Platelet-derived Growth Factor Proteins Research Articles

Antioxidant and Anticancer Potentials of the Olive and Sesame Mixture against Dimethylhydrazine-Induced Colorectal Cancer in Wistar Rats.

Cancer is one of the leading causes of death worldwide, and natural agents have shown some promise in fighting it. Thus, the present study tried to evaluate the healing potential of an equal combination of olive and sesame extract (MOS) against the colorectal cancerous lesions that were induced by dimethylhydrazine (DMH) in male rats and also compare the anticarcinogenic potential of the MOS and vitamin E with each other. Therefore, the mixture of equal olive and sesame extract (MOS) was used as the main treatment, alongside vitamin E as a parallel treatment. This study examined the red blood cell (RBC) and white blood cell (WBC) levels, biochemical indices, lactate dehydrogenase (LDH), C-reactive protein (CRP), total protein (TP), creatine kinase (CPK), albumin, and the colon tissue pathology, as well as the level of protein expression of the adenomatous polyposis coli (APC), proliferating cell nuclear antigen (PCNA), carcinoembryonic antigen (CEA), and platelet-derived growth factor (PDGF). Also, the tissue stress markers including total antioxidant capacity (TAC), malondialdehyde (MDA), and superoxide dismutase (SOD) were analyzed. Overall, the results represented a significant reduction in the congestion, mitotic index, inflammation, and cell destruction in the MOS group compared to the DMH group. In terms of the oxidative stress level, a significant increase was observed in the DMH group in comparison with the DMH-MOS group (P < 0.05), and the MOS significantly increased TAC level (P < 0.05). Furthermore, the DMH+MOS-exposed group exhibited a significantly lower expression of the PCNA, CEA, and PDGF proteins than those of the DMH group. Overall, the MOS showed that it can effectively prevent DMH-induced colon lesions. This mixture, as a strong antioxidant agent, can be clinically applied for preventing and treating colorectal cancer, the effectiveness of which is higher than that of vitamin E.

Poster 101: Human pluripotent stem cell-derived chondroprogenitors committed to form distinct types of chondrocytes for the future cartilage tissue engineering

Objectives:Chondrogenic mesenchymal cells have been developed from human pluripotent stem cells (hPSCs) as a human model of embryonic chondrogenesis. These cells are expandable in the presence of fibroblast growth factor (FGF), and the types of cartilage they tend to generate have been characterized in vitro and in vivo. Resistance to endochondral ossification is one of the most important characteristics of the tissue engineered cartilage for cartilage regenerative therapy. Despite some reports on directed derivation of endochondral ossification resistant chondrocytes from hPSCs, their precursors (chondrogenic mesenchymal cells) have been poorly characterized. We have developed methods to generate two types of chondrogenic mesenchymal cells that lead to cartilage pellets expressing high or very low levels of hypertrophic chondrocyte markers in vitro. These cartilage pellets, when transplanted, became either fully mineralized bony tissue or remained unmineralized, respectively. Here, we present results from cell-type analyses of the two types of mesenchymal cells using the genome-wide RNA-seq technology, aiming to provide mechanistical insights into how the two types of mesenchymal cells prefer to form different types of cartilage.Methods:Human PSC lines were differentiated toward paraxial mesodermal progeny in a chemically-defined medium (CDM) as previously described (1, 2), and mesodermal cells were isolated by cell sorting (2, 3). The isolated cells were then cultured in CDM containing FGF2, platelet-derived growth factor (PDGF), transforming growth factor-beta (TGF-beta) inhibitor and glycogen synthase kinase (GSK) inhibitor (1), which were then subjected to CDM containing FGF2, TGFbeta inhibitor and GSK inhibitor (FSbC medium) as well as to CDM containing PDGF and BMP inhibitor (PN medium) to generate mesenchymal cells of distinct size and shape. The resulting cells were subjected to RT-PCR and bulk RNA-seq analyses for cell-type determination. Chondrogenesis was done by pellet culture using PDGF, TGF-beta3 and bone morphogenetic proteins, as described previously (1, 2). Some cartilage pellets were fixed, sectioned, and stained with Toluidine Blue and von Kossa, or immunostained with anti-collagen type I, II and X (COL1, 2, 10) antibodies. DNA, RNA, and sulfated glycosaminoglycan were isolated and quantified from unfixed cartilage pellets. Some cartilage pellets were also subcutaneously transplanted into NSG (severely immunocompromised) mice and the recovered cartilage pellets were similarly analyzed. All the methods involved were previously described in detail (1, 2).Results:The PN cultured mesenchymal cells (Fig. 1) developed COL2+ cartilage pellets that expressed COL10A1 at 10 to 100-fold lesser levels and PRG4 (Lubricin gene) at higher levels than those derived from the FSbC cultured mesenchymal cells, and were resistant to endochondral ossification after subcutaneous transplantation for 8 weeks (Fig. 2) (1, 2). In contrast, the FSbC cultured mesenchymal cells (Fig. 1) developed cartilage pellets that expressed COL10A1 at higher levels than those derived from the PN cultured mesenchymal cells, and were readily mineralized after subcutaneous transplantation (Fig. 2) (1, 2). The comparative RNA-seq analyses of the two mesenchymal cell populations showed that the FSbC cells resembled ectomesenchymal cells derived from neural crest progeny of hPSCs (1, 2) and the PN cells consisted of cells resembling mesenchymal stromal cells and ligament/tendon progenitors (Fig. 3).Conclusions:From hPSCs, chondrogenic mesenchymal cells of distinct chondrogenesis activity can be generated in vitro in a controlled fashion. One type leads to chondrocytes prone to endochondral ossification, and the other type leads to rather stable cartilage that seems to resist endochondral ossification. Our results suggest that the fate of hPSC-derived chondrocytes can be controlled at the mesenchymal cell (i.e., chondrocyte precursor)-stage, which seems to be dependent on how the mesenchymal cells are generated from hPSCs and maintained in culture. Further biological studies on these cells will not only enable hPSC-derived chondrocytes/chondroprogenitors to be used directly for cartilage regenerative therapy, but also may lead to a critical mechanism that allows therapeutically relevant adult chondrogenic cells such as skeletal stem cells to reproducibly regenerate hyaline permanent cartilage during cartilage repair.Fig. 1Fig. 2Fig. 3

Antioxidant and Anticarcinogenic Potentials of Propolis for Dimethylhydrazine-Induced Colorectal Cancer in Wistar Rats.

Propolis is a natural compound with anticarcinogenic properties. The present study aimed to compare the inhibitory effect of ethanolic extract of propolis (EEP) and vitamin E on dimethylhydrazine-induced colon lesions in rats. In this study, 60 rats were randomly categorized into six 10-member groups. After 13 weeks, blood and colon tissue were sampled to examine some factors. The parameters included red (RBC) and white (WBC) blood cell profile, lactate dehydrogenase (LDH), C-reactive protein (CRP), total protein (TP), creatine kinase (CPK), and albumin, as well as the extent of colon histological lesions, protein expression (adenomatous polyposis coli (APC), proliferating cell nuclear antigen (PCNA), carcinoembryonic antigen (CEA), and platelet-derived growth factor (PDGF)), and oxidative stress markers (total antioxidant capacity (TAC), malondialdehyde (MDA), and superoxide dismutase (SOD)) in colon tissue. A significant decrease was observed in congestion, mitotic index, inflammation, and cell destruction in colon tissue in dimethylhydrazine group in comparison with the control group (P < 0.05). The EEP exposed rats exhibited a significant lower oxidative stress than the DMH group (P < 0.05). Furthermore, the extract significantly affected TAC level (P < 0.05). While the expression level of APC rose substantially in the EEP-treated group compared to the DMH group, the level of PCNA, CEA, and PDGF proteins significantly reduced. It seems that the EEP can efficiently prevent DMH-induced colonic lesions. Furthermore, its effectiveness is more than the vitamin E, which is a strong antioxidant.

Increased growth factors play a role in wound healing promoted by noninvasive oxygen-ozone therapy in diabetic patients with foot ulcers.

Management of diabetic foot ulcers (DFUs) is a great challenge for clinicians. Although the oxygen-ozone treatment improves the diabetic outcome, there are few clinical trials to verify the efficacy and illuminate the underlying mechanisms of oxygen-ozone treatment on DFUs. In the present study, a total of 50 type 2 diabetic patients complicated with DFUs, Wagner stage 2~4, were randomized into control group treated by standard therapy only and ozone group treated by standard therapy plus oxygen-ozone treatment. The therapeutic effects were graded into 4 levels from grade 0 (no change) to grade 3 (wound healing). The wound sizes were measured at baseline and day 20, respectively. Tissue biopsies were performed at baseline and day 11. The expressions of vascular endothelial growth factor (VEGF), transforming growth factor-β (TGF-β), and platelet-derived growth factor (PDGF) proteins in the pathologic specimens were determined by immunohistochemical examinations. The effective rate of ozone group was significantly higher than that of control group (92% versus 64%, P < 0.05). The wound size reduction was significantly more in ozone group than in control group (P < 0.001). After treatment, the expressions of VEGF, TGF-β, and PDGF proteins at day 11 were significantly higher in ozone group than in control group. Ozone therapy promotes the wound healing of DFUs via potential induction of VEGF, TGF-β, and PDGF at early stage of the treatment. (Clinical trial registry number is ChiCTR-TRC-14004415).

Micromechanical PDGF recognition via lab-on-a-disc aptasensor arrays

A plug-and-play CD-like platform is used to perform a statistical detection of platelet derived growth factor (PDGF) proteins through aptamer-based surface functionalization of multiple microcantilever arrays. When PDGF proteins bind to aptamer coatings, the cantilevers deflect. The deflection response is monitored by optical read-out units from a commercial DVD-ROM device. We report on the use of an improved sensing platform which facilitates measurements under continuous liquid flow and with temperature control. Also, the mechanical wobbling of the DVD-ROM platform has been minimized and the scanning system has been optimized in order to detect cantilever deflections in liquid with nanometer scale resolution. The capability of the sensing platform is demonstrated by detection of clinically relevant concentrations of PDGF proteins. We present statistical measurements on 100 microcantilevers at different concentrations of PDGF, ranging from 10nM to 400nM. Hereby it is possible to reliably characterize the averaged mechanical response of cantilevers as a function of protein concentration.

Platelet-derived growth factor may be a potential diagnostic and prognostic marker for cholangiocarcinoma

Our previous report showed that platelet-derived growth factor (PDGF) and related genes were upregulated in a Syrian hamster model and could be detected in all human cholangiocarcinoma (CCA) tissues. We therefore hoped that PDGF could be used as a diagnostic and prognostic marker. We analyzed 78 samples of human CCA and adjacent tissues for PDGF and related gene expression, and localized PDGF protein expression. The mechanism of anti-cancer drugs on PDGF and related genes or proteins in CCA cell lines (OCA17, M156, and KKU100) was studied through MTT cell viability assay, quantitative real-time PCR, and immunoblotting. Mutagenesis of the PDGFRA coding region was analyzed. Moreover, the PDGFRA in sera of CCA patients and healthy controls was investigated. PDGFA was found to be upregulated in CCA tissue (84.6 %). Positive PDGFA immunohistochemical staining was significantly correlated with status (P = 0.000), stage of CCA (P = 0.013), metastasis (P = 0.017), and short survival rate (P = 0.005), and the multivariate analysis confirmed that PDGFA positive immunostaining had a higher likelihood of the risk of death (HR = 2.907, P = 0.016). For DNA point mutation of the PDGFRA sequence, silent mutations were found at tyrosine kinase 2 V824V (exon 18) and A603A (exon 13), and a missense mutation in S478P (exon 10); there was only a missense mutation in S478P (29 %) that has significant correlation with the histopathological grading (P = 0.037) and positive immunoreactive PDGFA (P = 0.021). In vitro cell line study by immunowestern blotting found that sunitinib malate had an inhibitory effect on the PDGFA pathway by decreasing p-PDGFRA, AKT, and p-AKT expression. The serum level of PDGFA in CCA patients was significantly higher than those of healthy control by 1.4-fold (P = 0.014). The present results suggest that PDGFA and PDGFRA may be used for CCA prognosis and/or as diagnostic candidate markers.

Gene therapy with adenoviral plasmids or naked DNA of vascular endothelial growth factor and platelet-derived growth factor accelerates healing of duodenal ulcer in rats.

After we demonstrated that daily intragastric administration of angiogenic growth factors like basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF), or vascular endothelial growth factor (VEGF) accelerated the healing of chronic duodenal ulcers in rats, we hypothesized that a single dose of gene therapy related to these growth factors may be enough to accelerate the healing of duodenal ulcers through enhancement of synthesis of endogenous angiogenic growth factors. Thus, we compared the effects of intraduodenal or intravenous adenoviral vectors and naked DNA transducing the genes for either VEGF or PDGF in experimental duodenal ulcers induced by cysteamine in rats. Sprague-Dawley female rats with confirmed duodenal ulcers were randomly divided into control and treatment groups. The controls received either intraduodenal injection of buffer or the beta-galactosidase-transducing adenoviral vector. Rats treated with a single or double dose of adenoviral vector or naked DNA of VEGF or PDGF had significantly smaller ulcers than the controls. Histologic analysis demonstrated that reepithelized granulation tissue with prominent angiogenesis replaced the ulcers. Western blotting, immunohistochemistry, and enzyme-linked immunosorbent assay of duodenal mucosa confirmed that the expression of VEGF or PDGF proteins was enhanced by the transgenes, whereas beta-galactosidase staining in multiple organs identified that the transgenes, especially after local administration were only localized in the duodenum, stomach, and jejunum. These results suggest that gene therapy with either VEGF or PDGF may be a rapid approach to achieve duodenal ulcer healing.

Biology of platelet-derived growth factors in development.

Platelet-derived growth factor (PDGF) was one of the first growth factors to be characterized, and the PDGF family of ligand and receptors has remained an archetype system for studies of the mechanisms of action of growth factors and receptor tyrosine kinases for more than two decades. The small size of the family has also facilitated genetic studies and, in particular, manipulations of the mouse PDGF and PDGF receptor genes have given important insights into the role of this family during mammalian development. These studies have shown that discrete populations of mesenchymal and neuroectodermal progenitor cells depend on PDGF signaling for their growth and distribution within developing organs. Other studies suggest that the same, or similar, cells may be targeted by exaggerated PDGF signaling in a number of pathological processes, including different types of cancer. The present review summarizes current views on the roles of PDGFs in developmental processes, and discusses the critical importance of the amount, spatial distribution, and bioavailability of the PDGF proteins for acquisition of the correct number and location of target cells.

Expression of platelet-derived growth factor (PDGF) in the epididymis and analysis of the epididymal development in PDGF-A, PDGF-B, and PDGF receptor beta deficient mice.

The platelet-derived growth factor (PDGF) family of ligands and receptors play a pivotal role in the development of various organs. The critical importance of the PDGF-mediated signaling during embryonic development and adult physiology of the kidney and the common mesonephric origin of the epididymis and kidney prompted us to investigate the immunohistochemical localization of PDGF A- and B-chain and PDGF receptor (PDGFR) alpha- and beta-subunit in rat and mouse epididymis, the expression profiles of the corresponding mRNAs, and the consequences of a loss-of-function mutation at the PDGF-A, PDGF-B, and PDGFR-beta loci on mouse epididymis phenotypic appearance. Prenatally, PDGF-A and PDGFR-alpha immunohistochemical staining was seen in both species, whereas PDGF-B and PDGFR-beta were absent. The cellular localization of PDGF-A within the epithelium and the alpha-receptor in the mesenchyme in either mouse or rat before birth suggests that the PDGF-A/PDGFR-alpha system might be involved in the epididymal epithelial-mesenchymal interaction during the fetal period of life. Postnatally, PDGF A- and B-ligand and PDGFR alpha- and beta-subunit were confined in the epithelium. The identity of PDGF and PDGFR proteins were further confirmed by immunoblotting. In line with the immunohistochemical studies, PDGF-A and PDGFR-alpha mRNAs were seen by reverse transcription-polymerase chain reaction in rat and mouse tissue before birth, whereas PDGF-B and PDGFR-beta were almost not detectable. During the first days of life, PDGF-B and PDGFR-beta genes started to appear, and the overall trend in mRNA expression throughout postnatal development showed that the transcripts levels for PDGF-A, PDGF-B, PDGFR-beta, and PDGFR-alpha were constant with the only exception of a progressive decrease of PDGFR-alpha in adult rats. The PDGF-A null mutation strongly influenced the epididymal phenotype starting from puberty; only fetal PDGF-B and PDGFR-beta -/- mice were available, and no differences were seen in the epididymis of these animals, compared with wild-type littermates. Taken together, these data indicate that the PDGF system is highly expressed in the epididymis and suggest that PDGF could be involved in the maintenance of morphological structure and functional control of this organ.

Expression of platelet-derived growth factor proteins and their receptor alpha and beta mRNAs during fracture healing in the normal mouse.

Platelet-derived growth factor (PDGF), abundant in bone tissue, has been reported to stimulate mesenchymal cell proliferation and migration. To elucidate the functional roles of PDGF during fracture healing, we investigated the expression of PDGF-A and -B chain proteins and receptor alpha and beta mRNAs in fractured mouse tibiae. Twelve-week-old male BALB/c mice were operated on to make a closed fracture on the proximal tibia. On days 2, 4, 7, 10, 14, 21, and 28 after the operation, the fractured tibiae were excised, fixed with 4% paraformaldehyde, decalcified with 20% EDTA, and embedded in paraffin to prepare 7-microm sections. Immunohistochemistry using polyclonal antibodies against human PDGF-A and -B chains was carried out by the avidin-biotin-peroxidase method. For in situ hybridization, we used digoxigenin-labeled single-stranded DNA probes specific for mouse PDGF receptors alpha and beta generated by unidirectional polymerase chain reaction. In the inflammatory phase on days 2-4 after the fracture, mesenchymal cells gathering at the fracture site expressed the PDGF-B chain and beta receptor mRNA. At the stage of cartilaginous callus formation on day 7, the immunoreactivity for PDGF-A and -B chains on proliferating and hypertrophic chondrocytes and the signals of alpha and beta receptor mRNAs on proliferating chondrocytes became manifest. At the stage of bony callus and bone remodeling on days 14-21, the predominant expression of the PDGF-B chain and beta receptor was observed on both osteoclasts and osteoblasts. On day 28, signals for PDGF ligand proteins and receptor mRNAs diminished. The coincidental localization of PDGF ligands and their receptors implies a paracrine and autocrine mechanism. Our data suggested that PDGF contributed in part to the promotion of the chondrogenic and osteogenic changes of mesenchymal cells from the early to the midphase of fracture healing; the functions mediated by the beta receptor, including cell migration, might be prerequisites to the recruitment of mesenchymal cells in the initial step and to the interaction between osteoclasts and osteoblasts in the bone remodeling phase.

The role of integrins in saphenous vein vascular smooth muscle cell migration

Smooth muscle cell (SMC) migration is an essential feature of the intimal hyperplastic process that so frequently limits the patency of vascular reconstructions. The purpose of this investigation was to evaluate the effect of a series of integrins, or cell surface receptors that mediate cellular attachment, on platelet-derived growth factor (PDGF) and extracellular matrix (ECM) protein-induced migration of human SMCs. Immunofluorescence staining was used to search for various integrins and subunits on the surface of SMCs derived from human saphenous vein. Chemotaxis and haptotaxis of SMCs to various matrix proteins and PDGF were assayed using a 48-well microchemotaxis chamber in the presence or absence of antibodies that blocked the function of these integrins. Several subunits (beta 1, alpha 2, alpha 5) and one integrin (alpha v beta 3) were identified in saphenous vein SMCs. The beta 1 integrin antibody inhibited chemotaxis to collagen I and IV, laminin, and PDGF. The alpha 2 integrin antibody inhibited collagen I and IV, and laminin-induced chemotaxis. The alpha 5 integrin antibody had no effect on SMC migration. The alpha v beta 3 integrin antibody inhibited chemotaxis to PDGF but not to the ECM proteins. Integrins are necessary for SMC migration induced by PDGF and ECM proteins. The integrin or subunits responsible for facilitating migration varies with the stimulant. Agonists designed to inhibit integrin function might be used to suppress SMC migration and suppress the formation of intimal hyperplasia.

Enhanced synthesis of platelet-derived growth factor following injury induced by 6-hydroxydopamine in rat brain

The kinetics of platelet-derived growth factor messenger RNA synthesis in the substantia nigra and in the striatum, before and after unilateral intranigral 6-hydroxydopamine injection, was studied and compared with that after sham operation by a quantitative reverse transcription–polymerase chain reaction. The kinetics of brain-derived neurotrophic factor messenger RNA was studied as a comparison. Furthermore, the expression of platelet-derived growth factor A- and B-chain proteins was analysed by enzyme-linked immunosorbent assay and immunohistochemistry. In the ipsilateral striatum of 6-hydroxydopamine-lesioned rats, the signal density of messenger RNA for both A- and B-chains had already increased at one day and remained at an elevated level during the observation period of four weeks. In the substantia nigra ipsilateral to the lesion, a strongly increased level of B-chain and, to a lesser extent, of A-chain messenger RNA was already detected at 4 h, reaching a maximal level at one day. No significant increase was seen either in sham-operated rats or in the contralateral striatum and substantia nigra. Amounts of platelet-derived growth factor proteins were examined separately by enzyme-linked immunosorbent assay in both sides of the substantia nigra, striatum and cortex. Three days after 6-hydroxydopamine lesions the levels of both platelet-derived growth factor A- and B-chains increased in the ipsilateral striatum, substantia nigra, and cortex. An increase in the A-chain was also observed in the contralateral side of the brain. The signal for brain-derived neurotrophic factor messenger RNA increased in the striatum in the lesioned side and, to a lesser extent, in the contralateral side, as well as the in the substantia nigra, where a significant difference was observed when compared with the contralateral side. Semiquantitative immunohistochemical analysis on the substantia nigra confirmed the enhanced platelet-derived growth factor expression, revealing that the majority of the platelet-derived growth factor-producing cells were neurons. In summary, we have shown that platelet-derived growth factor messenger RNA as well as its protein are induced after injury to dopaminergic cells. These data indicate an important role of platelet-derived growth factor in the dopaminergic system.

Localization of platelet-derived growth factor and insulin-like growth factor I in the fibrotic lung.

To better understand the mechanisms responsible for the increase in numbers of fibroblasts and increased collagen synthesis in fibrotic intestitial lung diseases, platelet-derived growth factor (PDGF)-A and PDGF-B, PDGF receptor-alpha and -beta, insulin-like growth factor I (IGF-I), and IGF-I receptor were evaluated immunohistochemically. Additionally, the messenger ribonucleic acids (mRNAs) for PDGF-A and PDGF-B, PDGF receptor-alpha and -beta, and IGF-I were investigated by in situ hybridization in alveolar macrophages and lung tissues from patients with interstitial lung disease. In specimens of bronchoalveolar lavage fluid (BALF), PDGF-A, PDGF-B, and IGF-I were local in alveolar macrophages in patients with idiopathic pulmonary fibrosis (IPF), patients with sarcoidosis (Sar), and normal individuals. Although there were no differences between IPF and Sar in terms of the staining intensity or number of positive cells, the number of such cells was smaller in the normal controls. In lung tissues with early-stage IPF, PDGF and IGF-I proteins were localized exclusively in alveolar macrophages, mononuclear phagocytes, fibroblasts, alveolar Type II cells, vascular endothelial cells, and vascular smooth-muscle cells. In lung tissues with late-stage IPF and those from normal controls, only alveolar macrophages contained PDGF and IGF-I proteins. Interestingly, the cellular localizations of PDGF receptor-alpha and -beta, and of IGF-I receptor were the same as those of the PDGF and IGF-I proteins in early-stage IPF, whereas these cells were not positive for any of these substances in late-stage IPF or normal controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of tumor necrosis factor and interferon-gamma on platelet-derived growth factor (PDGF) A and B gene expression in THP-1 monocytic cells

Platelet-derived growth factor (PDGF) is a potent mitogen of mesenchymal cells. A and B chains of PDGF are transcribed in atherosclerotic plaques but details of the mechanism of the expression of either chains in vivo are not understood. It has been suggested that differentiated macrophages, which are the major origin of foam cells, but not monocytes hardly secrete PDGF in vitro. In the present study, we investigated the effect of 1) tumor necrosis factor-alpha (TNF-alpha), a macrophage derived cytokine which promote the differentiation from monocytes to macrophages, and 2) interferon-gamma (IFN-gamma), a T cell derived macrophage activating factor, on the production and mRNA expression of PDGF A and B chains in a human monocytic leukemia cell line THP-1. THP-1 cells were cultured in the presence of one of the experimental cytokines with or without the addition of phorbol 12-myristate 13-acetate (PMA). PMA is known to induce the differentiation of monocytes (including THP-1 cells) to macrophages within a period of 24 hours. Thus the treatment of THP-1 cells with a cytokine and/or PMA in vitro was expected to mimic the effect of the cytokine during the differentiation of monocytes to macrophages in vivo. Unstimulated THP-1 cells expressed no detectable amount of PDGF A or B mRNA. The treatment of THP-1 cells with PMA resulted in the strong expression of both PDGF A and B mRNAs. Stimulated with TNF-alpha (50 U/ml) on day 0, THP-1 cells expressed PDGF A mRNA significantly from 1 up to 3 days after the treatment, whereas the expression of PDGF B and mRNA was only weakly detected on day 2. The maximum signal of PDGF A mRNA was ten times stronger than that of PDGF B mRNA. Within the same period, relatively small number of TNF-alpha treated THP-1 cells had differentiated to macrophages as detected by the reduction of nitro blue tetrazolium (NBT) (35% of total cells) and by the counting of adherent cells (35% of total cells). THP-1 cells simultaneously treated with TNF-alpha and PMA strongly expressed PDGF A and B gene. Most remarkably, the expression of PDGF B mRNA was greatly enhanced with compared to the treatment with PMA alone. In addition, TNF-alpha augmented the secretion of PDGF proteins in PMA stimulated THP-1 cells. In contrast, INF gamma per se had no effect on PDGF gene expression, NBT reducing activity and cell adherence in unstimulated THP-1 cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Receptors for platelet-derived growth factor on microvascular endothelial cells.

Endothelial cells are widely thought to be unresponsive to platelet-derived growth factor (PDGF) and devoid of PDGF receptors. However, in examining the growth factor responses of microvascular endothelial cells isolated from human omental adipose tissue, we detected PDGF-induced tyrosine phosphorylation of an 180-kD glycoprotein, subsequently identified as the cellular receptor for PDGF by specific immunoprecipitation. Scatchard analysis of 125I-PDGF binding to human microvascular endothelial cells revealed 30,000 PDGF receptors/cell with a kD of 0.14 nM. PDGF stimulated tyrosine phosphorylation of PDGF and other cellular proteins in a dose- and time-dependent manner, with half-maximal receptor phosphorylation occurring at 0.3 nM recombinant human PDGF-BB within 1 min of PDGF exposure. Normal cellular consequences of receptor activation were also observed, including tyrosine phosphorylation of a 42-kD protein and serine phosphorylation of ribosomal protein S6. Furthermore, PDGF was mitogenic for these cells. The finding of functional PDGF receptors on human microvascular endothelial cells suggests an important direct role for PDGF in neovascularization.

Human iliac artery endothelial cells express both genes encoding the chains of platelet‐derived growth factor (PDGF) and synthesize PDGF‐like mitogen

In human umbilical vein and bovine aortic endothelial cells in culture c-sis gene expression and secretion of platelet-derived growth factor (PDGF) has been previously demonstrated. We now report the presence of PDGF-1 and PDGF-2/sis mRNA transcripts in primary cultures of human iliac artery endothelial cells (HIA-EC). Concomitantly, these cells synthesize and secrete PDGF-like proteins identified by direct immunoprecipitation with specific PDGF antiserum. The PDGF proteins secreted by HIA-EC have molecular weights of 31 and 35 kd under nonreducing conditions. Upon reduction these proteins are converted to the monomeric 15- and 16-kd forms. Conditioned media derived from HIA-EC stimulated the incorporation of 3H-thymidine by 3T3 cells and competed with 125I-PDGF for its binding to 3T3 cell membrane receptors. The biologic activity was stable to heating at 100 degrees C for 10 min and sensitive to reducing agents, properties similar to those of authentic PDGF. Production of PDGF-like mitogen by the human arterial endothelial cells may play an important role in the paracrine modulation of arterial wall regeneration following vascular injury.