A role of heparin-binding epidermal growth factor-like growth factor in cardiac remodeling after myocardial infarction

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is known to induce cell growth in various cell types via transactivation of epidermal growth factor receptor (EGFR). To investigate the involvement of HB-EGF and EGFR in cardiac remodeling after myocardial infarction (MI), we examined the expressions of mRNA and protein in rat hearts 6 weeks after MI-induction. Where increased expressions of HB-EGF mRNA and protein were observed, infarcted myocardium was replaced by extracellular matrix and interstitial fibroblasts. EGFR mRNA and protein expression did not show significant changes in sham-operated heart tissues, non-infarcted region, and infarcted region. In vitro study demonstrated that HB-EGF mRNA was expressed mainly in cultured fibroblasts rather than in myocytes. We suggest that the interaction between HB-EGF and EGFR transactivation is closely related to the proliferation of cardiac fibroblasts and cardiac remodeling after MI in an autocrine, paracrine, and juxtacrine manner.     

Platelet-derived growth factor.

Platelet-derived growth factor (PDGF) is a potent activator for cells of mesenchymal origin. Two different PDGF chains termed A and B encoded by different genes have been identified leading to three different PDGF isoforms, the AA and BB homodimers and the AB heterodimer. All three forms have been observed in vivo and possess biological activity in vitro with the AA homodimer being the poorest cellular mitogen. The availability of highly purified recombinant PDGF isoforms was the initial basis for comparative studies in order to specify the different spectra of activity of the various PDGF species. This review is particularly focused on the AB heterodimer as from the standpoint of heterologous gene expression, this species is the one with the highest demands concerning expression and purification protocols. This explains the fact that, in comparison to PDGF-BB, only very limited data on the in vivo application of PDGF-AB are available so far.     

Proteomic analysis reveals late exercise effects on cardiac remodeling following myocardial infarction

Exercise has been shown to improve function of the left ventricle (LV) following myocardial infarction (MI). The mechanisms to explain this benefit have not been fully delineated, but may involve improved mechanics resulting in unloading effects and increased endothelial nitric oxide synthase levels [1,2]. Accordingly, the goal of this study was to determine how the LV infarct proteome is altered by a post-MI exercise regimen. Sprague–Dawley rats underwent ligation of the left descending coronary artery to induce MI. Exercise training was initiated four weeks post-MI and continued for 8 weeks in n = 12 rats. Compared with the sedentary MI group (n = 10), the infarct region of rats receiving exercise showed 20 protein spots with altered intensities in two-dimensional gels (15 increased and 5 decreased; p < 0.05). Of 52 proteins identified in 20 spots, decreased levels of voltage-dependent anion-selective channel 2 and increased levels of glutathione perioxidase and manganese superoxide were confirmed by immunoblotting. Cardiac function was preserved in rats receiving exercise training, and the beneficial effect was linked with changes in these 3 proteins. In conclusion, our results suggest that post-MI exercise training increases anti-oxidant levels and decreases ion channel levels, which may explain, in part, the improved cardiac function seen with exercise.     

Platelet-derived endothelial cell growth factor.

Platelet-derived endothelial cell growth factor (PD-ECGF) is a 45 kDa single chain polypeptide which stimulates endothelial cell growth and chemotaxis in vitro and angiogenesis in vivo. Analysis of a full length PD-ECGF cDNA revealed an open reading frame coding for 482 amino acids without homology to other known proteins. No signal sequence was observed, and analysis of the biosynthesis and processing of PD-ECGF in a thyroid carcinoma cell line revealed that PD-ECGF is released only very slowly. PD-ECGF becomes covalently associated with nucleotide triphosphates (e.g., ATP) in vivo, as well as in vitro. The physiological significance of this posttranslational modification remains to be elucidated. The tissue distribution and target cell specificity of PD-ECGF suggest roles in angiogenesis (e.g., during wound healing and in the developing placenta), as well as in the maintenance of the integrity of the endothelial cell lining of large vessels.     

血小板源生长因子受体与肿瘤

血小板源生长因子(platelet-derived growth factor,PDGF)经由其受体(platelet-derived growth fac tor receptor,PDGFR)表现细胞效应。PDGF和PDGFR涉及多种肿瘤的发病机制并在血管生成中起重要作用。PDGF在肿瘤中的自分泌刺激、PDGFR的过表达或过度活化或者刺激肿瘤内血管生成都会促进肿瘤生长;PDGFR的阻断可以降低实体瘤中组织间质液压而增强药物传送。这些机制可能提示在肿瘤治疗中PDGFR抑制剂单用、与化疗药物或者和其他靶点药物联合用药的可能性和可行性。随着PDGFR拮抗剂,如imatinib的上市,PDGFR作为抗肿瘤药物的靶点备受瞩目。     

Platelet-derived growth factor signaling and human cancer

Platelet-derived growth factor (PDGF) is a critical regulator of mesenchymal cell migration and proliferation. The vital functions of PDGFs for angiogenesis, as well as development of kidney, brain, cardiovascular system and pulmonary alveoli during embryogenesis, have been well demonstrated by gene knock-out approaches. Clinical studies reveal that aberrant expression of PDGF and its receptor is often associated with a variety of disorders including atherosclerosis, fibroproliferative diseases of lungs, kidneys and joints, and neoplasia. PDGF contributes to cancer development and progression by both autocrine and paracrine signaling mechanisms. In this review article, important features of the PDGF isoforms and their cell surface receptor subunits are discussed, with regards to signal transduction, PDGF-isoform specific cellular responses, and involvement in angiogensis, and tumorstromal interactions.     

Platelet-derived growth factor in human malignancy.

Platelet-derived growth factor (PDGF) was first implicated in the process of transformation when one of its peptide chains was found to be homologous to the viral sis oncogene (v-sis). Since that time, there have been multiple demonstrations of the transforming activity of v-sis in fibroblasts. Because of the near identity of the v-sis protein with the PDGF B chain, v-sis is thought to transform through an autocrine stimulatory mechanism of cell growth. Consistent with this view are studies which demonstrate inhibition of v-sis-mediated transformation by anti-PDGF antibodies. Expression of the cellular sis gene (c-sis) and its receptors, and secretion of PDGF-like factors have been demonstrated in many types of human malignant cells. Nevertheless, a causative role for c-sis in inducing or maintaining the transformed phenotype in human malignancies remains to be established. There are significant differences in structure between v-sis and c-sis. Studies of transforming ability have yielded conflicting results in transfection models, depending on the transfected vector and target cell type utilized. While there is compelling evidence for the involvement of PDGF in an autocrine growth mechanism in transformed fibroblasts, the evidence in human epithelial tumor types is less convincing because PDGF receptors are usually not detectable on the cell surface. The recent demonstration of intracellular co-localization of active PDGF precursors and PDGF receptors, however, supports the existence of an internal autocrine pathway independent of PDGF secretion. Further investigation of such a mechanism in de novo human malignancies is warranted to establish the role of PDGF in the development of these neoplasms.     

Platelet-derived growth factor in chemotactic for fibroblasts

Chemotaxis assays in modified Boyden chambers were used to detect fibroblast chemoattractants in materials released from early-stage inflammatory cells, namely, mast cells, platelets, and neutrophils. Strong attractant activity was found in substances released from platelets. This activity was accounted for mainly by the platelet- derived growth factor (PDGF), which is released from the platelets and which was active as a chemoattractant at 0.5-1.0 mitogenic units/ml. The mitogenic activity of purified PDGF, measured by [3H]thymidine incorporation, occurs at a similar concentration range. By varying the gradient of PDGF, we demonstrated that PDGF stimulates chemotaxis rather than random motility. Preincubation of suspensions of fibroblasts in the presence of PDGF decreased the subsequent migration of cells to a gradient of PDGF as well as to a gradient of fibronectin, which is also in attractant for fibroblasts. The chemotactic response of fibroblasts to PDGF was not inhibited by hydroxyurea or azidocytidine but was inhibited by actinomycin D and cycloheximide, suggesting that synthesis of RNA and proteins but not of DNA is required for the chemotactic response to occur. Fibroblast growth factor, epidermal growth factor, nerve growth factor, and insulin were not chemotactic for human skin fibroblasts, suggesting that the chemoattractant activity of PDGF for fibroblasts is not a general property of growth factors and mitogens. These results suggest that PDGF could have two functions in wound healing: to attract fibroblasts to migrate into the clot and then to induce their proliferation.     

Transplanted embryonic stem cells following mouse myocardial infarction inhibit apoptosis and cardiac remodeling

We have previously shown that mouse embryonic stem (ES) cells transplanted following myocardial infarction (MI) differentiate into the major cell types in the heart and improve cardiac function. However, the extent of regeneration was relatively meager compared with the observed functional improvement. Therefore, we hypothesize that mechanisms in addition to regeneration contribute to the functional improvement from ES cell therapy. In this study, we examined the effect of mouse ES cells transplanted post-MI on cardiac apoptosis, fibrosis, and hypertrophy. MI was produced by left coronary artery ligation in C57BL/6 mice. Two different mouse ES cell lines, expressing enhanced green fluorescent protein and beta-galactosidase, respectively, were tested. Post-MI intramyocardial injection of 3 x 10(4) ES cells was compared with injection of medium alone. Terminal deoxynucleotidyl nick end labeling (TUNEL), immunofluorescence, and histology were used to examine the effect of transplanted ES cells on apoptosis, fibrosis, and hypertrophy. Two weeks post-MI, ES cell-transplanted hearts exhibited a significant decrease in TUNEL-stained nuclei (mean +/- SE; MI+medium = 12 +/- 1.5%; MI+ES cells = 6.6 +/- 1%, P < 0.05). TUNEL-positive nuclei were confirmed to be apoptotic by colabeling with a caspase-3 antibody. Cardiac fibrosis was 57% less in the MI+ES cell group compared with the MI + medium group (P < 0.05) as shown with Masson's trichrome staining. Picrosirius red staining confirmed a decreased amount of collagen present in the MI+ES cell group. Cardiomyocyte hypertrophy was significantly decreased following ES cell transplantation compared with medium control animals. In conclusion, transplanted mouse ES cells in the infarcted heart inhibit apoptosis, fibrosis, and hypertrophy, thereby reducing adverse remodeling.     

Platelet-derived growth factor or basic fibroblast growth factor induce anchorage-independent growth of human fibroblasts

Anchorage-independent growth, i.e., growth in semi-solid medium is considered a marker of cellular transformation of fibroblast cells. Diploid human fibroblasts ordinarily do not exhibit such growth but can grow transiently when medium contains high concentrations of fetal bovine serum. This suggests that some growth factor(s) in serum is responsible for anchorage-independent growth. Much work has been done to characterize the peptide growth factor requirements of various rodent fibroblast cells for anchorage-independent growth; however, the requirements of human fibroblasts are not known. To determine the peptide growth factor requirements of human fibroblasts for anchorage-independent growth, we used medium containing serum that had had its peptide growth factors inactivated. We found that either platelet-derived growth factor (PDGF) or the basic form of fibroblast growth factor (bFGF) induced anchorage-independent growth. Epidermal growth factor (EGF) did not enhance the growth induced by PDGF, or did so only slightly. Transforming growth factor beta (TGF-beta) decreased the growth induced by PDGF. EGF combined with TGF-beta induced colony formation in semi-solid medium at concentrations at which neither growth factor by itself was effective, but the combination was much less effective in stimulating anchorage-independent growth than PDGF or bFGF. This work showed that PDGF, or bFGF, or EGF combined with TGF-beta can stimulate anchorage-independent growth of nontransformed human fibroblasts. The results support the idea that cellular transformation may reduce or eliminate the need for exogenous PDGF or bFGF.     

Platelet-derived growth factor receptor-mediated signal transduction from endosomes

Although accumulated evidence supports the concept of endosomal signaling of receptor tyrosine kinases, most results are generated from studies of epidermal growth factor receptor (EGFR). It is not clear whether the concept of endosomal signaling could be generally applied to the other receptor tyrosine kinases. For example, platelet-derived growth factor receptor (PDGFR) is very similar to EGFR in terms of both signaling and trafficking; however, little is known about the endosomal signaling of PDGFR. In this research, we applied the same approaches from our recent studies regarding EGFR endosomal signaling to investigate the endosomal signaling of PDGFR. We showed in this communication that we are able to establish a system that allows the specific activation of endosome-associated PDGFR without the activation of the plasma membrane-associated PDGFR and without disrupting the overall endocytosis pathway. By using this system, we showed that endosomal activation of PDGFR recruits various signaling proteins including Grb2, SHC, phospholipase C-gamma1, and the p85alpha subunit of phosphatidylinositol 3-kinase into endosomes and forms signaling complexes with PDGFR. We also showed that endosomal PDGFR signaling is sufficient to activate the major signaling pathways implicated in cell proliferation and survival. Moreover, we demonstrate that endosomal PDGFR signaling is sufficient to generate physiological output including cell proliferation and cell survival.     

Platelet-derived growth factor receptor regulates salivary gland morphogenesis via fibroblast growth factor expression

A coordinated reciprocal interaction between epithelium and mesenchyme is involved in salivary gland morphogenesis. The submandibular glands (SMGs) of Wnt1-Cre/R26R mice have been shown positive for mesenchyme, whereas the epithelium is beta-galactosidase-negative, indicating that most mesenchymal cells are derived from cranial neural crest cells. Platelet-derived growth factor (PDGF) receptor alpha is one of the markers of neural crest-derived cells. In this study, we analyzed the roles of PDGFs and their receptors in the morphogenesis of mouse SMGs. PDGF-A was shown to be expressed in SMG epithelium, whereas PDGF-B, PDGFRalpha, and PDGFRbeta were expressed in mesenchyme. Exogenous PDGF-AA and -BB in SMG organ cultures demonstrated increased levels of branching and epithelial proliferation, although their receptors were found to be expressed in mesenchyme. In contrast, short interfering RNA for Pdgfa and -b as well as neutralizing antibodies for PDGF-AB and -BB showed decreased branching. PDGF-AA induced the expression of the fibroblast growth factor genes Fgf3 and -7, and PDGF-BB induced the expression of Fgf1, -3, -7, and -10, whereas short interfering RNA for Pdgfa and Pdgfb inhibited the expression of Fgf3, -7, and -10, indicating that PDGFs regulate Fgf gene expression in SMG mesenchyme. The PDGF receptor inhibitor AG-17 inhibited PDGF-induced branching, whereas exogenous FGF7 and -10 fully recovered. Together, these results indicate that fibroblast growth factors function downstream of PDGF signaling, which regulates Fgf expression in neural crest-derived mesenchymal cells and SMG branching morphogenesis. Thus, PDGF signaling is a possible mechanism involved in the interaction between epithelial and neural crest-derived mesenchyme.     

Electrotonic remodeling following myocardial infarction in dogs susceptible and resistant to sudden cardiac death.

Passive electrical remodeling following myocardial infarction (MI) is well established. These changes can alter electrotonic loading and trigger the remodeling of repolarization currents, a potential mechanism for ventricular fibrillation (VF). However, little is known about the role of passive electrical markers as tools to identify VF susceptibility post-MI. This study investigated electrotonic remodeling in the post-MI ventricle, as measured by myocardial electrical impedance (MEI), in animals prone to and resistant to VF. MI was induced in dogs by a two-stage left anterior descending (LAD) coronary artery ligation. Before infarction, MEI electrodes were placed in remote (left circumflex, LCX) and infarcted (LAD) myocardium. MEI was measured in awake animals 1, 2, 7, and 21 days post-MI. Subsequently, VF susceptibility was tested by a 2-min LCX occlusion during exercise; 12 animals developed VF (susceptible, S) and 12 did not (resistant, R). The healing infarct had lower MEI than the normal myocardium. This difference was stable by day 2 post-MI (287 +/- 32 Omega vs. 425 +/- 62 Omega, P < 0.05). Significant differences were observed between resistant and susceptible animals 7 days post-MI; susceptible dogs had a wider electrotonic gradient between remote and infarcted myocardium (R: 89 +/- 60 Omega vs. S: 180 +/- 37 Omega). This difference increased over time in susceptible animals (252 +/- 53 Omega at 21 days) due to post-MI impedance changes on the remote myocardium. These data suggest that early electrotonic changes post-MI could be used to assess later arrhythmia susceptibility. In addition, passive-electrical changes could be a mechanism driving active-electrical remodeling post-MI, thereby facilitating the induction of arrhythmias.     

Connective tissue growth factor and cardiac fibrosis after myocardial infarction.

The temporal and spatial expression of transforming growth factor (TGF)-beta(1) and connective tissue growth factor (CTGF) was assessed in the left ventricle of a myocardial infarction (MI) model of injury with and without angiotensin-converting enzyme (ACE) inhibition. Coronary artery ligated rats were killed 1, 3, 7, 28, and 180 days after MI. TGF-beta(1), CTGF, and procollagen alpha1(I) mRNA were localized by in situ hybridization, and TGF-beta(1) and CTGF protein levels by immunohistochemistry. Collagen protein was measured using picrosirius red staining. In a separate group, rats were treated for 6 months with an ACE inhibitor. There were temporal and regional differences in the expression of TGF-beta(1), CTGF, and collagen after MI. Procollagen alpha1(I) mRNA expression increased in the border zone and scar peaking 1 week after MI, whereas collagen protein increased in all areas of the heart over the 180 days. Expression of TGF-beta(1) mRNA and protein showed major increases in the border zone and scar peaking 1 week after MI. The major increases in CTGF mRNA and protein occurred in the viable myocardium at 180 days after MI. Long-term ACE inhibition reduced left ventricular mass and decreased fibrosis in the viable myocardium, but had no effect on cardiac TGF-beta(1) or CTGF. TGF-beta(1) is involved in the initial, acute phase of inflammation and repair after MI, whereas CTGF is involved in the ongoing fibrosis of the heart. The antifibrotic benefits of captopril are not mediated through a reduction in CTGF.     

Platelet-derived growth factor oncoprotein detection using three-dimensional carbon microarrays

The potential of aptamers as ligand binding molecule has opened new avenues in the development of biosensors for cancer oncoproteins. In this paper, a label-free detection strategy using signaling aptamer/protein binding complex for platelet-derived growth factor (PDGF-BB) oncoprotein detection is reported. The detection mechanism is based on the release of fluorophore (TOTO intercalating dye) from the target binding aptamer's stem structure when it captures PDGF. Amino-terminated three-dimensional carbon microarrays fabricated by pyrolyzing patterned photoresist were used as a detection platform. The sensor showed near linear relationship between the relative fluorescence difference and protein concentration even in the sub-nanomolar range with an excellent detection limit of 5pmol. This detection strategy is promising in a wide range of applications in the detection of cancer biomarkers and other proteins.