor, are (like PDGF) potent inducers of competence. Growth factors with potent progression activity have little or no competence activity and vice versa. In contrast, simian virus 40 provides both competence and progression activity. Coordinate control of BALB/c 3T3 cell growth in vitro by competence factors and somatomedins may be a specific example of a common pattern of growth regulation in animal tissues.