Platelet-derived Growth factor-B Overexpression Research Articles

PROX1 Inhibits PDGF-B Expression to Prevent Myxomatous Degeneration of Heart Valves.

Cardiac valve disease is observed in 2.5% of the general population and 10% of the elderly people. Effective pharmacological treatments are currently not available, and patients with severe cardiac valve disease require surgery. PROX1 (prospero-related homeobox transcription factor 1) and FOXC2 (Forkhead box C2 transcription factor) are transcription factors that are required for the development of lymphatic and venous valves. We found that PROX1 and FOXC2 are expressed in a subset of valvular endothelial cells (VECs) that are located on the downstream (fibrosa) side of cardiac valves. Whether PROX1 and FOXC2 regulate cardiac valve development and disease is not known. We used histology, electron microscopy, and echocardiography to investigate the structure and functioning of heart valves from Prox1ΔVEC mice in which Prox1 was conditionally deleted from VECs. Isolated valve endothelial cells and valve interstitial cells were used to identify the molecular mechanisms in vitro, which were tested in vivo by RNAScope, additional mouse models, and pharmacological approaches. The significance of our findings was tested by evaluation of human samples of mitral valve prolapse and aortic valve insufficiency. Histological analysis revealed that the aortic and mitral valves of Prox1ΔVEC mice become progressively thick and myxomatous. Echocardiography revealed that the aortic valves of Prox1ΔVEC mice are stenotic. FOXC2 was downregulated and PDGF-B (platelet-derived growth factor-B) was upregulated in the VECs of Prox1ΔVEC mice. Conditional knockdown of FOXC2 and conditional overexpression of PDGF-B in VECs recapitulated the phenotype of Prox1ΔVEC mice. PDGF-B was also increased in mice lacking FOXC2 and in human mitral valve prolapse and insufficient aortic valve samples. Pharmacological inhibition of PDGF-B signaling with imatinib partially ameliorated the valve defects of Prox1ΔVEC mice. PROX1 antagonizes PDGF-B signaling partially via FOXC2 to maintain the extracellular matrix composition and prevent myxomatous degeneration of cardiac valves.

Abstract P2-21-05: Vascular effects of Platelet-derived growth factor-B (PDGFB) in breast cancer metastasis to the brain

Abstract Brain metastases are associated with shortened survival and poorer quality of life in breast cancer patients. Recently, the importance of platelet-derived growth factor-B (PDGFB) and its stromally localized cognate receptor, platelet-derived growth factor receptor-beta (PDGFRβ) in promoting breast cancer metastasis to the brain was identified by our group. Historically, PDGFB-to-PDGFRβ signaling is known to promote angiogenesis and vessel maturation both in normal physiology and in cancer, but the mechanism(s) behind this tumor stromal interplay in brain metastasis is poorly understood. Based on our previous work and the work of others, we hypothesized that breast tumor-derived PDGFB directly modulates both brain endothelial and blood brain barrier (BBB) function by altering angiogenesis and endothelial permeability. To test this directly, we over-expressed PDGFB ligand in MDA-MB-231 and HCC1187 triple negative breast cancer cell lines and compared the angiogenic potential of the conditioned medium (CM) from these cells. We utilized both a 3D in vitro (human brain microvascular endothelial cell, HBMEC) spheroid angiogenesis model and a tri-culture BBB system (HBMECs in direct interaction with human astrocytes and human pericytes) to test the BBB vasculature. CM harvested from PDGFB overexpressing cells significantly increased the sprouting angiogenesis potential of 3D HBMECs and the migration of 3D tri-culture BBB spheroids compared to CM obtained from parental cells. In addition, CM from PDGFB overexpressing cells caused the surface of BBB spheroids to exhibit significantly increased permeability to high molecular weight dextran compared to CM from parental cells. In agreement with these findings, the tight junction protein ZO1 was diminished. Importantly, in vivo BBB permeability was also tested upon intracardiac injection of MDA-MB-231 cells with and without PDGFB overexpression. These studies revealed that Evans blue dye fluorescence intensity was significantly higher in the brains of mice injected with PDGFB overexpressing cells compared to parental cells. Overall, these studies reveal a previously unrecognized role for PDGFB-to-PDGFRβ signaling in brain vascular remodeling and BBB permeability, which has clinical implications for women with PDGFB-positive breast cancer. Citation Format: Sajita Shah, Rebecca Packard, Sarah Steck, Alexis Mossing, Yalini Ramamoorthy, Katie Thies, Alexander Didier, Jonathan Spehar, Dillon Richardson, Tasneem Arsiwala, Jonathan Godbout, Steven Sizemore, Paul Lockman, Gina Sizemore. Vascular effects of Platelet-derived growth factor-B (PDGFB) in breast cancer metastasis to the brain [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P2-21-05.

Platelet-derived growth factor-B signalling might promote epithelial-mesenchymal transition in gastric carcinoma cells through activation of the MAPK/ERK pathway.

IntroductionEpithelial-mesenchymal transition (EMT) is important in the metastasis of tumours and is triggered by several key growth factors, including platelet-derived growth factor-B (PDGF-B). But, whether PDGF-B signalling promotes EMT in gastric carcinoma cells is still unknown.Material and methodsWe established 2 gastric carcinoma cell lines (MKN28 and MKN45) to stably overexpress PDGF-B by lentiviral vectors, and expression of E-cadherin, N-cadherin, and ERK-1 were detected by western blot assay. Then, PDGF-B overexpression and normal MKN28 and MKN45 cells were cocultured with PDGFR-b positive fibroblast (hs738) and MAPK inhibitors were added; also, the expressions of ERK-1, E-cadherin, and N-cadherin were detected by western blot assay.ResultsAfter being cocultured with hs738 cells, expressions of ERK-1 and N-cadherin protein in PDGF-B overexpression MKN28 and MKN45 cells were much higher than normal MKN28 and MKN45 cells (p < 0.05), and those could be decreased by MAPK inhibitor. Also, expressions of E-cadherin protein in PDGF-B overexpression MKN28 and MKN45 cells were much lower than normal MKN28 and MKN45 cells (p < 0.05), and they could be increased by MAPK inhibitor.ConclusionsOur data indicate that PDGF-B signalling can induce EMT in gastric carcinoma cells. Thr tumour microenvironment is imperative in the process of PDGF-B signalling inducing EMT in gastric carcinoma cells. Also, activation of MAPK/ERK pathway, which is a downstream pathway of PDGF-B signalling, might participate in this process.

Aggravated pulmonary injury after subarachnoid hemorrhage in PDGF-Bret/ret mice

BackgroundRecent advances in surgical and neuroprotective strategies could effectively manage the pathophysiological progression of subarachnoid hemorrhage (SAH). However, pulmonary dysfunction frequently occurs in SAH patients with an increased risk of unsatisfactory outcomes. Based on the similar microvascular structures in the blood-air barrier and blood-brain barrier and possible brain-lung crosstalks, we believe that pericytes may be involved in both neurological and pulmonary dysfunction after SAH.MethodsIn our experiments, platelet-derived growth factor B (PDGF-B) retention motif knockout (PDGF-Bret/ret) mice and adeno-associated virus PDGF-B were employed to show the involvement of pericyte deficiency and PDGF-B expression. Neurological score, SAH grade, hematoxylin-eosin staining, and PaO2/FiO2 ratio analysis were performed to evaluate the neurological deficits and pulmonary functions in endovascular perforation SAH models at 24 h after surgery, as well as western blotting and immunofluorescence staining for underlying molecular expressions.ResultsWe found that neonatal PDGF-Bret/ret mice exhibited pulmonary atelectasis 12 h after birth. Further investigation showed a decrease in PaO2/FiO2 and lung-specific surfactant proteins in adult PDGF-Bret/ret mice. These dysfunctions were much worse than those in wild-type mice at 24 h after SAH. PDGF-B overexpression alleviated pulmonary dysfunction after SAH.ConclusionsThese results suggested pulmonary dysfunction after SAH and the pivotal role of PDGF-B signaling for the pathophysiological process and future therapeutic targets of pulmonary injury treatment after SAH. Further studies are needed for pathophysiological investigations and translational studies on pulmonary injuries after SAH.

NCK-dependent pericyte migration promotes pathological neovascularization in ischemic retinopathy

Pericytes are mural cells that surround capillaries and control angiogenesis and capillary barrier function. During sprouting angiogenesis, endothelial cell-derived platelet-derived growth factor-B (PDGF-B) regulates pericyte proliferation and migration via the platelet-derived growth factor receptor-β (PDGFRβ). PDGF-B overexpression has been associated with proliferative retinopathy, but the underlying mechanisms remain poorly understood. Here we show that abnormal, α-SMA-expressing pericytes cover angiogenic sprouts and pathological neovascular tufts (NVTs) in a mouse model of oxygen-induced retinopathy. Genetic lineage tracing demonstrates that pericytes acquire α-SMA expression during NVT formation. Pericyte depletion through inducible endothelial-specific knockout of Pdgf-b decreases NVT formation and impairs revascularization. Inactivation of the NCK1 and NCK2 adaptor proteins inhibits pericyte migration by preventing PDGF-B-induced phosphorylation of PDGFRβ at Y1009 and PAK activation. Loss of Nck1 and Nck2 in mural cells prevents NVT formation and vascular leakage and promotes revascularization, suggesting PDGFRβ-Y1009/NCK signaling as a potential target for the treatment of retinopathies.

Thalidomide Reduces Hemorrhage of Brain Arteriovenous Malformations in a Mouse Model.

Brain arteriovenous malformation (bAVM) is an important risk factor for intracranial hemorrhage. Current treatments for bAVM are all associated with considerable risks. There is no safe method to prevent bAVM hemorrhage. Thalidomide reduces nose bleeding in patients with hereditary hemorrhagic telangiectasia, an inherited disorder characterized by vascular malformations. In this study, we tested whether thalidomide and its less toxic analog, lenalidomide, reduce bAVM hemorrhage using a mouse model. bAVMs were induced through induction of brain focal activin-like kinase 1 (Alk1, an AVM causative gene) gene deletion and angiogenesis in adult Alk1-floxed mice. Thalidomide was injected intraperitoneally twice per week for 6 weeks, starting either 2 or 8 weeks after AVM induction. Lenalidomide was injected intraperitoneally daily starting 8 weeks after AVM induction for 6 weeks. Brain samples were collected at the end of the treatments for morphology, mRNA, and protein analyses. The influence of Alk1 downregulation on PDGFB (platelet-derived growth factor B) expression was also studied on cultured human brain microvascular endothelial cells. The effect of PDGFB in mural cell recruitment in bAVM was explored by injection of a PDGFB overexpressing lentiviral vector to the mouse brain. Thalidomide or lenalidomide treatment reduced the number of dysplastic vessels and hemorrhage and increased mural cell (vascular smooth muscle cells and pericytes) coverage in the bAVM lesion. Thalidomide reduced the burden of CD68+ cells and the expression of inflammatory cytokines in the bAVM lesions. PDGFB expression was reduced in ALK1-knockdown human brain microvascular endothelial cells and in mouse bAVM lesion. Thalidomide increased Pdgfb expression in bAVM lesion. Overexpression of PDGFB mimicked the effect of thalidomide. Thalidomide and lenalidomide improve mural cell coverage of bAVM vessels and reduce bAVM hemorrhage, which is likely through upregulation of Pdgfb expression.

Smooth muscle cell recruitment to lymphatic vessels requires PDGFB and impacts vessel size but not identity.

Tissue fluid drains through blind-ended lymphatic capillaries, via smooth muscle cell (SMC)-covered collecting vessels into venous circulation. Both defective SMC recruitment to collecting vessels and ectopic recruitment to lymphatic capillaries are thought to contribute to vessel failure, leading to lymphedema. However, mechanisms controlling lymphatic SMC recruitment and its role in vessel maturation are unknown. Here, we demonstrate that platelet-derived growth factor B (PDGFB) regulates lymphatic SMC recruitment in multiple vascular beds. PDGFB is selectively expressed by lymphatic endothelial cells (LECs) of collecting vessels. LEC-specific deletion of Pdgfb prevented SMC recruitment causing dilation and failure of pulsatile contraction of collecting vessels. However, vessel remodelling and identity were unaffected. Unexpectedly, Pdgfb overexpression in LECs did not induce SMC recruitment to capillaries. This was explained by the demonstrated requirement of PDGFB extracellular matrix (ECM) retention for lymphatic SMC recruitment, and the low presence of PDGFB-binding ECM components around lymphatic capillaries. These results demonstrate the requirement of LEC-autonomous PDGFB expression and retention for SMC recruitment to lymphatic vessels, and suggest an ECM-controlled checkpoint that prevents SMC investment of capillaries, which is a common feature in lymphedematous skin.

The methylation status of the platelet-derived growth factor-B gene promoter and its regulation of cellular proliferation following folate treatment in human glioma cells

Platelet-derived growth factor-B (PDGF-B) is a growth factor that regulates cell migration, proliferation, and differentiation, and is involved in several physical and pathological processes. The overexpression of PDGF-B in glioma surgical samples revealed its effect on tumorigenesis. In this study, we determined that the expression of PDGF-B in 54 glioma samples varied among different grades and was correlated with the cell proliferation marker, Ki-67. Using pyrosequencing, we quantitatively assessed PDGF-B gene methylation levels and determined that hypomethylation promotes increased expression of PDGF-B in higher grade gliomas. Furthermore, we treated two glioma cell lines with a demethylating agent (5-aza-2׳-deoxycitidine, 5-aza-dC) or a remethylating agent (folate) to alter the methylation status of PDGF-B. The epigenetic regulation of the PDGF-B gene not only modulated the expression levels of PDGF-B but also affected the cellular proliferation induced by TGFβ-Smad activity and the PDGF-B peptide itself. Our work showed the importance of the methylation status of the PDGF-B gene promoter, and suggests that the epigenetic regulation of the PDGF-B gene may serve as a potential therapeutic target for the inhibition of glioma proliferation.

Overexpression of platelet-derived growth factor-B increases the growth, invasion, and angiogenesis of gastric carcinoma cells through protein kinase B

Platelet-derived growth factor-B (PDGF-B) promotes tumor metastasis by inducing tumor cell growth, invasion, and angiogenesis in several cancers. However, the roles of PDGF-B in gastric carcinoma are largely unknown. We established two gastric carcinoma cell lines, SGC7901 and BGC823, to stably overexpress PDGF-B by lentiviral vectors, and determined their growth, invasion and angiogenesis. Overexpression of PDGF-B significantly enhanced the cell proliferation, invasion and angiogenesis of both SGC7901 and BGC823 cells, accompanied with increased activation of AKT, which is a downstream target of PDGF signaling pathway. Consequently, an AKT kinase inhibitor abolished the PDGF-B overexpression-mediated up-regulation of growth, invasion and angiogenesis. These results indicate that PDGF-B signaling may promote the metastasis of gastric carcinoma through AKT signaling. Targeting the PDGF-B pathway may be an alternative strategy for the development of therapies for gastric cancers.

Clinicopathological significance of platelet-derived growth factor B, platelet-derived growth factor recpertor-β, and E-cadherin expression in gastric carcinoma

Aim of the studyPlatelet-derived growth factor B (PDGF-B), a vital growth factor which can induce angiogenesis and epithelial-mesenchymal transition (EMT), is important in the metastasis of many tumors. However, the roles of PDGF-B in gastric carcinoma are largely unknown. We investigated the correlation between PDGF-B, PDGFR-β and E-cadherin expression with the clinical features of gastric carcinoma patients to evaluate the relationship between PDGF-B signaling, E-cadherin and metastasis of gastric carcinoma, the correlation between PDGF-B and E-cadherin expression to assess the roles of PDGF-B signaling in metastasis of gastric carcinoma..Material and methodsWe detected expressions of PDGF-B, PDGFR-β and E-cadherin in gastric carcinoma tissues and normal gastric mucosa tissues of 64 patients with gastric carcinoma who had undergone surgical resection, and investigated their relationships with clinical features and the relationships between PDGF-B and E-cadherin expression in gastric carcinoma.ResultsIn surgical specimens, tumor cells expressed PDGF-B, and PDGFR-β was expressed by tumor stromal cells. E-cadherin was expressed by both tumor cells and normal gastric mucosa cells. Expressions of PDGF-B and PDGFR-β were increased in gastric carcinoma tissues (p < 0.05) and were positively correlated with the depth of cancer invasion, lymph node metastasis and TNM stage (p < 0.05). The expression of E-cadherin was reduced in gastric carcinoma tissues (p < 0.05) and was negatively correlated with the depth of cancer invasion, lymph node metastasis and TNM stage (p < 0.05). The correlation between PDGF-B and E-cadherin expression was negative (p < 0.05).ConclusionOur data indicate that either the overexpression of PDGF-B and PDGFR-β or the underexpression of E-cadherin is correlated with cancer progression and lymphogenous metastasis of gastric carcinoma. The PDGF-B signal pathway might induce EMT by down-regulating expression of E-cadherin to promote metastasis of gastric carcinoma.

PDGF-B-mediated downregulation of miR-21: new insights into PDGF signaling in glioblastoma

Glioblastoma (GBM) is a highly heterogeneous type of tumor characterized by genomic and signaling abnormalities affecting pathways involved in control of cell fate, including tumor-suppressor- and growth factor-regulated pathways. An aberrant miRNA expression has been observed in GBM, being associated with impaired cellular functions resulting in malignant transformation, proliferation and invasion. Here, we demonstrate for the first time that platelet-derived growth factor-B (PDGF-B), a potent angiogenic growth factor involved in GBM development and progression, promotes downregulation of pro-oncogenic (miR-21) and anti-oncogenic (miR-128) miRNAs, as well as upregulation/downregulation of several miRNAs involved in GBM pathology. Retrovirally mediated overexpression of PDGF-B in U87 human GBM cells or their prolonged exposure, as well as that of F98 rat glioma cells to this ligand, resulted in decreased miR-21 and miR-128 levels, which was associated with increased cell proliferation. Furthermore, siRNA-mediated PDGF-B silencing led to increased levels of miR-21 and miR-128, while miRNA modulation through overexpression of miR-21 did not alter the levels of PDGF-B. Finally, we demonstrate that modulation of tumor suppressors PTEN and p53 in U87 cells does not affect the decrease in miR-21 levels associated with PDGF-B overexpression. Overall, our findings suggest that, besides its role in inducing GBM tumorigenesis, PDGF-B may enhance tumor proliferation by modulating the expression of oncomiRs and tumor suppressor miRNAs in U87 human GBM cells.

Placental Gene Transfer

Katz, A.B.; Keswani, S.G.; Habli, M.; Lim, F.Y.; Zoltick, P.W.; Midrio, P.; Kozin, E.D.; Herlyn, M.; Crombleholme, T.M. Author Information

Placental gene transfer: transgene screening in mice for trophic effects on the placenta

We hypothesized that gene transfer of select growth factors to the placenta may enhance placental and fetal growth. Thus, we examined the effect of 8 growth factor transgenes on murine placenta. Adenoviral-mediated site-specific intraplacental gene transfer of 8 different growth factor transgenes at embryonic day (e) 14 was performed. Transgenes included angiopoietin-1, angiopoietin-2 (Ang-2), basic fibroblast growth factor, hepatocyte growth factor, insulin-like growth factor-1 (IGF-1), placenta growth hormone, platelet-derived growth factor-B (PDGF-B), and vascular endothelial growth factor(121). Fetuses and placentas were harvested at e17 and assessed for survival, gene transfer efficiency, placenta area, and fetal and placental weights. Efficient gene transfer to the placenta was detected with minimal dissemination to the fetus. Overexpression of IGF-1, PDGF-B, and Ang-2 resulted in an increase in placenta cross-sectional area. Only Ang-2 gene transfer resulted in increased fetal weight, and only Ang-2 and basic fibroblast growth factor resulted in a change in placental weight. Site-specific placental gene transfer results in efficient gene transfer with minimal dissemination to the fetus. Adenoviral-mediated IGF-1, adenoviral-mediated PDGF-B, and adenoviral-mediated Ang-2 significantly increase placenta growth.

Association of the T1799A BRAF mutation with tumor extrathyroidal invasion, higher peripheral platelet counts, and over-expression of platelet-derived growth factor-B in papillary thyroid cancer

The relationship among BRAF mutation, platelet counts, and platelet-derived growth factor (PDGF) with respect to clinicopathological outcomes of papillary thyroid cancer (PTC) may play a role in PTC pathogenesis but remains undefined. We examined the T1799A BRAF mutation by direct genomic DNA sequencing in 108 primary PTC samples from a Chinese cohort and analyzed its relationship with clinicopathological, hematological, and other laboratory results as well as the levels of expression of PDGF in tumors. We found that the BRAF mutation was significantly associated with extrathyroidal invasion and advanced tumor stages III and IV. Specifically, extrathyroidal invasion was seen in 30/54 (56%) PTC with BRAF mutation versus 18/54 (33%) PTC without the mutation (P=0.02). Tumor stages III and IV were seen in 16/54 (30%) PTC with BRAF mutation versus 7/54 (13%) PTC without the mutation (P=0.04). The BRAF mutation was also significantly associated with a higher platelet count, with 249.28+/-53.76 x 10(9)/l in the group of patients with BRAF mutation versus 207.79+/-58.98 x 10(9)/l in the group without the mutation (P=0.001). An association of higher platelet accounts with extrathyroidal invasion was also seen, with 242.66+/-51.85 x 10(9)/l in patients with extrathyroidal invasion versus 218.49+/-59.10 x 10(9)/l in patients without extrathyroidal invasion (P=0.03). The BRAF T1799A-positive PTC tissues harbored a significantly higher level of PDGF-B than BRAF T1799A-negative PTC tissues. The data suggest that the BRAF T1799A mutation is associated with aggressive pathological outcomes of PTC in which high platelet counts and increased PDGF production may play a role.