Staining For Platelet-derived Growth Factor Research Articles

ANNEXIN A2 REGULATES ANGIOGENESIS AND INVASION PHENOTYPES OF MALIGNANT GLIOMA

BACKGROUND: Despite advances in multimodal therapy, patients with malignant glioma have a dismal prognosis. Aberrant angiogenesis and diffuse glioma cell invasion are major obstacles for effective treatment. Therefore, investigation of angiogenesis and invasion is essential for the development of a curative therapy. We established animal models that histologically mimic two invasive and angiogenic phenotypes of glioma, ie, angiogenesis-dependent invasion (J3T-1) and angiogenesis-independent invasion (J3T-2). In our previous study, comparative proteomic analysis showed that annexin A2 was more highly expressed in J3T-1 than in J3T-2. In this study, the function of annexin A2 in malignant glioma in relation to angiogenesis and invasion was investigated using these animal models. METHODS: Annexin A2-downregulated J3T-1 cells (J3T-1shA) and annexin A2-upregulated J3T-2 cells (J3T-2A) were generated by gene transfection. Quantitative reverse transcription polymerase chain reaction (QRT-PCR) and immunohistochemical staining of annexin A2 and vascular endothelial growth factor (VEGF) were performed on cultured cells. J3T-1, J3T-1shA, J3T-2, and J3T-2A brain tumors were established in athymic rats. Brain sections were evaluated histopathologically. Human glioblastoma specimens were stained for annexin A2, VEGF, and platelet-derived growth factor (PDGF). RESULTS: QRT-PCR and immunohistochemical staining of cultured cells revealed that expression of VEGF was upregulated in accordance with the expression of annexin A2 in J3T-1 and J3T-2A cells. J3T-1 tumors showed remarkable angiogenic activity and invasion around neovasculature, whereas J3T-1shA tumors, including J3T-2 tumors, showed no angiogenic activity, but diffuse single-cell infiltration of tumor cells into normal parenchyma. J3T-2A tumors were highly angiogenic and showed angiogenesis-dependent invasion. Human glioblastoma specimens revealed that annexin A2 was positive in clusters of tumor cells around dilated vessels (25/30 cases). The intensity of VEGF and PDGF staining corresponded to the expression of annexin A2 (correlation coefficients: annexin A2/VEGF: R = 0.73, P < 0.05; annexin A2/PDGF: R = 0.37, P < 0.05). CONCLUSIONS: Our results revealed that the phenotype of glioma invasion is closely related to angiogenesis, and annexin A2 is one of the factors that regulates angiogenesis and invasion phenotype of malignant glioma. SECONDARY CATEGORY: Neuropathology & Tumor Biomarkers.

Role of fibrogenic markers in chronic hepatitis C and associated hepatocellular carcinoma

Detection and follow up of fibrogenesis in chronic hepatitis C (CHC) is mandatory for early treatment and risk stratification. The current study included 120 patients with CHC, of whom 30 had liver cirrhosis (LC) and 30 had hepatocellular carcinoma (HCC). 15 wedge liver biopsies, taken during laparoscopic cholecystectomy, were included as normal controls. Cases were subjected to laboratory investigations, serologic markers for viral hepatitis and assessment of circulating levels of hyaluronic acid (HA) and platelet-derived growth factor (PDGF). Immunohistochemical expression of connective tissue growth factor (CTGF), PDGF and transforming growth factor-β1 (TGF-β1) was also carried out. A significant increase (p < 0.01) in serum HA was noticed in CHC, LC and HCC compared to controls. Although, a significant decrease in serum PDGF was detected in CHC and LC compared to controls, HCC values were comparable. A significant up-regulation of CTGF was detected in CHC, LC and HCC (p < 0.01) in contrast to its limited mild expression in normal livers. Intense PDGF positive staining was noticed in CHC, LC and HCC compared to scattered faint expression in controls. The significant expression and marked intensity of PDGF staining matched the progress to tumorigenesis. A positive TGF-β1 immunostaining was also noticed in CHC, LC and HCC. An intense and extensive cytoplasmic expression of TGF-β1 was encountered in patients with LC revealing that CTGF, PDGF and TGF-β1 act synergistically in LC. Data revealed that HA and CTGF may be implicated as important diagnostic parameters for assessment of hepatic fibrosis and PDGF for monitoring malignant transformation in CHC.

Platelet Derived Growth Factor Secretion and Bone Healing After Er:YAG Laser Bone Irradiation

Er:YAG laser irradiation has been reported to enhance wound healing. However, no studies have evaluated the synthesis of growth factors after laser irradiation. The present study investigated the effects of laser irradiation on the amount of secretion of platelet derived growth factor (PDGF) in the wound, clarifying the effects of the Er:YAG laser on the bone healing. Osteotomies were prepared in the tibiae of 28 rats using an Er:YAG laser (test group). Maximum power of 8 watts, energy per pulse of 700mJ, and frequency up to 50Hz were used. The laser was used with external water irrigation, a spot size of 2mm, energy per pulse of 500 to 1000mJ/pulse, and energy density of 32J/cm(2). Twenty eight additional rats served as a control group and their osteotomies were prepared with a drill 1.3mm in diameter at 1000rpm, with simultaneous saline irrigation. Two rats from the tested group and 2 from the control group were sacrificed on each day following surgery (1-14days), and the tissue specimens were prepared for histologic evaluation. Immunohistochemical staining with anti-PDGF was performed after histologic examination. The difference between the PDGF staining intensities of the 2 treatment groups was analyzed using a multivariate logistic regression test. A significant rise in PDGF staining occurred in both groups 2-3days following surgery. However, while high PDGF counts remained for the 2-week experimental period in the laser group, PDGF levels in the control group returned to baseline levels 8days post surgery. The 2 groups (laser and control) were found to be different throughout the experiment, and the rat type was found to be a significant predictor (P = .000011). The present study demonstrated that Er:YAG laser irradiation seems to stimulate the secretion of PDGF in osteotomy sites in a rat model. It is possible that the high levels of PDGF are part of the mechanism that Er:YAG irradiation enhances and improves the healing of osteotomy sites.

Growth factor expression following clinical mandibular distraction osteogenesis in humans and its comparison with existing animal studies

Lengthening the mandible by distraction osteogenesis (DO) is nowadays a well recognized technique in maxillofacial surgery. In this study growth factor expression profiles were examined in biopsies taken from six patients undergoing mandibular DO and compared with findings from a sheep model for mandibular DO. In all patients (and sheep), the ascending ramus was distracted 10-15 mm at a rate of 1mm/day using an intraoral device. Biopsies were taken from the centre of the distraction zone 21 days after completion of distraction. Using standard immunohistochemical techniques, samples were stained for platelet-derived growth factor (PDGF), transforming growth factor beta (TGF-beta), basic fibroblast growth factor (bFGF) and bone morphogenetic proteins-2, -4 and -7 (BMP-2, -4, -7), matrix metalloproteinases-1 and -3 (MMP-1, -3), the vascular endothelial growth factor (VEGF), a marker for endothelial cells (CD-31) and type IV collagen (Col IV). Positive staining for PDGF, bFGF, TGF-beta, BMP-2, -4, and -7 was noted in cells and matrix components. There was intense staining for MMP-1. Strong staining for CD-31 and COL IV was observed adjacent to vessels. VEGF staining was less specific. Similar findings were noted in the sheep model. Growth factor expression in the human distraction site is similar to that in the sheep model.

Abundant expression of platelet-derived growth factor in spiral arteries in decidua associated with pregnancy-induced hypertension and its relevance to atherosis.

To elucidate the role of platelet-derived growth factor (PDGF) in the pathophysiology of pregnancy-induced hypertension (PIH) in which the spiral arteries of the decidua demonstrate the atherosclerotic change. We determined serum levels of PDGF and PDGF expression in the decidua as well as serum levels of 17 beta-estradiol (E2) and progesterone (P4) both in normotensive cases and in PIH cases. Furthermore, we investigated whether sex steroid hormones could interact with PDGF in cultured vascular smooth muscle cells (SMC) by immunohistochemical staining for proliferating cell nuclear antigen. Serum PDGF levels were higher (P<0.01) but serum E2 levels were lower (P<0.01) in PIH cases compared with normotensive cases. There was no statistically significant difference between serum P4 levels in PIH cases and those in normotensive cases. Immunohistochemical staining for PDGF in SMC of spiral arteries was more prominent in PIH cases than in normotensive cases. The proliferative potential of cultured SMC was stimulated by PDGF, but inhibited by concomitant treatment with PDGF and E2. PDGF is suggested to play an important role in the pathophysiology of PIH through its stimulatory effect on vascular SMC proliferation which may elicit the atherosclerotic change in the spiral arteries of the placenta.

Immunohistochemical analysis of platelet-derived growth factor and basic fibroblast growth factor in cardiac biopsy and autopsy specimens of heart transplant patients

The purposes of this study were to examine 250 heart biopsy specimens and 20 autopsy specimens from heart transplant patients for the presence and localization of platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF) and to correlate these findings with the histologic features of rejection and the autopsy findings of graft coronary vasculopathy and global ischemia. Positive specimen staining was significantly more prevalent for PDGF (78% of specimens) than for bFGF (54% of specimens) (p < 0.001). PDGF was distributed more in an interstitial (53%) than a vascular (28%) pattern and was associated with macrophages, whereas bFGF was distributed more in a vascular (50%) than an interstitial (12%) pattern. The prevalence of PDGF (but not bFGF) staining was significantly greater in biopsy specimens with at least grade 2 vascular rejection changes (81%) than in those without vascular rejection changes (58%) (p < 0.001). In autopsy specimens, PDGF staining was present in the hearts of all 5 patients (100%) who died of graft failure from coronary vasculopathy and was also present in all 11 hearts (100%) with global ischemic changes, but in only 4 of 9 (44%) of the hearts without global ischemia (p < 0.01). PDGF staining was absent in nontransplanted heart specimens, whereas bFGF staining in nontransplanted heart specimens was similar to that in transplanted hearts. We conclude that PDGF is increased in transplanted hearts, is distributed more in an interstitial pattern, and is associated with macrophages. Furthermore, PDGF staining is increased in transplanted hearts with evidence of vascular rejection, coronary vasculopathy, or global ischemia.

Isoforms of platelet-derived growth factor and its receptors in epiretinal membranes: Immunolocalization to retinal pigmented epithelial cells

Epiretinal membranes (ERMs) form on the inner surface of the retina in conjunction with various ocular disease processes, but the factors controlling their development are not understood. The predominant cell types involved are retinal pigmented epithelial (RPE) cells and retinal glia. Cultured RPE cells secrete platelet-derived growth factor (PDGF), which is chemotactic and mitogenic for both RPE cells and retinal glia and, therefore, could be involved in the development of ERMs. In the present study, we performed immunohistochemical staining for PDGF A chain (PDGF-A), PDGF B chain (PDGF-B), and both types of PDGF receptors (PDGFr alpha and PDGFr beta) on ERMs associated with various disease processes. PDGF-A is detected in most ERMs, regardless of the associated disease process, and it appears to be localized predominantly in RPE cells, recognized by the presence of pigment and the immunohistochemical demonstration of some or all of the following RPE-associated epitopes: class III beta-tubulin, keratin, the 65-kDa microsomal protein recognized by the RPE9 antibody, and cellular retinaldehyde-binding protein. PDGF-B is found only in minor subpopulations of cells in about half of the ERMs evaluated and, with only occasional exceptions, appears to be localized almost entirely in blood-borne cells found in and around vessels in vascularized ERMs. Both PDGFr alpha and PDGFr beta are demonstrated in most ERMs with neither isotype consistently predominating: they are found predominantly on RPE cells with many cells expressing both receptor types. ERMs with little or no RPE cell component contain little or no PDGF and PDGF receptor, whereas those in which the RPE cell represents the major cell type, have widespread PDGF and PDGF receptor positivity. These findings show that RPE cells in ERMs produce PDGF-A and PDGF alpha and PDGF beta receptors and suggest that autocrine and paracrine stimulation with PDGF may be involved in ERM pathogenesis.

Immunohistologic demonstration of platelet-derived growth factor(PDGF) in skin biopsies from acute stage of Kawasaki disease

The involvement of endothelial cells and vascular smooth muscle cells in the initiation, formation and development of aneurysmal dilatation of coronary arteries in Kswasaki disease(KD) have been emphasized. It has been thought that specific growth factors may have an effect on anintimal thickening and vascular dysfunction. Plateletderived growth factor(PDGF) is reported to be involved in various clinical situations such as wound healing,atherogenesis,embryogenesis, and infectious diseases. In this regard, we explored the localization of PDGF in KD. Sections of skin biopsies from 8 KD patients were investigated using the mouse anti-human PDGF monoclonalantibody. Samples obtained in the acute stage were stained with ABC technique. As a result, PDGF was detected in the dermis and epidermis. The prominent staining for PDGF was found in the endothelial lining of dilated qapillaries and in the cytoplasm of macrophages~ Positive staining for PDGF was particularly associated with macrophage infiltrates. In conclus{on, it is postulated that PDGF may play an importantrole in the initiation and/or healing process of vasculitis, and skin lesion in KD. { 28

Expression of platelet-derived growth factor and its receptor in AIDS-related Kaposi sarcoma in vivo suggests paracrine and autocrine mechanisms of tumor maintenance.

As previously described, proliferation of Kaposi sarcoma (KS)-derived cells in vitro is dependent on the presence of platelet-derived growth factor (PDGF). To test the hypothesis that PDGF may also be a major growth factor for KS cells in vivo, we performed in situ hybridization and immunohistochemical staining for PDGF and PDGF receptors in tissue sections of AIDS-related KS. The data suggest that KS consists of two types of tumor cells. (i) The main population are spindle-shaped cells with elongated nuclei (KS-s cells). They reveal a strong expression of PDGF beta receptors but do not express the PDGF-A and PDGF-B isoforms. (ii) A minor population of KS cells express PDGF beta receptor as well as PDGF-A and PDGF-B (KS-p cells). These cells are often grouped in whorls and surrounding vascular slits. They reveal spherical nuclei with evenly distributed chromatin and inconspicuous nucleoli. PDGF alpha receptor is not expressed in either form of KS cells. The results suggest that the isoforms of PDGF and the PDGF beta receptor are differentially expressed in two different cell types in KS and that PDGF isoforms may contribute to the pathogenesis of KS.

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.