Background: The platelet derived growth factor-D (PDGF-D) plays an important role in breast tumor aggressiveness. However, limited study has investigated the effect of silencing PDGF-D on the biological function of breast cancer. The purpose of this study is to clarify the potential value of PDGF-D as a target for breast cancer treatment.

Methods: Reverse transcription-polymerase chain reaction and western blot were used to detect PDGF-D expression in 5 different breast cancer cells. The lentiviral vector was usd to silence PDGF-D in MDA-MB-231 cells. Then, Methyl Thiazolyl Tetrazolium was used to detect cell viability, 5-Ethynyl-2'- deoxyuridine and a soft agar assay were used to detect cell proliferation and clonality. Additionally, cell apoptosis after PDGF-D knockdown was measured by Annexin V/ Prodium Iodide staining, and cell migration was detected by trans-well assay. Survival rate and tumor size were measured by nude mice transplantation.

Results: The MDA-MB-231 and SK-BR-3 cell lines showed higher PDGF-D expression than the MCF7 cell lines (P<.05). After the PDGF-D gene was silenced, the growth and colony forming abilitys ignificantly decreased (P<.05) together with the induction of apoptosis in MDA-MB-231 cells (P<.05). Moreover, MDA-MB-231 cells with PDGF-D silencing showed significantly diminished aggressive migration and invasion potential compared to other cells (P<.05). In vivo experiments also indicated that PDGF-D silencing inhibited tumor growth and improved the survival rate of tumor-bearing mice.

Conclusion: Downregulation of PDGF-D had dramatic effects on breast cancer cell proliferation, apoptosis and migration, which indicates that it plays an important role in breast cancer development and progression.

Keywords: Apoptosis; Breast carcinoma; Migration; PDGF-D; Proliferation.

The family of vascular endothelial growth factors (VEGFs) includes 5 members (VEGF-A to -D, and placenta growth factor), which regulate several critical biological processes. VEGF-A exerts a variety of biological effects through high-affinity binding to tyrosine kinase receptors (VEGFR-1, -2 and -3), co-receptors and accessory proteins. In addition to its fundamental function in angiogenesis and endothelial cell biology, VEGF/VEGFR signalling also plays a role in other cell types including epithelial cells. This review provides an overview of VEGF signalling in biliary epithelial cell biology in both normal and pathologic conditions. VEGF/VEGFR-2 signalling stimulates bile duct proliferation in an autocrine and paracrine fashion. VEGF/VEGFR-1/VEGFR-2 and angiopoietins are involved at different stages of biliary development. In certain conditions, cholangiocytes maintain the ability to secrete VEGF-A, and to express a functional VEGFR-2 receptor. For example, in polycystic liver disease, VEGF secreted by cystic cells stimulates cyst growth and vascular remodelling through a PKA/RAS/ERK/HIF1α-dependent mechanism, unveiling a new level of complexity in VEFG/VEGFR-2 regulation in epithelial cells. VEGF/VEGFR-2 signalling is also reactivated during the liver repair process. In this context, pro-angiogenic factors mediate the interactions between epithelial, mesenchymal and inflammatory cells. This process takes place during the wound healing response, however, in chronic biliary diseases, it may lead to pathological neo-angiogenesis, a condition strictly linked with fibrosis progression, the development of cirrhosis and related complications, and cholangiocarcinoma. Novel observations indicate that in cholangiocarcinoma, VEGF is a determinant of lymphangiogenesis and of the immune response to the tumour. Better insights into the role of VEGF signalling in biliary pathophysiology might help in the search for effective therapeutic strategies.

Keywords: ADPKD, adult dominant polycystic kidney disease; Anti-Angiogenic therapy; BA, biliary atresia; BDL, bile duct ligation; CCA, cholangiocarcinoma; CCl4, carbon tetrachloride; CLDs, chronic liver diseases; Cholangiocytes; Cholangiopathies; DP, ductal plate; DPM, ductal plate malformation; DRCs, ductular reactive cells; Development; HIF-1α, hypoxia-inducible factor type 1α; HSCs, hepatic stellate cells; IHBD, intrahepatic bile ducts; IL-, interleukin-; LECs, lymphatic endothelial cells; LSECs, liver sinusoidal endothelial cells; Liver repair; MMPs, matrix metalloproteinases; PBP, peribiliary plexus; PC, polycystin; PDGF, platelet-derived growth factor; PIGF, placental growth factor; PLD, polycystic liver diseases; Polycystic liver diseases; SASP, senescence-associated secretory phenotype; TGF, transforming growth factor; VEGF, vascular endothelial growth factors; VEGF-A; VEGF/VEGFR-2 signalling; VEGFR-1/2, vascular endothelial growth factor receptor 1/2; mTOR, mammalian target of rapamycin.

Canine prostate cancer (PC) presents a poor antitumor response, usually late diagnosis and prognosis. Toceranib phosphate (TP) is a nonspecific inhibitor of receptor tyrosine kinases (RTKs), including vascular endothelial growth factor receptor (VEGFR), platelet-derived growth factor receptor (PDGFR), and c-KIT. This study aimed to evaluate VEGFR2, PDGFR-β, and c-KIT protein expression in two established canine PC cell lines (PC1 and PC2) and the transcriptome profile of the cells after treatment with TP. Immunofluorescence (IF) analysis revealed VEGFR2 and PDGFR-β protein expression and the absence of c-KIT protein expression in both cell lines. After TP treatment, only the viability of PC1 cells decreased in a dose-dependent manner. Transcriptome and enrichment analyses of treated PC1 cells revealed 181 upregulated genes, which were related to decreased angiogenesis and cell proliferation. In addition, we found upregulated PDGFR-A, PDGFR-β, and PDGF-D expression in PC1 cells, and the upregulation of PDGFR-β was also observed in treated PC1 cells by qPCR. PC2 cells had fewer protein-protein interactions (PPIs), with 18 upregulated and 22 downregulated genes; the upregulated genes were involved in the regulation of parallel pathways and mechanisms related to proliferation, which could be associated with the resistance observed after treatment. The canine PC1 cell line but not the PC2 cell line showed decreased viability after treatment with TP, although both cell lines expressed PDGFR and VEGFR receptors. Further studies could explain the mechanism of resistance in PC2 cells and provide a basis for personalized treatment for dogs with PC.

Keywords: animal model; antitumor response; dog; microarray; prostate.

Cyclophilin D (CypD) is an important component in mitochondrial-dependent tubular cell death in acute kidney injury. However, it is not known whether CypD contributes to tubular cell damage in chronic interstitial fibrosis. We investigated this question in the unilateral ureter obstruction (UUO) model of renal interstitial fibrosis. Groups of CypD-/- and wild type (WT) mice were killed 7 or 12 days after UUO surgery. The significant tubular cell apoptosis seen in WT UUO was significantly reduced in CypD-/- UUO based on TUNEL and cleaved caspase 3 staining. Other markers of tubular cell damage; loss of E-cadherin and AQP1 expression, were also reduced in the CypD-/- UUO kidney. This reduced tubular damage was associated with less inflammation and a partial protection against loss of peritubular capillaries. The prominent accumulation of α-SMA+ myofibroblasts and interstitial collagen deposition seen in WT UUO was significantly reduced in CypD-/- UUO on day 12, but not day 7. Activation of several pro-fibrotic signalling pathways (p38 MAPK, JNK and Smad3) was unaltered in CypD-/- UUO, arguing that CypD acts independently to promote renal fibrosis. CypD deletion in cultured tubular cells attenuated oxidative stress-induced pro-inflammatory, pro-fibrotic and apoptotic responses; however, responses to angiotensin II and LPS were unaffected. In contrast, CypD deletion in cultured renal fibroblasts did not affect PDGF-induced proliferation or TGF-β1-induced collagen I expression, suggesting no direct role of CypD in the fibroblast response. In conclusion, we have identified a role for CypD in chronic tubular cell damage and in the development of renal interstitial fibrosis.

Portal hypertension, defined as increased pressure in the portal vein, develops as a consequence of increased intrahepatic vascular resistance due to the dysregulation of liver sinusoidal endothelial cells (LSECs) and hepatic stellate cells (HSCs), frequently arising from chronic liver diseases. Extrahepatic haemodynamic changes contribute to the aggravation of portal hypertension. The pathogenic complexity of portal hypertension and the unsuccessful translation of preclinical studies have impeded the development of effective therapeutics for patients with cirrhosis, while counteracting hepatic and extrahepatic mechanisms also pose a major obstacle to effective treatment. In this review article, we will discuss the following topics: i) cellular and molecular mechanisms of portal hypertension, focusing on dysregulation of LSECs, HSCs and hepatic microvascular thrombosis, as well as changes in the extrahepatic vasculature, since these are the major contributors to portal hypertension; ii) translational/clinical advances in our knowledge of portal hypertension; and iii) future directions.

Keywords: ACE2, angiogenesis-converting enzyme 2; ACLF, acute-on-chronic liver failure; AT1R, angiotensin II type I receptor; CCL2, chemokine (C-C motif) ligand 2; CCl4, carbon tetrachloride; CLD, chronic liver disease; CSPH, clinically significant portal hypertension; Dll4, delta like canonical Notch ligand 4; ECM, extracellular matrix; EUS, endoscopic ultrasound; FXR; FXR, farnesoid X receptor; HCC, hepatocellular carcinoma; HRS, hepatorenal syndrome; HSC; HSCs, hepatic stellate cells; HVPG, hepatic venous pressure gradient; Hsp90, heat shock protein 90; JAK2, Janus kinase 2; KO, knockout; LSEC; LSEC, liver sinusoidal endothelial cells; MLCP, myosin light-chain phosphatase; NET, neutrophil extracellular trap; NO; NO, nitric oxide; NSBB; NSBBs, non-selective beta blockers; PDE, phosphodiesterase; PDGF, platelet-derived growth factor; PIGF, placental growth factor; PKG, cGMP-dependent protein kinase; Rho-kinase; TIPS; TIPS, transjugular intrahepatic portosystemic shunt; VCAM1, vascular cell adhesion molecule 1; VEGF; VEGF, vascular endothelial growth factor; angiogenesis; eNOS, endothelial nitric oxide synthase; fibrosis; liver stiffness; statins; β-Arr2, β-arrestin 2; β1-AR, β1-adrenergic receptor; β2-AR, β2-adrenergic receptor.

OBJECTIVE

To compare the biological characteristics of colorectal cancer associated fibroblasts (CAFs) with normal fibroblasts (NFs).

METHODS

CAFs and NFs were isolated from fresh specimens of colorectal cancer and their paired normal colon tissue and cultured by tissue explant method. Light microscopy, quantitative polymerase chain reaction (qPCR), Western blot, immunofluorescence microscopy, electron microscopy and flow cytometry were used to identify isolated fibroblasts and to explore their characteristics of activation and growth.

RESULTS

Primary colorectal CAFs and NFs were isolated and cultured successfully. NFs showed spindled morphology and were arranged in interlacing or spiral bundles. CAFs were polygonal or spindle, but were fatter than NFs. They were distributed randomly and arranged irregularly, and had obvious actin expression. CAFs and NFs both expressed fibronectin, but not E-cadherin, CDDF-1) and platelet derived growth factor (PDGF)-D in CAFs were lower while that for PDGFC was higher than that in NFs. That indicated the proliferation of CAFs was inhibited and the secretion of some cytokines was different when compared with NFs.

CONCLUSIONS

CAFs show differences with NFs in morphology, characteristics of activation and secretion of some cytokines. The proliferation of CAFs is down regulated as compared with NFs.

Na⁺/H⁺ exchanger regulatory factor 1 (NHERF1; also known as ezrin-radixin-moesin-binding phosphoprotein 50) is a PSD-95, disc large, zona occludens-1 adapter that acts as a scaffold for signaling complexes and cytoskeletal-plasma membrane interactions. NHERF1 is crucial to β-2-adrenoceptor (β₂AR)-mediated activation of cystic fibrosis transmembrane conductance regulator (CFTR) in epithelial cells, and NHERF1 has been proposed to mediate the recycling of internalized β₂AR back to the cell membrane. In the current study, we assessed the role of NHERF1 in regulating cAMP-mediated signaling and immunomodulatory functions in airway smooth muscle (ASM). NHERF1 knockdown attenuated the induction of (protein kinase A) phospho-vasodilator-stimulated phosphoprotein (p-VASP) by isoproterenol (ISO), prostaglandin E₂ (PGE₂), or forskolin (FSK) as well as the induction of p-heat shock protein 20 after 4 h of stimulation with ISO and FSK. NHERF1 knockdown fully abrogated the ISO-, PGE₂-, and FSK-induced IL-6 gene expression and cytokine production without affecting cAMP-mediated phosphodiesterase 4D (PDE4D) gene expression, phospho-cAMP response element-binding protein (p-CREB), and cAMP response element (CRE)-Luc, or PDGF-induced cyclin DDE4D gene expression. The differential regulation of cAMP-induced signaling and gene expression in our study indicates a role for NHERF1 in the compartmentalization of cAMP signaling in ASM.-Pera, T., Tompkins, E., Katz, M., Wang, B., Deshpande, D. A., Weinman, E. J., Penn, R. B. Specificity of NHERF1 regulation of GPCR signaling and function in human airway smooth muscle.

BACKGROUND

Brain pericytes ensheathe the endothelium and contribute to formation and maintenance of the blood-brain-barrier. Additionally, pericytes are involved in several aspects of the CNS immune response including scarring, adhesion molecule expression, chemokine secretion, and phagocytosis. In vitro cultures are routinely used to investigate these functions of brain pericytes, however, these are highly plastic cells and can display differing phenotypes and functional responses depending on their culture conditions. Here we sought to investigate how two commonly used culture media, high serum containing DMEM/F12 and low serum containing Pericyte Medium (ScienCell), altered the phenotype of human brain pericytes and neuroinflammatory responses.

METHODS

Pericytes were isolated from adult human brain biopsy tissue and cultured in DMEM/F12 (D-pericytes) or Pericyte Medium (P-pericytes). Immunocytochemistry, qRT-PCR, and EdU incorporation were used to determine how this altered their basal phenotype, including the expression of pericyte markers, proliferation, and cell morphology. To determine whether culture media altered the inflammatory response in human brain pericytes, immunocytochemistry, qRT-PCR, cytometric bead arrays, and flow cytometry were used to investigate transcription factor induction, chemokine secretion, adhesion molecule expression, migration, phagocytosis, and response to inflammatory-related growth factors.

RESULTS

P-pericytes displayed elevated proliferation and a distinct bipolar morphology compared to D-pericytes. Additionally, P-pericytes displayed lower expression of pericyte-associated markers NG2, PDGFRβ, and fibronectin, with notably lower αSMA, CDDGF-BB.

CONCLUSIONS

Despite differences in their phenotype and magnitude of response, both P-pericytes and D-pericytes responded similarly to all examined functions, indicating that the neuroinflammatory phenotype of these cells is robust to culture conditions.

Neovascularization and re-endothelialization rely on endothelial progenitor cells (EPCs). However, the recruitment and angiogenic roles of EPCs are subject to regulation through the vascular microenvironment, which remains largely unknown. Platelet‑derived growth factor D (PDGF‑D) is a new member of the PDGF family that binds the PDGFR‑β homodimer. However, it remains unknown whether and how it affects the angiogenic capacity of EPCs and participates in tube‑like formation. EPCs were derived from rat bone marrow cells, and the gain‑of‑function approach was used to study the effects of PDGF‑D on the biological activities of EPCs. EPCs that stably express PDGF‑D were generated by lentiviral‑mediated transduction, and the expression levels were evaluated by western blotting and reverse transcription, followed by real‑time quantitative polymerase chain reaction (RT‑qPCR). The biological activities of EPCs evaluated in the present study included proliferation, adhesion, migration, tube formation and senescence. Furthermore, the downstream signaling of PDGF‑D was explored by western blot analysis. The results revealed that the lentiviral‑mediated expression of PDGF‑D in the microenvironment promoted the migration, proliferation, adhesion and tube formation of EPCs. In addition, PDGF‑D suppressed the senescence of EPCs. Mechanistically, PDGF‑D induced the phosphorylation of several signaling molecules, including STAT3, AKT, ERK1/2, mTOR and GSK‑3β, suggesting that PDGF‑D enhanced the angiogenic function of EPCs through PDGF receptor‑dependent and ‑independent signaling pathways. In conclusion, PDGF‑D promotes the angiogenic capacity of EPCs, including proliferation, migration, adhesion and tube formation, and thereby contributes to angiogenesis.

Background: Full thickness burn wounds are lack of angiogenesis, cell migration, epithelialisation and finally scar tissue formation. Tissue engineered composite graft can provide sustained release of growth factor and promote the wound healing by cell migration, early angiogenesis and proliferation of extracellular matrix and wound remodeling. The objective of this study was to evaluate the gene embedded (pDNA-platelet-derived growth factor, PDGF-B) porcine acellular urinary bladder matrix with transfected mesenchymal stem cells (rBMSC) on healing of full thickness burn wound in rat model.

Methods: Full thickness burn wound of 2 × 2 cm size was created in dorsum of rat model under general anesthesia. Burn wounds were treated with silver sulfadiazine; porcine acellular urinary bladder matrix (PAUBM); PAUBM transfected with pDNA-PDGF-B; PAUBM seeded with rBMSC; PAUBM seeded with rBMSC transfected with pDNA-PDGF-B in groups A, B, C, D and E respectively. The wound healing was assessed based on clinical, macroscopically, immunologically, histopathological and RT-qPCR parameters.

Results: Wound was significantly healed in group E and group D with early extracellular matrix deposition, enhanced granulation tissue formation and early angiogenesis compared to all other groups. The immunologic response against porcine acellular matrix showed that PDGF-B gene activated matrix along with stem cell group showed less antibody titer against acellular matrix than other groups in all intervals. PDGF gene activated matrix releasing the PDGF-B and promote the healing of full thickness burn wound with neovascularization and neo tissue formation. PDGF gene also enhances secretion of other growth factors results in PDGF mediated regenerative activities. This was confirmed in RT-qPCR at various time intervals.

Conclusion: Gene activated matrix encoded for PDGF-B protein transfected stem cells have been clinically proven for early acceleration of angiogenesis and tissue regeneration in burn wounds in rat models. Evaluation of PDGF-B gene-activated acellular matrix and mesenchymal stem cell in full thickness skin burn wound in rat.

Keywords: Acellular matrix; Burn wound; Gene therapy; PDGF-B; Stem cell.

Previous epidemiological studies showed that chronic arsenic exposure is related to increased cardiovascular disease incidence. The detailed biochemical mechanisms by which arsenic exerts its effects remain unknown. Vascular disease progression is characterized by smooth muscle cell (SMC) phenotypic switching, vessel wall reorganization, and platelet-derived growth factor (PDGF) production. The objective of this study was to examine early biochemical and structural changes in the aortas of ICR mice systemically exposed to arsenic. Animals were fed sodium arsenite (20 mg/kg) via gavage 5 days/week or Milli-Q water only (control) for 8 weeks. Aortic proteins were subjected to two-dimensional (2-D) differential gel electrophoresis and proteomic studies. Two 2-D gel protein spots were identified as the same protein, smooth muscle (SM)22α, using proteomics. SM22α and Rho kinase 2 gene and protein expression were significantly decreased in the aortic tissue of arsenic-exposed mice compared with that of control mice. No atherosclerotic lesion formation or tissue injury was detected in the aortic wall of either the arsenic-fed or the control group. However, the percent (%) SMC area of the aortic wall was significantly decreased in arsenic-fed mice compared with that in control mice. Additionally, the expression levels of PDGF-BB and early growth response-1 (Egr-1) were significantly higher in the arsenic group than that in the control group. These findings reveal biochemical alterations of SM22α, PDGF, and Egr-1 in conjunction with decreased SMC area in the aortic wall of arsenic-fed mice. Arsenic may initiate aortic SMC alterations that subsequently lead to vascular dysfunction.

Intestinal resection and anastomosis are performed in over a million people with various bowel diseases annually. Excessive fibrosis and anastomotic site leakage are the main complications of anastomosis surgery, despite great improvements in operative technique and equipment in recent years. In this study, cRGD modified poly(p-dioxanone-co-l-Phe) (PDPA) membranes are designed and applied in intestinal anastomosis to simultaneously solve the two aforementioned complications. cRGD is modified onto PDPA membranes through both physical absorption and π-π accumulation between d-Phe of cRGD and l-Phe of PDPA. Although cRGD modification enhanced the biocompatibility of PDPA membranes, cRGD modified PDPA membrane suppresses fibroblast proliferation both in vitro and in vivo as a result of degradation and subsequent release of fibroblast suppressive l-Phe from PDPA. Meanwhile, platelets are entrapped by cRGD modified PDPA membranes through the specific binding of cRGD and platelet GP_IIbIIIa . cRGD modified PDPA membranes are applied in rat intestinal anastomosis, and both adhesion and stenosis are successfully prevented at anastomotic sites. At the same time, bursting pressure, which represents healing intensity at anastomotic sites, is promoted. The gathering and activation of platelets on PDPA membranes induce secretion of autologous PDGF and VEGF to facilitate angiogenesis and subsequent healing of anastomotic sites.

Platelet-derived growth factor is commonly known as a mitogen. There are four members of PDGF family known as PDGF-A chain, PDGF-B chain, PDGF-C chain and PDGF-D chain, which in active forms are dimers. As far as two receptors PDGF-alphaR and PDGF-betaR are known to bind PDGF There is a difference in binding affinity of various forms of PDGF by those receptors. Two different transcripts are derived from PDGF-A gene by alternative splicing. Transcription of PDGF-A gene is under control of many factors. Many research data suggest a role for PDGF-A in smooth muscle cell hyperplasia in hypertension and atherosclerosis. Since inhibition of PDGF-A transcription by a specific ribozyme represses smooth muscle cell proliferation in arteries, introduction of gene therapy might be of value in the treatment of hypertension and its complications.

OBJECTIVE

To study the efficacy of 40-O-(2-hydroxyethyl)-rapamycin (SDZ RAD) in preventing chronic renal allograft rejention in rats.

METHODS

The rat model of chronic renal allograft rejection was made with micro-surgery. The recipients were divided into two groups. The recipients in the study group were treated with SDZ RAD at a dose of 0.5 mg x kg(-1) x d(-1) by means of gavage and the controls with vehicle.

RESULTS

After 24 weeks of treatment, proteinuria, glomerular sclerosis, vascular intimal thickening and the infiltration of monocytesmacrophages and lymphocytes in the study group were decreased, in consistence with the reduced expression of ICAM-1, VCAM-1, TGF-beta(1) and PDGF-AA mRNA.

CONCLUSIONS

It is suggested that SDZ RAD can prevent chronic renal allograft rejection and the reduced expression of above adhesion molecules and growth factors may be involved in the mechanism.

The objective of this study was to investigate the association of cell orientation around a biomaterial with expression of a contractile actin isoform. Selected cytokines and a fungal metabolite known to alter the cytoskeleton were used to modulate the fibroblast orientation around titanium in vitro and the synthesis of a specific muscle actin in order to reveal an association between these processes. A novel culture system using a fibronectin-coated silicone surface was employed to evaluate the orientation of human gingival fibroblasts around titanium discs. Round glass cover slips, 25 mm in diameter, were coated with polydimethylsiloxane. During the heat-induced polymerization process, two commercially pure titanium discs, 5 mm in diameter, were placed on the silicone at a distance of approximately 0.5 mm apart. The rubbery consistency of the silicone stabilized the metal discs on the cover slip and eliminated the risk of developing a lip at the edge of the titanium sample. The cover slip was then heated to complete polymerization of the silicone and subsequently coated with fibronectin. One hundred thousand human gingival fibroblasts were plated onto each glass cover slip containing the titanium discs. The cells were treated with one of the following prior to seeding on the cover slips: transforming growth factor-beta1 (TGF-beta1), platelet-derived growth factor-BB (PDGF-BB), interferon-gamma (IFN-gamma) for cytochalasin-D. Untreated cells served as controls. The orientation of the cells at the surface of the titanium discs was evaluated microscopically and the cell content of alpha-smooth muscle actin (SMA) was determined by Western blot analysis and immunohistochemistry. A notable finding was the high correlation between the percentage of cells oriented perpendicular to the titanium surface and SMA synthesis. TGF-beta1, IFN-gamma and cytochalasin-D increased synthesis of SMA while PDGF-BB decreased it. The findings support the proposition that SMA-enabled cell contraction may play a role in the orientation of cells to a biomaterial surface.

We previously reported that corneal fibroblasts within 3D fibrin matrices secrete, bind, and organize fibronectin into tracks that facilitate cell spreading and migration. Other cells use these fibronectin tracks as conduits, which leads to the development of an interconnected cell/fibronectin network. In this study, we investigate how cell-induced reorganization of fibrin correlates with fibronectin track formation in response to two growth factors present during wound healing: PDGF BB, which stimulates cell spreading and migration; and TGFβ1, which stimulates cellular contraction and myofibroblast transformation. Both PDGF BB and TGFβ1 stimulated global fibrin matrix contraction (p < 0.005); however, the cell and matrix patterning were different. We found that, during PDGF BB-induced cell spreading, fibronectin was organized simultaneously with the generation of tractional forces at the leading edge of pseudopodia. Over time this led to the formation of an interconnected network consisting of cells, fibronectin and compacted fibrin tracks. Following culture in TGFβ1, cells were less motile, produced significant local fibrin reorganization, and formed fewer cellular connections as compared to PDGF BB (p < 0.005). Although bands of compacted fibrin tracks developed in between neighboring cells, fibronectin labeling was not generally present along these tracks, and the correlation between fibrin and fibronectin labeling was significantly less than that observed in PDGF BB (p < 0.001). Taken together, our results show that cell-induced extracellular matrix (ECM) reorganization can occur independently from fibronectin patterning. Nonetheless, both events seem to be coordinated, as corneal fibroblasts in PDGF BB secrete and organize fibronectin as they preferentially spread along compacted fibrin tracks between cells, producing an interconnected network in which cells, fibronectin and compacted fibrin tracks are highly correlated. This mechanism of patterning could contribute to the formation of organized cellular networks that have been observed following corneal injury and refractive surgery.

Keywords: 3-D matrices; collective cell migration; corneal fibroblasts; fibrin; fibronectin.

The selective blockage of platelet-derived growth factor BB (PDGF-BB), basic fibroblast growth factor (bFGF), and transforming growth factor beta1 (TGF-beta1) by specific antibodies coated into expanded polytetrafluoroethylene (ePTFE) grafts may diminish neointimal hyperplasia. Sixty pigs were divided into two groups (n = 30 each) and then further divided into five subgroups. Group 1 had a bilateral iliac artery ePTFE interposition graft precoated with Matrigel. Three subgroups (A, B, and C) received a specific monoclonal antibody against PDGF-BB, bFGF, or TGF-beta1. One (D) received all antibodies, and one served as control (nonimmune immunoglobulin G [IgG] isotypes) (E). Group 2 had a bilateral iliac artery endothelial cell (EC)-seeded ePTFE interposition graft precoated with Matrigel. Three subgroups (A, B, and C) received a specific antibody against PDGF-BB, bFGF, or TGF-beta1. One (D) received all antibodies, and one served as control (nonimmune IgG isotypes) (E). Light microscopy and immunohistochemical stain showed that neointimal hyperplasia formation was significantly reduced in subgroups D compared to the others (p < 0.05). In subgroups D, the different precoating influenced neointimal hyperplasia formation. It was more pronounced in the prosthesis precoated with EC and Matrigel (p < 0.05). In organ culture, the amount of PDGF-BB, bFGF, and TGF-beta1 release was reduced in subgroup D animals compared to the others (p < 0.05). In subgroups D, the release of PDGF-BB, bFGF, and TGF-beta1 depended on ePTFE seeding. A higher amount of these growth factors was released in the prostheses precoated with EC and Matrigel (p < 0.05), and the bromodeoxyuridine labeling index confirmed higher incorporation in this subgroup (p < 0.001). The combined use of locally administered anti-PDGF-BB, bFGF, and TGF-beta1 monoclonal antibodies reduces neointimal hyperplasia formation.

A series of 2-phenyl-1H-benzo[d]imidazole-4,7-diones were synthesized and tested for their inhibitory activity on the PDGF-stimulated proliferation of rat aortic vascular smooth muscle cells. Among the tested compounds, 6-arylthio-5-chloro-2-phenyl-1H-benzo[d]imidazole-4,7-diones exhibited an potent antiproliferative activity.

OBJECTIVE

To detect the relationship between platelet-derived growth factor-A (PDGF-A), basic fibroblast growth factor (bFGF), and proliferation of smooth muscle cells in vein grafts.

METHODS

An animal model of autogenous vein graft was established by transplanting internal branch of the jugular vein to the common iliac artery by end-to-end anastomosis. 25 rats were used. The grafted veins were harvested at 6 h, 2 d, 1 w, 2 w and 4 w respectively after the operation. The expression of PDGF-A and bFGF in different stages after grafting procedure was observed immunohistochemically.

RESULTS

The expression of PDGF-A peaked at 1 w, paralleled with proliferation of SMC, and was significantly higher than that at 6 h (P < 0.01). The expression of PDGF-A diminished gradually. At 4 w, it was significantly lower than that at 1 w (P < 0.01). bFGF demonstrated bimodal pattern of the expression: the first peak of expression occurred by 2 d and the second peak by 2 w. The difference was significantly compared with other stages.

CONCLUSIONS

The expression of PDGF-A 1 contributed to migration and proliferation of SMC. Except for contributing to proliferation of SMC, bFGF perhaps took part in endothelization.

Serum-deprived (0.2%) resting NIH 3T3 mouse fibroblasts, stimulated for 0.5-4.0 h by serum (10%) or PDGF (beta-beta homodimer) (early cells), were fused with stimulated cells taken 10 h after changing the medium to one containing 10% serum DNA synthesis was investigated in the nuclei of monokaryons, homodikaryons and heterodikaryons, using radioautography with the double-labeling technique. Preincubation of resting cells with serum for 3 h or with the competence factor PDGF for 1.0-1.5 h abolished their ability to suppress DNA synthesis in late stimulated nuclei in heterodikaryons. Actomycin D restores the inhibitory capacity of resting cells stimulated by PDGF but not by serum. The results give evidence that the acquirement by resting cells of competence for DNA replication includes as a necessary step the down-regulation of intracellular growth inhibitors, and thus serum can overcome the influence of endogenous growth inhibitors, at any rate, by two ways.

Guanxinshutong capsule (GXST), which consists of five traditional Chinese medicines, has been used for a long time in China for the treatment of cardiovascular diseases, such as coronary artery disease and myocardial infarction. However, the effects on GXST on myocardial injury (MI) have not been studied in detail. In these experiments, we found that GXST administration decreased MI-associated ventricular remodeling (VR) with a reduction in interventricular septal thickness in diastole (IVSd), left ventricular posterior wall diameter in systole (LVPWs), and left ventricular posterior wall diameter in diastole (LVPWd) to ameliorate cardiac function and architecture, as measured by echocardiography. Furthermore, histological analysis showed that GXST could ameliorate pathological alterations in the myocardium. And Sirius red staining, wheat germ agglutinin staining and inflammation-related immunohistochemistry results showed that GXST ameliorated the fibrosis areas, cardiac hypertrophy and inflammation (IL-6 and TNF-α). In addition, GXST upregulated intercellular junction proteins (N-cad and Cx-43) and downregulated the angiogenesis-related proteins (PDGF and VEGFA), myocardial fibrosis-related proteins (TGF-β1), and matrix metalloproteinase (MMP-2 and MMP-9). We also found that GXST medium-dose group (1 g/kg/d) dosage was the most efficacious. In conclusion, GXST protected cardiac tissues against MI by reducing VR, thus indicating the potential application of GXST in the treatment of MI.

Keywords: Angiogenesis; Guanxinshutong capsule; Intercellular junction; Myocardial fibrosis; Myocardial injury; Ventricular remodeling.