Tryptase-Positive Mast Cells Promote Adipose Fibrosis in Secondary Lymphedema through PDGF.

Lymphedema is a chronic and progressive condition that causes physical disfigurement and psychological trauma due to the accumulation of lymphatic fluid in the interstitial space. Once it develops, lymphedema is difficult to treat because it leads to the fibrosis of adipose tissue. However, the mechanism behind this remains unclear. The purpose of this study was to investigate the involvement of mast cells (MCs) in the adipose tissues of patients with lymphedema. We found that fibrosis spread through blood vessels in the adipose tissues of lymphedema patients, and the expression of the collagen I and III genes was significantly increased compared to that of those in normal adipose tissue. Immunostaining of vimentin and α-smooth muscle actin showed that fibroblasts were the main cellular components in severely fibrotic regions. Toluidine blue staining confirmed a significant increase in the number of MCs in the adipose tissues of lymphedema patients, and immunostaining of serial sections of adipose tissue showed a significant increase in the number of tryptase-positive cells in lymphedema tissues compared with those in normal adipose tissues. Linear regression analyses revealed significant positive correlations between tryptase and the expressions of the TNF-α, platelet-derived growth factor (PDGF)-A, and PDGFR-α genes. PDGF-A-positive staining was observed in both fibroblasts and granules of tryptase-positive MCs. These results suggest that MC-derived tryptase plays a role in the fibrosis of adipose tissue due to lymphedema directly or in cooperation with other mediators.

In vitro Pretreatment with Zinc Alleviates the Adverse Effects of Tetrahydrocannabinol on Cultured Mouse Sertoli Cells: Role of Anti-apoptotic and Antioxidant Activities.

BACKGROUND: Global rise in cannabis abuse during reproductive years has placed a large number of men at risk for the adverse consequences of δ-9-tetrahydrocannabinol (THC), the primary active component of cannabis. It has been reported that THC affects male fertility and causes testicular cell dysfunction and apoptosis. This study aimed to investigate the possible protective role of zinc pretreatment against the toxic effects of THC in cultured mouse Sertoli cells and the underlying mechanism. METHODS: The Mus Musculus Sertoli cell line (TM4) was cultured, exposed to THC alone (470 µM, 24 h), co-administered with zinc (8 µM, 48 h), and investigated in three groups: control, THC, and THC + zinc. The MTT was performed to evaluate cell viability. TUNEL assay was also applied for the detection of cell apoptosis and a western blot was performed for measuring protein expression levels of Caspase3, Pro-caspase3, SOD, and PDGF-A. RESULTS: THC significantly decreased cell viability (p < 0.001) and expression levels of SOD, PDGF-A, and pro-caspase3 proteins (p < 0.05 for all), whereas increased Sertoli cells apoptosis (p < 0.001) and expression level of cleaved caspase3 protein (p < 0.001). Pretreatment with zinc reversed THC-induced apoptotic and oxidative effects and reduced cleaved caspase3/pro-caspase3 ratio but could not reverse THC-induced reduction of PDGF-A expression level in TM4 cells. CONCLUSION: The present data suggest that THC induces Sertoli cell damage through a multitarget mechanism. Zinc was reported to protect against THC-induced Sertoli cell damage due to its antiapoptotic and antioxidant activities, indicating its clinical importance against THC-induced testicular toxicity among addicted men.

Telocytes in the human ascending aorta: Characterization and exosome-related KLF-4/VEGF-A expression.

Telocytes (TCs), a novel interstitial cell entity promoting tissue regeneration, have been described in various tissues. Their role in inter-cellular signalling and tissue remodelling has been reported in almost all human tissues. This study hypothesizes that TC also contributes to tissue remodelling and regeneration of the human thoracic aorta (HTA). The understanding of tissue homeostasis and regenerative potential of the HTA is of high clinical interest as it plays a crucial role in pathogenesis from aortic dilatation to lethal dissection. Therefore, we obtained twenty-five aortic specimens of heart donors during transplantation. The presence of TCs was detected in different layers of aortic tissue and characterized by immunofluorescence and transmission electron microscopy. Further, we cultivated and isolated TCs in highly differentiated form identified by positive staining for CD34 and c-kit. Aortic-derived TC was characterized by the expression of PDGFR-α, PDGFR-ß, CD29/integrin ß-1 and αSMA and the stem cell markers Nanog and KLF-4. Moreover, TC exosomes were isolated and characterized for soluble angiogenic factors by Western blot. CD34+ /c-kit+ TCs shed exosomes containing the soluble factors VEGF-A, KLF-4 and PDGF-A. In summary, TC occurs in the aortic wall. Correspondingly, exosomes, derived from aortic TCs, contain vasculogenesis-relevant proteins. Understanding the regulation of TC-mediated aortic remodelling may be a crucial step towards designing strategies to promote aortic repair and prevent adverse remodelling.

Platelet derived growth factor-A (Pdgf-a) gene transfer modulates scar composition and improves left ventricular function after myocardial infarction.

BACKGROUND: Novel therapies that can limit or reverse damage caused by myocardial infarction (MI) could ease the increasing burden of heart failure. In this regard Platelet Derived Growth Factor (PDGF) has been previously shown to contribute to cardiac repair after MI. Here, we use a rodent model of MI and recombinant adeno-associated virus 9 (rAAV9)-mediated gene transfer to overexpress Pdgf-a in the injured heart and assess its therapeutic potential. METHODS AND RESULTS: Sprague Dawley rats underwent temporary occlusion of the left anterior descending coronary artery, followed immediately by systemic delivery of 1 × 10^11 vector genomes of either rAAV9 Pdgf-a or rAAV9 Empty vector (control). At day 28 post-MI echocardiography showed significantly improved left ventricular (LV) function (fractional shortening) after rAAV9 Pdgf-a (0.394 ± 0.019%) treatment vs control (0.304 ± 0.018%). Immunohistochemical analysis demonstrated significantly increased capillary and arteriolar density in the infarct border zone of rAAV9 Pdgf-a treated hearts together with a significant reduction in infarct scar size (rAAV9 Pdgf-a 6.09 ± 0.94% vs Empty 12.45 ± 0.92%). Western blot and qPCR analyses confirmed overexpression of PDGF-A and showed upregulation of smooth muscle alpha actin (Acta2), collagen type III alpha 1 (Col3a1) and lysyl oxidase (Lox) genes in rAAV9 Pdgf-a treated infarcts. CONCLUSION: Overexpression of Pdgf-a in the post-MI heart can modulate scar composition and improve LV function. Our study highlights the potential of rAAV gene transfer of Pdgf-a as a cardio-reparative therapy.

Lung developmental arrest caused by PDGF-A deletion: consequences for the adult mouse lung.

PDGF-A is a key contributor to lung development in mice. Its expression is needed for secondary septation of the alveoli and deletion of the gene leads to abnormally enlarged alveolar air spaces in mice. In humans, the same phenotype is the hallmark of bronchopulmonary dysplasia (BPD), a disease that affects premature babies and may have long lasting consequences in adulthood. So far, the knowledge regarding adult effects of developmental arrest in the lung is limited. This is attributable to few follow-up studies of BPD survivors and lack of good experimental models that could help predict the outcomes of this early age disease for the adult individual. In this study, we used the constitutive lung-specific Pdgfa deletion mouse model to analyze the consequences of developmental lung defects in adult mice. We assessed lung morphology, physiology, cellular content, ECM composition and proteomics data in mature mice, that perinatally exhibited lungs with a BPD-like morphology. Histological and physiological analyses both revealed that enlarged alveolar air spaces remained until adulthood, resulting in higher lung compliance and higher respiratory volume in knockout mice. Still, no or only small differences were seen in cellular, ECM and protein content when comparing knockout and control mice. Taken together, our results indicate that Pdgfa deletion-induced lung developmental arrest has consequences for the adult lung at the morphological and functional level. In addition, these mice can reach adulthood with a BPD-like phenotype, which makes them a robust model to further investigate the pathophysiological progression of the disease and test putative regenerative therapies.

Sphingosine-1-phosphate promotes PDGF-dependent endothelial progenitor cell angiogenesis in human chondrosarcoma cells.

The malignant bone tumors that are categorized as chondrosarcomas display a high potential for metastasis in late-stage disease. Higher-grade chondrosarcomas contain higher levels of expression of platelet-derived growth factor (PDGF) and its receptor. The phosphorylation of sphingosine by sphingosine kinase enzymes SphK1 and SphK2 generates sphingosine-1-phosphate (S1P), which inhibits human chondrosarcoma cell migration, while SphK1 overexpression suppresses lung metastasis of chondrosarcoma. We sought to determine whether S1P mediates levels of PDGF-A expression and angiogenesis in chondrosarcoma. Surprisingly, our investigations found that treatment of chondrosarcoma cells with S1P and transfecting them with SphK1 cDNA increased PDGF-A expression and induced angiogenesis of endothelial progenitor cells (EPCs). Ras, Raf, MEK, ERK and AP-1 inhibitors and their small interfering RNAs (siRNAs) inhibited S1P-induced PDGF-A expression and EPC angiogenesis. Our results indicate that S1P promotes the expression of PDGF-A in chondrosarcoma via the Ras/Raf/MEK/ERK/AP-1 signaling cascade and stimulates EPC angiogenesis.

Role of PDGF-A-Activated ERK Signaling Mediated FAK-Paxillin Interaction in Oligodendrocyte Progenitor Cell Migration.

Oligodendrocyte progenitor cells (OPCs) originate from the sub-ventricular zone of the developing brain. They migrate and proliferate to occupy the white matter tracts of the central nervous system and transform into myelinating oligodendrocytes. Along their route of migration, OPCs are guided and controlled by several growth factors and chemokines. PDGF-A (platelet-derived growth factor), a growth factor, serves as a monogenic and mitogenic cue during the process and activates intracellular signaling pathways inside the cell. Activation of extracellular signal regulated kinase (ERK) signaling is one of the mechanisms by which PDGF-A induces the migration of OPCs. However, the mechanisms governing the PDGF-A-induced ERK-driven OPCs migration are still unclear. In the current study, we investigated further the role of PDGF-A-induced ERK signaling in OPC migration. First, we confirmed the role of PDGF-A-activated ERK signaling in OPC migration using the pharmacological inhibitor U0126, or siRNA-mediated suppression of ERK expression. Then, we demonstrated that PDGF-A-induced actin reorganization and interaction of focal adhesion kinase (FAK), Paxillin, and pERK signals are impaired in OPCs treated with the MEK inhibitor U0126. Thus, our findings demonstrated that PDGF-A induces OPC migration in an ERK-dependent mechanism via regulation of actin reorganization and FAK-Paxillin interaction.

PDGF-A suppresses contact inhibition during directional collective cell migration.

The leading-edge mesendoderm (LEM) of the Xenopus gastrula moves as an aggregate by collective migration. However, LEM cells on fibronectin in vitro show contact inhibition of locomotion by quickly retracting lamellipodia upon mutual contact. We found that a fibronectin-integrin-syndecan module acts between p21-activated kinase 1 upstream and ephrin B1 downstream to promote the contact-induced collapse of lamellipodia. To function in this module, fibronectin has to be present as puncta on the surface of LEM cells. To overcome contact inhibition in LEM cell aggregates, PDGF-A deposited in the endogenous substratum of LEM migration blocks the fibronectin-integrin-syndecan module at the integrin level. This stabilizes lamellipodia preferentially in the direction of normal LEM movement and supports cell orientation and the directional migration of the coherent LEM cell mass.

CDK4/6 and PDGFRA Signaling as Therapeutic Targets in Diffuse Intrinsic Pontine Glioma.

Diffuse intrinsic pontine gliomas (DIPGs) are incurable childhood brain tumors, whereby the standard of care is focal radiation, a treatment that provides temporary relief for most patients. Surprisingly, decades of clinical trials have failed to identify additional therapies that can prolong survival in this disease. In this conference manuscript, we discuss how genetic engineered mouse modeling techniques with the use of a retroviral gene delivery system can help dissect the complex pathophysiology of this disease. With this approach, autochthonous murine DIPG models can be readily induced to (1) help interrogate the function of novel genetic alterations in tumorigenesis, (2) identify candidate cells of origin for this disease, (3) address how region-specific differences in the central nervous system influence the process of gliomagenesis, and (4) evaluate novel therapeutics in an immunocompetent model.

PDGF-A signaling is required for secondary alveolar septation and controls epithelial proliferation in the developing lung.

Platelet-derived growth factor A (PDGF-A) signaling through PDGF receptor α is essential for alveogenesis. Previous studies have shown that Pdgfa-/- mouse lungs have enlarged alveolar airspace with absence of secondary septation, both distinctive features of bronchopulmonary dysplasia. To study how PDGF-A signaling is involved in alveogenesis, we generated lung-specific Pdgfa knockout mice (Pdgfafl/-; Spc-cre) and characterized their phenotype postnatally. Histological differences between mutant mice and littermate controls were visible after the onset of alveogenesis and maintained until adulthood. Additionally, we generated Pdgfafl/-; Spc-cre; PdgfraGFP/+ mice in which Pdgfra+ cells exhibit nuclear GFP expression. In the absence of PDGF-A, the number of PdgfraGFP+ cells was significantly decreased. In addition, proliferation of PdgfraGFP+ cells was reduced. During alveogenesis, PdgfraGFP+ myofibroblasts failed to form the α-smooth muscle actin rings necessary for alveolar secondary septation. These results indicate that PDGF-A signaling is involved in myofibroblast proliferation and migration. In addition, we show an increase in both the number and proliferation of alveolar type II cells in Pdgfafl/-; Spc-cre lungs, suggesting that the increased alveolar airspace is not caused solely by deficient myofibroblast function.

FoxM1 promotes breast tumorigenesis by activating PDGF-A and forming a positive feedback loop with the PDGF/AKT signaling pathway.

The autocrine platelet-derived growth factor (PDGF)/PDGF receptor (PDGFR) signaling pathway promotes breast cancer tumorigenesis, but the mechanisms for its dysregulation in breast cancer are largely unknown. In the study, we identified PDGF-A as a novel transcriptional target of FoxM1. FoxM1 directly binds to two sites in the promoter of PDGF-A and activates its transcription. Mutation of these FoxM1-binding sites diminished PDGF-A promoter activity. Increased FoxM1 resulted in the upregulation of PDGF-A, which led to activation of the AKT pathway and increased breast cancer cell proliferation and tumorigenesis, whereas knockdown of FoxM1 does the opposite. Blocking AKT activation with a phosphoinositide 3-kinase/AKT inhibitor decreased FoxM1-induced cell proliferation. Moreover, PDGF/AKT pathway upregulates the expression of FoxM1 in breast cancer cells. Knockdown of PDGF-A or blockade of AKT activation inhibited the expression of FoxM1 in breast cancer cells. Furthermore, expression of FoxM1 significantly correlated with the expression of PDGF-A and the activated AKT signaling pathway in human breast cancer specimens. Our study demonstrates a novel positive regulatory feedback loop between FoxM1 and the PDGF/AKT signaling pathway; this loop contributes to breast cancer cell growth and tumorigenesis.

Effect of IGF-I on Hair Growth Is Related to the Anti-Apoptotic Effect of IGF-I and Up-Regulation of PDGF-A and PDGF-B.

BACKGROUND: Insulin-like growth factor-I (IGF-I) shares a high degree of structural and functional homology with insulin and is a potent mitogen supporting cell growth and survival in many kinds of the tissues and cells. It also plays a role in some differentiation and anti-apoptotic functions. In previous reports, it has been shown that IGF-I stimulates hair follicle (HF) growth, maintains the anagen stage, and postpones the catagen stage. OBJECTIVE: The exact mechanism of the effect of IGF-I on HF growth is not yet established. Therefore, we investigated the relationships between IGF-I and various other factors (i.e. apoptosis related molecules, pro-inflammatory cytokines, other growth factors, etc.) in the control of HF growth. METHODS: The effect of IGF-I on human hair growth was measured using an organ culture model of human HFs and compared with a control group that did not receive IGF-I. We also measured mRNA expression of factors related to hair growth and apoptosis (which was determined by reverse transcription polymerase chain reaction (RT-PCR). RT-PCR was done on days 2, 4, 6, and 8 of organ culture. RESULTS: In organ cultured human hair follicles, IGF-I had a positive effect on the rate of linear hair growth. IGF-I maintained the anagen phase. IGF-I increased the expression of platelet-derived growth factor (PDGF)-A, PDGF-B and the expression ratio of Bcl-2/Bax. CONCLUSION: The effect of IGF-I on hair growth appears to be related to the upregulation of PDGF-A and PDGF-B and to the anti-apoptotic effect of IGF-I.

Effect of IGF-I on Hair Growth Is Related to the Anti-Apoptotic Effect of IGF-I and Up-Regulation of PDGF-A and PDGF-B

BACKGROUND: Insulin-like growth factor-I (IGF-I) shares a high degree of structural and functional homology with insulin and is a potent mitogen supporting cell growth and survival in many kinds of the tissues and cells. It also plays a role in some differentiation and anti-apoptotic functions. In previous reports, it has been shown that IGF-I stimulates hair follicle (HF) growth, maintains the anagen stage, and postpones the catagen stage. OBJECTIVE: The exact mechanism of the effect of IGF-I on HF growth is not yet established. Therefore, we investigated the relationships between IGF-I and various other factors (i.e. apoptosis related molecules, pro-inflammatory cytokines, other growth factors, etc.) in the control of HF growth. METHODS: The effect of IGF-I on human hair growth was measured using an organ culture model of human HFs and compared with a control group that did not receive IGF-I. We also measured mRNA expression of factors related to hair growth and apoptosis (which was determined by reverse transcription polymerase chain reaction (RT-PCR). RT-PCR was done on days 2, 4, 6, and 8 of organ culture. RESULTS: In organ cultured human hair follicles, IGF-I had a positive effect on the rate of linear hair growth. IGF-I maintained the anagen phase. IGF-I increased the expression of platelet-derived growth factor (PDGF)-A, PDGF-B and the expression ratio of Bcl-2/Bax. CONCLUSION: The effect of IGF-I on hair growth appears to be related to the upregulation of PDGF-A and PDGF-B and to the anti-apoptotic effect of IGF-I.

FGF-2, TGFâ-1, PDGF-A and respective receptors expression in pleomorphic adenoma myoepithelial cells: an in vivo and in vitro study

Myoepithelial cells have an important role in salivary gland tumor development, contributing to a low grade of aggressiveness of these tumors. Normal myoepithelial cells are known by their suppressor function presenting increased expression of extracellular matrix genes and protease inhibitors. The importance of stromal cells and growth factors during tumor initiation and progression has been highlighted by recent literature. Many tumors result from the alteration of paracrine growth factors pathways. Growth factors mediate a wide variety of biological processes such as development, tissue repair and tumorigenesis, and also contribute to cellular proliferation and transformation in neoplastic cells. OBJECTIVES: This study evaluated the expression of fibroblast growth factor-2 (FGF-2), transforming growth factor â-1 (TGFâ-1), platelet-derived growth factor-A (PDGF-A) and their respective receptors (FGFR-1, FGFR-2, TGFâR-II and PDGFR-á) in myoepithelial cells from pleomorphic adenomas (PA) by in vivo and in vitro experiments. MATERIAL AND METHODS: Serial sections were obtained from paraffin-embedded PA samples obtained from the school's files. Myoepithelial cells were obtained from explants of PA tumors provided by surgery from different donors. Immunohistochemistry, cell culture and immunofluorescence assays were used to evaluate growth factor expression. RESULTS: The present findings demonstrated that myoepithelial cells from PA were mainly positive to FGF-2 and FGFR-1 by immunohistochemistry and immunofluorescence. PDGF-A and PDGFR-á had moderate expression by immunohistochemistry and presented punctated deposits throughout cytoplasm of myoepithelial cells. FGFR-2, TGFâ-1 and TGFâR-II were negative in all samples. CONCLUSIONS: These data suggested that FGF-2 compared to the other studied growth factors has an important role in PA benign myoepithelial cells, probably contributing to proliferation of ...

Construction and identification of recombinant adenovirus expressing hPDGF-A and hBD-2 / 第三军医大学学报

Objective To further determine their possible synergistic effect on accelerating wound healing, adenovirus vector containing recombinant human hPDGF-A and hBD2 genes was constructed and the expression of exogenous genes in transformed mesenchymal stem cells derived from rat bone marrow was observed. Methods By putting IRES in the middle of hPDGF-A and hBD2, these two genes were expected to be expressed individually. The shuttle vector was named as pAdTrack-hPDGF-A-IRES2-hBD2, which homologously recombinated with Adeasy-1 in BJ5183 cells and formed the mammalian expression vector pAdeasy-hPDGF-A-IRES2-hBD2. Furthermore, the recombinant vector was packaged in 293 cells into infectious recombinant adenovirus, which were used to infect BMSCs. The expression of hPDGF-A and hBD2 in BMSCs was detected by RT-PCR. Results We successfully constructed recombinant adenovirus vector that simultaneously expressed hPDGF-A and hBD2. The expressions of hPDGF-A and hBD2 were confirmed by RT-PCR on transformed BMSCs. Conclusion The established BMSCs that overexpressed hPDGF-A and hBD2 provide a new strategy of combining cell therapy and gene therapy to promote wound healing, especially the chronic one.

Effect of Unilateral Ureteral Obstruction(UUO) on the Expression of Platelet-Derived Growth Factor(PDGF) in the Sprague-Dawley Rat / 대한신장학회잡지

Unilateral ureteral obstruction(UUO) results in severe renal vascular constriction through the activation of renin-angiotensin system, which causes progressive tubulointerstitial fibrosis. Platelet-derived growth factor(PDGF) plays an important role in stimulating myofibroblasts and regulating synthesis of extracellular matrix in renal interstitial proliferation and fibrosis. This study was designed to investigate the relationship between unilateral ureteral obstruction and PDGF expression in tubulointerstitial fibrosis of the kidney. Eleven adult male Spraugue-Dawley rats were carried out unilateral ureteral ligation and sham operation. After 14 days, control kidney, UUO kidney and intact opposite(IO) kidney were harvested. Tissue fibrosis was quantified morphologically using the point detection method after Masson-Trichrome stain. Expression of PDGF-A and B was determined by immunohistochemical staining, RT-PCR and Western blot assay. Results were as follows: 1) UUO and IO group resulted in reduced kidney weight compared with control group(p<0.05). 2) Collagen deposition was increased in the renal cortex of UUO group(p<0.05). 3) PDGF-A and B mRNA expression was increased significantly compared with control and IO group(p< 0.05). 4) PDGF-A and B protein expression were increased in UUO and IO group(p<0.05). 5) On the immunohistochemical staining for PDGF- A and B, staining intensity was increased significantly at the renal cortex, interstitium and tubular epithelial cells in the UUO group. This results indicated that PDGF-A and B plays important role in tubulointerstitial fibrosis developed after unilateral ureteral obstruction and compensatory fibroproliferative growth in contralateral kidney.