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High levels of PDGFR expression in primary rhabdomyosarcoma (RMS) have been associated with disease progression. To date however, there are no reports on the activity of imatinib mesylate, a selective PDGFR inhibitor, in RMS preclinical models. A panel of 5 RMS cell lines was used to investigate the expression of PDGFRalpha and PDGFRbeta, c-Kit and the multidrug transporter ABCG2 (also inhibited by imatinib). In vitro and in vivo experiments were performed using RD (embryonal) and RH30 (alveolar) cell lines to determine the efficacy of imatinib as single agent and in combination with topotecan (TPT). PDGFRbeta was significantly expressed in all cell lines, with the highest levels in RD, while PDGFR alpha and ABCG2 were significantly expressed only in RH30 and RMZ-RC2. c-Kit was not detected. PDGFRbeta signaling was active in RD but not in RH30, whilst PDGFRalpha signaling was not active in either cell lines. Significant ABCG2-mediated extrusion of Hoechst 33342 was demonstrated in RH30 but not in RD, and was inhibited by imatinib and the specific ABCG2 inhibitor Ko143. In vitro, imatinib was not active as a single agent at therapeutic concentrations, but significantly potentiated TPT antitumor activity in both cell lines. In vivo experiments using tumor xenografts confirmed the synergistic interaction in both cell lines. These results suggest that at least 2 different mechanisms--inhibition of ABCG2 and/or PDGFRbeta--are involved in the synergistic interaction between imatinib and TPT, and support the use of this combination for the treatment of high-risk RMS patients., (Copyright 2006 Wiley-Liss, Inc.)

The platelet-derived growth factor receptor (PDGFR) is a tyrosine kinase, implicated in the development and progression of different tumors, including gliomas. Chemoresistance is a common feature of malignant gliomas. Since receptor tyrosine kinases contribute to chemoresistance in tumors, we addressed whether PDGFR signaling might confer selective growth advantage to chemoresistant cells. The effects of the PDGFR inhibitor STI571 on proliferation and PDGFR signaling were compared in chemosensitive and cisplatin-selected, chemoresistant sublines derived from glioma and from two other PDGFR-expressing tumors (ovarian carcinoma and neuroblastoma). The chemoresistant glioma U87/Pt cells were twofold more sensitive to STI571 growth-inhibitory effects than the chemosensitive U87 cells, and two- to threefold more sensitive than five unrelated glioma cell lines. The other two paired cell lines were equally responsive. Sensitization of U87/Pt cells correlated with upregulation of the PDGF-B isoform and with PDGF-BB-induced Akt overactivation, which was prevented by STI571. STI571 specifically inhibited PDGF-BB-, but not PDGF-AA- or stem cell factor-mediated signaling. In serum-containing medium, STI571 decreased phospho-Akt in U87/Pt cells, but not in U87, while activating extracellular signal-regulated kinase (Erk) in both. STI571 antiproliferative effects were partially reverted by constitutively active Akt. Cotreatment with inhibitors of phosphatidylinositol 3'-kinase (PI3K) or mitogen-activated protein kinase kinase (MEK) resulted in enhanced growth inhibition in glioma cells. Our results suggest that increased PDGF-BB signaling may sensitize chemoresistant glioma cells to STI571, suggesting a therapeutic potential for STI571 in patients with malignant gliomas refractory to chemotherapy. Simultaneous blockade of PDGFR and PI3K or Erk pathway may enhance therapeutic targeting in gliomas., (Copyright 2006 Wiley-Liss, Inc.)

The immunocytochemical localization of insulin-like growth factor I (IGF-I) and platelet-derived growth factor A (PDGF-A) chain ligands, PDGF-chain receptors, and the intermediate filaments cytokeratin, desmin, and vimentin in the tissue reorganization and development during the establishment of the porcine epitheliochorial placenta was studied at light and electron microscopic levels in sections of endometrium and/or placenta of gilts during early pregnancy up to day 40 postmating. The endometrial epithelia (surface and glandular) as well as the trophoblast showed PDGF-A and IGF-I labelling. The only temporal difference was seen regarding IGF-I labelling, this being more uniform in the glands during precontact and early contact stages (days 7-12) compared to later stages. Cytokeratin labelling was conspicuous in all epithelia, including the trophoblast. The endometrial stroma showed strong labelling for the PDGF receptors and the intermediate filament vimentin, staining being enhanced along with the increase in the vascular bed during the establishment of the placenta. The maternal endothelium immunoreacted to IGF-I, to both PDGF-A and PDGF receptors, to vimentin and to von Willebrand factor (Factor VIII). Desmin was also expressed in the capillary bed underneath the maternal surface epithelium. In larger blood vessels, desmin was detected in the smooth muscle layer but not in the endothelium. The vascular smooth muscle also reacted with IGF-I, PDGF-A, and PDGF-receptor antibodies. The results suggest the involvement of both growth factors, IGF-I and PDGF, in the early nutrition of the pig embryo and the tissue reorganization that encompasses placentation. IGF-I appears to be related to the secretory tissue compartment (formation of histotrophe), whereas PDGF might play a role in the reorganization of the stroma, particularly during placental angiogenesis. Desmin and von Willebrand factor immunolabelling appears to be useful to monitor the development of the subepithelial capillary bed in the porcine placenta.

Background: The objective of this study was to evaluate the expression of platelet-derived growth factor (PDGF A, PDGF B) and their receptors (PDGF alpha, PDGF beta) in prostatic intraepithelial neoplasia (PIN) and adenocarcinoma., Methods: Peroxidase-antiperoxidase immunoperoxidase labeling was used to detect the extent of antibody labeling in 29 different high grade PIN specimens., Results: PDGF A and PDGF alpha were uniformly expressed in glandular epithelial and stromal cells, whereas PDGF B was not expressed by either tissue. However, PDGF beta was lightly expressed in a uniform manner by both the glandular epithelial and stromal cells., Conclusions: The data suggest that an autocrine loop may exist in which the epithelial cells of high-grade PIN express PDGF AA and PDGF alpha to modulate growth of precancerous lesions.

This study evaluates the distribution of receptors for platelet-derived growth factor (PDGF) on central nervous cells maintained in vitro using colloidal gold-labeled immunocytochemical markers at the electron microscopic level. Platelet-derived growth factor receptors were found to be sparsely distributed over the surface of type 1 astrocytes, apparent type 2 astrocytes, and neurons. Receptors appeared to be preferentially associated with filopodia-like extensions of the cell membrane. The existence of functional receptors was confirmed using the impermeant, water-soluble affinity cross-linking agent bis(sulfosuccinimidyl)suberate to covalently link radiolabeled PDGF to its receptor. The PDGF/receptor complexes could also be immunoprecipitated with the same antibody used in immunocytochemical experiments. The improved resolution of these techniques allows definitive identification of PDGF receptors on cultured mammalian central nervous system cells other than oligodendrocytes. These data expand the range of possible roles of PDGF during nervous system development. Receptors for PDGF are likely to play a key role in the differentiation of cells in the central nervous system.

Established cell lines derived from human malignant astrocytomas typically express a combination of platelet-derived growth factor (PDGF) and PDGF receptor which could form an autocrine loop. In this study, we screened for the essential components of a PDGF autocrine loop in fresh surgical isolates of human astrocytomas, using in situ hybridization and immunohistochemical techniques. Eight malignant astrocytomas (6 glioblastomas and 2 anaplastic astrocytomas), 5 low-grade astrocytomas and 4 non-neoplastic glial specimens (mesial temporal sclerosis) were evaluated. Malignant astrocytomas, and to a lesser extent low-grade astrocytomas, expressed more PDGF-A and PDGF-B than non-neoplastic glia. PDGF-alpha-receptor expression was elevated both in malignant and in low-grade astrocytomas. These data support the argument that PDGF autocrine loops contribute to the unregulated growth of human astrocytomas. Expression of PDGF and PDGF receptor in low-grade astrocytomas suggests that activation of PDGF autocrine loops may be an early event in the pathogenesis of malignant astrocytomas.

We have examined the state of tyrosine phosphorylation of ligand-bound, internalized platelet-derived growth factor (PDGF) beta-receptors. Analysis by immunofluorescence staining of cells stimulated with PDGF-BB at 37 degrees C indicated colocalization of phosphotyrosine, PDGF beta-receptors, and PDGF-BB in endosome-like vesicles. Treatment of cells with an acidic buffer, which removed cell surface-bound PDGF-BB, led to a considerable decrease in phosphorylation and kinase activity of cell surface localized PDGF beta-receptors, but not of internalized receptors. Immunoprecipitations using antisera against phosphotyrosine and the PDGF beta-receptor from metabolically labeled cells showed that a major fraction of the tyrosine-phosphorylated pool of receptors were still accessible to the acid buffer treatment after 10 min of incubation of the cells at 37 degrees C. Under these conditions, about 20-25% of the total pool of tyrosine-phosphorylated receptors were intracellular, since they remained tyrosine phosphorylated after the acid buffer treatment. A considerable pool of tyrosine-phosphorylated, internalized receptors, after 10 min of incubation of the cells at 37 degrees C, could also be detected by immunoblotting analysis, using antisera against the PDGF beta-receptor and phosphotyrosine. Analysis by in vitro kinase assays of immunoprecipitated PDGF beta-receptors, obtained from PDGF-BB-stimulated cells different times after acid wash, showed that the internalized receptors retained kinase activity. These data suggest that a pool of internalized PDGF beta-receptors remain active and may participate in signalling a considerable time after internalization.