Expression Of Platelet-derived Growth Factor B-chain Research Articles

Syndecan-1

Arterial injury induces smooth muscle cell (SMC) proliferation, migration, and intimal accumulation of cells and extracellular matrix. These processes are regulated by the administration of the glycosaminoglycans heparin and heparan sulfate, but little is known about the role of endogenous heparan sulfate proteoglycans in the vessel wall. We investigated the response to carotid injury of syndecan-1-null mice to assess the function of one of a conserved family of transmembrane heparan and chondroitin sulfate proteoglycans. Syndecan-1-null mice developed a large neointimal lesion after injury, whereas wild-type mice made little or none. This was accompanied by a significant increase in both medial and intimal SMC replication. Cultured syndecan-1-null SMCs showed a significant increase in proliferation in response to PDGF-BB, thrombin, FGF2, EGF, and serum. In response to thrombin, PDGF-BB, and serum syndecan-1-null SMCs expressed more PDGF-B chain message than did wild-type SMCs. Downregulation of PDGF-BB or PDGFRbeta inhibited thrombin-, PDGF-BB-, and serum-induced DNA synthesis in syndecan-1-null SMCs. These results suggest the possibility that syndecan-1 may limit intimal thickening in injured arteries by suppressing SMC activation through inhibition of SMC PDGF-B chain expression and PDGFRbeta activation.

Association of Platelet-Derived Growth Factor-B Chain with Simian Human Immunodeficiency Virus Encephalitis

Chemokines and cytokines play a critical role in HIV infection, serving both to modulate virus replication and to recruit target cells to the site of infection. Platelet-derived growth factor (PDGF), a mitogen and chemoattractant for a wide variety of cells, is secreted by macrophages. Since macrophages are the target cells for lentiviral infection in the brain and PDGF is a known inducer of macrophage chemoattractant protein-1 (MCP)-1, a potent chemokine closely associated with HIV encephalitis, we investigated the association of PDGF-B chain (PDGF-B) with encephalitis in macaques caused by simian human immunodeficiency virus (SHIV), a chimera of HIV and SIV. Northern blot analysis confirmed elevated expression of PDGF-B chain mRNA in the brains from encephalitic macaques. Validation of these in vivo studies was confirmed in rhesus macrophage cultures infected with SHIV(KU2) in which we demonstrated heightened expression of PDGF-B chain mRNA. Nuclear run-off analysis established transcriptional up-regulation of PDGF-B chain in virus-inoculated macrophage cultures. Reciprocally, addition of exogenous PDGF enhanced virus replication and MCP-1 expression in these cells. Inhibition of virus replication by tyrosine kinase inhibitor, STI-571, and by PDGF-B antisense oligonucleotides confirmed the specificity of the PDGF effect. Relevance of these findings was confirmed by analysis of archival brain tissue from SHIV encephalitic and non-encephalitic macaques for PDGF-B chain expression. PDGF-B chain protein expression was observed in the virus-infected cells in microglial nodules in the brains of SHIV-encephalitic macaques.

The PDGF B-chain is involved in the ontogenic susceptibility of the developing rat brain to NMDA toxicity

Hypoxic–ischemic (H–I) injury to neonatal brains can cause a life-long neuronal deficit because of increased susceptibility in the neonatal period. Excitotoxicity due to overstimulation of the N-methyl- d-aspartate receptor (NMDAR) is assumed to be the basis of the injury. However, the ontogenic profile of the susceptibility does not directly correlate with the levels of NMDAR expression. Platelet-derived growth factor B-chain (PDGF-B) has been reported to protect neurons by suppressing the NMDA-evoked current and translocating the glutamate transporter to the cell membrane. Thus, we assessed the relationship between the susceptibility to H–I injury and the expression of PDGF-B in neonatal rat brain. PDGF-B infusion before and after an intrastriatal NMDA injection significantly reduced the size of the lesions in 7-day-old rats, when they are most susceptible and the neuronal expression of PDGF-B is low. Fourteen-day-old neonatal rats were found to be resistant to NMDA injury, even though NMDARs are expressed at high levels in the brain at this age. Inhibition of PDGF-B protein synthesis by antisense oligodeoxynucleotides increased the size of the NMDA-induced lesions up to 6-fold at postnatal day 14, when PDGF-B is expressed at high levels in neurons. These data suggest that PDGF-B is an important physiological modulator of NMDAR excitability in the developing brain, and that the balance between the expression of NMDAR and PDGF-B partly determines the ontogenic susceptibility to brain injury. Enhancement of the PDGF-B/receptor signal pathway might rescue neonatal brains at risk of H–I injury.

Growth factor expression in a murine model of cryoglobulinemia

Increased expression of growth factors including platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta) are thought to play pivotal roles during mesangial expansion and glomerulosclerosis. Thymic stromal lymphopoietin (TSLP) transgenic mice develop mixed cryoglobulinemia and a membranoproliferative glomerulonephritis (MPGN). Here we describe the renal expression of isoforms of PDGF and TGF-beta in relation to changes in extracellular matrix (ECM) components and markers of cell proliferation and activation in this model. A total of 123 mice, including 61 TSLP transgenic mice and 62 wild-type controls, were sacrificed at defined intervals. PDGF-A chain, -B chain, PDGF alpha- and beta-receptor (beta-R) and TGF-beta1 mRNA were analyzed by in situ hybridization. Expression of alpha smooth muscle actin (alphaSMA), collagen type I, collagen type IV, laminin, and a marker of proliferating cells (PCNA) were assessed by immunohistochemistry. Slides also were studied by combined immunohistochemistry and in situ hybridization with an antibody that recognizes monocytes/macrophage and with riboprobes that detect PDGF B-chain, PDGF beta-R or TGF-beta1 mRNA. Increased numbers of proliferating glomerular cells appeared early in the disease course, associated with de novo expression of alphaSMA. Expression of PDGF B-chain and beta-R mRNA was increased in the mesangium and in parietal epithelial cells of TSLP transgenic mice and correlated with the number of PCNA positive cells. Increased TGF-beta1 mRNA expression paralleled the deposition of type IV collagen. A significant proportion of Mac-2 positive macrophages expressed TGF-beta1 mRNA, while only a small percentage of glomerular macrophages expressed PDGF B-chain mRNA. No PDGF beta-R mRNA expression by macrophages was detected. TSLP transgenic mice develop a membranoproliferative glomerulonephritis in which glomerular cell proliferation and matrix deposition are associated with an increased expression of PDGF B-chain, PDGF beta-R and TGF-beta1. These findings extend the paradigms covering these growth factors established in the rat Thy 1 model of mesangiolysis and repairs to a murine model of progressive glomerulonephritis closely resembling human MPGN.

Localization of SPARC in developing, mature, and chronically injured human allograft kidneys

The matricellular protein SPARC (secreted protein acidic and rich in cysteine) is expressed during development, tissue remodeling and repair. It functions as an endogenous inhibitor of cell proliferation, regulates angiogenesis, regulates cell adhesion to extracellular matrix, binds cytokines such as platelet derived growth factor and stimulates transforming growth factor-beta (TGF-beta) production. This study describes the expression of SPARC during human renal development, in normal kidneys and during renal allograft rejection. A total of 60 renal specimens, including normal areas from tumor nephrectomies (N = 24), fetal kidneys (N = 27) and explanted renal allografts (N = 9), were included in the study. SPARC protein was localized by immunohistochemistry using two different antibodies. On consecutive sections SPARC mRNA was detected by in situ hybridization. In the normal adult kidney SPARC protein was expressed by visceral and parietal epithelial cells, collecting duct epithelium (CD), urothelium, smooth muscle cells of muscular arteries and focally in interstitial cells. During renal development immature glomeruli demonstrated a polarized SPARC expression in visceral epithelial cells at their surface abutting the capillary basement membranes. In the fully differentiated glomeruli the expression pattern mirrored that of the adult kidney. Furthermore, SPARC was abundantly expressed by derivatives of the ureteric bud, and smooth muscle cells of arterial walls. During chronic allograft rejection SPARC is expressed in neointimal arterial smooth muscle cells, infiltrating inflammatory cells as well as by interstitial myofibroblasts in areas of interstitial fibrosis. SPARC mRNA synthesis detected by in situ hybridization mirrored these protein expression patterns. These studies co-localize SPARC to several sites of renal injury previously shown to be sites of PDGF B-chain expression and/or activity. We speculate that SPARC could function as an accessory molecule in chronic PDGF-mediated sclerosing interstitial and vascular injury. SPARC localization to glomerular epithelial cells corresponds to similar findings in rodents, and may reflect its role in cell adhesion and /or regulation of cell shape.

Von Hippel-Lindau tumor suppressor protein represses platelet-derived growth factor B-chain gene expression via the Sp1 binding element in the proximal PDGF-B promoter.

VHL is the causative gene for von Hippel-Lindau disease and sporadic clear cell renal cancer. It has been shown that pVHL can suppress the expression of certain genes that are overexpressed in renal carcinomas. One such gene is that encoding the potent mitogen and chemoattractant, platelet-derived growth factor B-chain (PDGF-B). The regulatory mechanisms underlying pVHL suppression of PDGF-B expression, however, are completely unknown. This understanding would shed vital light on the control of growth factor gene expression by this tumor suppressor. Here we report that pVHL can repress both endogenous steady-state PDGF-B mRNA expression and PDGF-B promoter-dependent transcription in WKY12-22 cells. Transient transfection analysis utilizing PDGF-B promoter-chloramphenicol acetyl transferase (CAT) reporter constructs revealed that pVHL inhibition of PDGF-B expression is mediated via the Sp1-binding element in the proximal region of the PDGF-B promoter. Recent studies have demonstrated a physical interaction between pVHL and Sp1, which activates the PDGF-B promoter. We show that Sp1 can rescue PDGF-B promoter activity and endogenous PDGF-B mRNA expression from pVHL repression. These findings thus demonstrate a pivotal role for Sp1 in pVHL inhibition of PDGF-B transcription.

Induction of Platelet-derived Growth Factor B-chain Expression by Transforming Growth Factor-β Involves Transactivation by Smads

Transforming growth factor-beta (TGF-beta) regulates a diverse array of biological processes, such as proliferation, differentiation, extracellular matrix production, and apoptosis. In cultured vascular endothelial cells, TGF-beta induces the expression of platelet-derived growth factor (PDGF) B-chain, a mitogen and chemoattractant, at the level of transcription. The molecular mechanism(s) underlying this process are not presently understood. In this study, we performed serial 5' deletion and transient transfection analysis to define a region in the PDGF-B promoter mediating inducible responsiveness to TGF-beta. This region contains an atypical nucleotide recognition element for the Smad family of transcriptional regulators. Electrophoretic mobility shift analysis revealed that nuclear proteins bound to this site in a transient and specific manner. Supershift studies demonstrated the physical association of Smad4 with the promoter. Overexpression of Smad4 activated the PDGF-B promoter and superinduced PDGF-B promoter-dependent expression in cells exposed to TGF-beta. Moreover, simultaneous cotransfection of Smad3 and Smad4 activated the PDGF-B promoter. This effect was attenuated when Smad4 was substituted with its dominant negative counterpart. Mutation of the (-81)CAGA(-78) motif in the PDGF-B promoter abrogated Smad-inducible promoter-dependent expression. Overexpression of Smad2 and Smad3 transactivated the PDGF-B promoter in a synergistic manner. These findings demonstrate the existence of a novel, functional binding element in the proximal region of the PDGF-B promoter mediating responsiveness to TGF-beta.

Immunohistochemical characterization of glomerular PDGF B-chain and PDGF β-receptor expression in diabetic rats

Platelet-derived growth factor (PDGF) was found to contribute to the pathophysiological process in the development and progression of glomerulosclerosis characterized by mesangial cell proliferation and accumulation of extracellular matrix. To examine the role of PDGF in the development of diabetic nephropathy, we conducted immunohistochemical analysis for PDGF B-chain (PDGF-B) and PDGF β-receptor (PDGFR-β) in the glomeruli of streptozotocin-induced diabetic rats. At 2, 4, and 12 weeks after the onset of diabetes, the expression of PDGF-B in glomeruli of diabetic rats was increased significantly as compared to control or diabetic rats treated with insulin. Similar changes were observed on PDGFR-β immunostaining. The immunostaining of mirror sections revealed the existence of PDGF-B or PDGFR-β not only in mesangial cells but also in visceral epithelial cells. Glomerular volume was significantly increased in diabetes. This early glomerular abnormality was prevented by an inhibition of PDGF system with trapidil as well as by the treatment of insulin. Our results suggest that the activation of the PDGF system in glomerular cells might play an important role in the development of early glomerular lesion in diabetes.

Role of PDGF B-Chain and PDGF Receptors in Rat Tubular Regeneration after Acute Injury

Various polypeptide growth factors are generally considered to be involved in the regulation of the nephrogenic process both after acute renal injury and during renal development. Because platelet-derived growth factor B-chain (PDGF-B) has been reported to be expressed in immature tubulus of the developing kidney, PDGF-B could play a role in the process of tubulogenesis. We examined the expression of PDGF-B and PDGF receptors alpha and beta and their localization in kidneys after ischemia/reperfusion injury. The mRNA expressions of PDGF-B, PDGFR-alpha, and PDGFR-beta were enhanced after injury. In the immunohistochemical analysis and/or in situ hybridization, PDGF-B and PDGFR-alpha, beta were expressed after reperfusion in the S3 segment of the proximal tubuli, where they were not expressed normally. The expressions of proliferating cell nuclear antigen and vimentin were concomitantly observed with PDGF-B and PDGFRs in the tubular cells of injured S3 segment at 48 hours after injury. Next, the inhibition of the PDGF-B/PDGFRs axis with either Trapidil or Ki6896, which was found to inhibit the phosphorylation of PDGFR-beta selectively, resulted in a rise of serum creatinine, higher mortality rate, abnormal regenerating process, and suppressed proliferation of tubular epithelial cells. These findings suggest that the PDGF-B/PDGFRs axis is involved in the proliferation of injured tubular cells and plays an important role in the regeneration of tubular cells from acute ischemic injury.

Angiotensin II stimulates platelet-derived growth factor-B chain expression in newborn rat vascular smooth muscle cells and neointimal cells through Ras, extracellular signal-regulated protein kinase, and c-Jun N-terminal protein kinase mechanisms.

Platelet-derived growth factors (PDGFs) have been implicated in the pathogenesis of vascular proliferative disorders. Vascular smooth muscle cells (VSMCs) are one of the cell types that produce PDGF-B chain in proliferative lesions, although the mechanism of regulation of PDGF-B chain production in these cells is not well understood. In the present study, we demonstrate that angiotensin II (Ang II), which is also implicated in vascular stenosis after angioplasty and atherosclerosis, markedly stimulates PDGF-B chain mRNA expression in cultured newborn rat medial VSMCs and neointimal VSMCs via an AT(1), but not in adult rat VSMCs. In newborn rat VSMCs, Ang II activates extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal protein kinase (JNK), and p38 mitogen-activated protein kinase. The mitogen-activated protein/ERK (MEK) inhibitor PD98059, but not the p38 inhibitor SB203580, abrogates Ang II-induced PDGF-B mRNA expression. Transient transfection analysis using a PDGF-B promoter-luciferase gene reporter construct reveals that Ang II induces transcriptional activation of PDGF-B chain gene, which is abolished by the expression of a dominant negative form of either ERK or JNK, but not of p38. The expression of a dominant negative form of Ras abolishes the stimulatory effects of Ang II on ERK activity and PDGF-B mRNA expression. In adult rat VSMCs, Ang II activates ERK and JNK, but weakly induces Egr-1, a transcription factor implicated in PDGF-B chain gene expression, compared with newborn VSMCs. These data indicate that Ang II activates PDGF-B chain gene expression in VSMCs through mechanisms involving Ras-ERK and JNK.

Quantitative analysis of growth-related factors in human pituitary adenomas: Lowered insulin-like growth factor-I and its receptor mRNA in growth hormone-producing adenomas

To elucidate the contribution of growth factors to the development, growth and behavior of human pituitary adenomas, the authors used competitive reverse transcription-polymerase chain reactions to quantify expression of mRNAs for growth factors extracted from pituitary adenomas. As previously diagnosed by endocrinologic evaluation, the pituitary adenomas in this study consisted of six prolactin-producing, six growth hormone (GH)-producing, four follicle-stimulating hormone producing and six nonfunctioning adenomas. The mRNAs examined included those for platelet-derived growth factor (PDGF) B-chain, transforming growth factor (TGF)-β1, epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and insulin-like growth factor (IGF)-I and -II; proliferating cell nuclear antigen (PCNA) as an indicator of cell proliferation; and pituitary-specific transcription factor-1 (Pit-1) which is a nuclear transcription factor expressed in the anterior pituitary. All factors except the last were expressed in all adenomas, and expression of PDGF B-chain, TGF-β1, EGF, bFGF and IGF-II did not differ between the four adenoma varieties. Pit-1 was expressed only in GH- and prolactin-producing adenomas. PCNA expression also showed no differences. However, IGF-I mRNA in GH-producing adenomas was significantly lower than in prolactin-producing and nonfunctioning adenomas despite high serum IGF-I levels (1121±253 ng/ml). The analysis on IGF-I receptor mRNA was significantly lowered in GH-producing adenoma compared with the other types of adenoma. These findings suggest that the attenuation of negative feedback through the pituitary GH–IGF-I axis may be involved in development of GH-producing adenoma.

Effect of simvastatin on proliferative nephritis and cell-cycle protein expression

Mesangial cell proliferation is important in subsequent mesangial matrix expansion in glomerular injury. Therefore, the regulation of mesangial cell proliferation may be critical in the treatment of glomerulonephritis. Inhibition of 3-hydro-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibits the production of mevalonate and has been shown to suppress proliferation in many cell types, including mesangial cells in vitro. It is expected that HMG-CoA reductase inhibitor may suppress mesangial cell proliferation and subsequent progression of glomerulonephritis. Recently, the tight relationship between cell-cycle regulatory protein expression and mesangial cell proliferation in experimental glomerulonephritis was demonstrated. The aim of the present study is to examine the effect of simvastatin, one of the HMG-CoA reductase inhibitors, on the glomerular cell proliferation and on the expression of CDK2 or p27Kip1 in mesangial cells in experimental glomerulonephritis in vivo. The effect of simvastatin on a rat mesangial proliferative glomerulonephritis induced by antithymocyte antibody (anti-Thy 1.1 GN) was studied. Administration of simvastatin or vehicle (for control GN) were started from two days before disease induction, and was continued to the day of nephrectomy. Nephrectomy was done at days 0, 2, 4, 7, 12 and 20 after disease induction. Immunohistochemistry for proliferating cells, macrophages, alpha-smooth muscle actin, type IV collagen and PDGF-B chain was performed, respectively, in addition to conventional periodic-acid Schiff staining. Double immunostaining for CDK2/OX-7 or p27Kip1/OX-7 was also done, respectively. There was no difference in the degree of the initial injuries between simvastatin-treated and control GN rats. The most pronounced feature of simvastatin-treated GN was the suppression of the early glomerular cell proliferation (about 70% of proliferation was suppressed at day 4). At day 4, alpha-smooth muscle actin expression was also decreased in simvastatin-treated GN rats. Inhibition of macrophage recruitment into glomeruli by simvastatin was also a prominent feature (about 30% decrease in the number of glomerular macrophages at day 2). Simvastatin significantly suppressed subsequent mesangial matrix expansion and type IV collagen accumulation in glomeruli. Although it might simply reflect the reduction in mesangial cells, glomerular PDGF-B chain expression was reduced. There was no significant difference in plasma lipids levels at day 2 and day 4. In vehicle-treated GN rats, the number of CDK2+/OX-7+ cells (CDK2-expressed mesangial cells) in glomeruli increased significantly from day 4 to day 7. Although simvastatin suppressed mesangial cell proliferation, the increase in the number of glomerular CDK2+/OX-7+ cells was also attenuated by simvastatin treatment. There was no difference in the number of p27Kip1+/OX-7+ cells (p27Kip1-expressed mesangial cells) in the glomerulus between vehicle-treated and simvastatin-treated GN rats. Simvastatin suppressed mesangial cell proliferation and subsequent matrix expansion, and macrophage infiltration into glomeruli in anti-Thy 1.1 GN rats. The antiproliferative effect of simvastatin in this model was also associated with the reduction of CDK2 expression in mesangial cells.

Expression of bcl-2 and bax in glomerular disease.

Bcl-2 may account in part for the maintenance of hypercellularity in human glomerular diseases through preventing cell death and by counteracting bax which may be expressed to regulate excessive proliferation. This process is associated with the effect of PDGF B-chain expression. Bax expression may be important in the cell loss leading to glomerulosclerosis and TGF-beta1 participates in this process by increasing bax expression. Thus, the balance of bcl-2/bax expression may be critical in the course of human glomerular diseases.

Expression of platelet-derived growth factor B-chain and platelet-derived growth factor β-receptor in fibroblasts of scleroderma

Scleroderma is a fibrotic disease occurring in a localized or systemic form. The pathogenesis of scleroderma remains poorly understood. Recent studies revealed that various cytokines and growth factors were involved in the development of scleroderma fibrosis. Platelet-derived growth factor (PDGF) is a potent growth factor for mesenchymal cells, especially fibroblasts. It can promote fibroblasts proliferation, enhance extracellular matrix synthesis. It is also a chemoattractant to fibroblasts. To better understand the role of PDGF in pathogenesis of scleroderma, we performed both in vivo studies on the expression of PDGF β-receptor protein in scleroderma tissue and in vitro studies on the expression of PDGF B-chain and PDGF β-receptor mRNA in cultured fibroblasts derived from both lesions of scleroderma and normal skin. Immunohistochemistry staining showed that PDGF β-receptor expression was greatly elevated in the dermis of scleroderma lesion whereas PDGF β-receptor were expressed at low levels in normal skin. Northern blot analysis showed that cultured fibroblasts from scleroderma had higher expression of PDGF B-chain and PDGF β-receptor mRNA than those from normal control. Two PDGF B-chain mRNA transcripts, 2.8 and 4.0 kb, were expressed. The 2.8 kb transcripts which had more efficient translation ability was the more predominantly expressed one. These results indicate that PDGF B-chain/PDGF β-receptor signal pathway might be involved in the development of fibrosis in scleroderma, and that the 2.8 kb PDGF B-chain mRNA transcript may be the main modulation gene.

Platelet-derived growth factor B-chain expression in the rat retina and optic nerve: distribution and changes after transection of the optic nerve.

To test the possible involvement of platelet-derived growth factor B-chain (PDGF-B) in anterograde and retrograde degenerations of the CNS neurons, we studied the changes of PDGF-B localization and its mRNA expression in the rat retina and optic nerve (ON) after unilateral ON transection, using immunohistochemistry and in situ hybridization. In the control retinas immunoreactivity for PDGF-B and its mRNA expression were localized in the retinal ganglion cells (RGCs) and the nerve fiber layer. After ON transection PDGF-B immunoreactivity in the nerve fiber layer started to decrease on post-injury day 3 or 4. Atrophic changes in the RGCs started on day 5 just after the decrease of PDGF expression, and thereafter the RGC number decreased. In the longitudinal section of the ON rostral to the transected site, swollen axons showed intense PDGF-B immunoreactivity and macrophages, and some glial cells revealed a significant increase in both immunoreactivity and hybridization signals. Based on these findings, we hypothesized that the decrease in PDGF-B in RGCs after axotomy causes the loss of RGCs, and that increased PDGF-B expression in the ON plays a role in the cascade of tissue reactions following ON transection.

C-type natriuretic peptide inhibits mesangial cell proliferation and matrix accumulation in vivo

Local C-type natriuretic peptide (CNP) production and CNP receptor expression have been demonstrated in glomeruli. However, the glomerular (patho-)physiological functions of CNP are largely unknown. We therefore investigated the effects of CNP on mesangial cell proliferation and matrix accumulation in the rat mesangioproliferative anti-Thy 1.1 model. Over seven days rats received a continuous infusion (1 microgram/kg/min) of either CNP (N = 6), an irrelevant control peptide (N = 3) or buffer alone (N = 6). Kidney biopsies were performed on days 2, 4 and 8. Few significant differences between the groups were noted on days 2 and 4. Compared to buffer treated rats on day 8, those receiving CNP showed a 35% reduction of glomerular mitoses, a 62% reduction of glomerular uptake of the thymidine analogue BrdU and a significant reduction in glomerular expression of PDGF B-chain. Double immunoperoxidase staining also revealed blunting of proliferating, activated mesangial cells (515 reduction of alpha-smooth muscle actin-/BrdU-positive cells) and macrophage influx. Moreover, there was a marked reduction of mesangial collagen IV and fibronectin accumulation at the protein and mRNA level. Rats receiving the control peptide were indistinguishable from buffer treated rats. Systemic blood pressure was reduced by 10 to 20% in both CNP and control peptide treated rats on day 8, excluding that the findings were due to hemodynamic effects of CNP. Our findings demonstrate that CNP is involved in the regulation of mesangial cell proliferation and matrix production in vivo. The data suggest the existence of a glomerular natriuretic peptide system that may regulate tissue homeostasis and contribute to resolution of mesangioproliferative diseases.

Clinical assessment of the significance of platelet-derived growth factor in patients with immunoglobulin A nephropathy

To examine the role of platelet-derived growth factor (PDGF) in the pathogenesis of IgA nephropathy (IgAN), we investigated the expression of PDGF and the PDGF receptor in glomeruli with immunohistochemistry, the plasma levels of PDGF with ELISA, and the expression of PDGF in peripheral blood monocytes (PBMCs) with reverse transcription polymerase chain reaction (RT-PCR). We also assessed the effect of corticosteroid therapy on the plasma levels of the PDGF B-chain. At the time of kidney biopsy, the expression of the PDGF B-chain and the PDGF β receptor in the glomeruli was up-regulated in patients with IgAN. In addition, the plasma concentration of the PDGF B-chain was significantly higher in patients with IgAN than in normal subjects. Moreover, mRNA expression of PDGF B-chain in PBMCs was up-regulated in patients with IgAN when compared with other patients with glomerulonephritis. We divided the patients into two groups according to the grade of urinary protein excretion (U p) after corticosteroid therapy. In patients in group 1 in whom U p was decreased by more than 50% or 1 gm/day after corticosteroid therapy, the expression of the PDGF B-chain and the PDGF β receptor in the glomeruli was up-regulated. Finally, corticosteroid therapy decreased the plasma levels of PDGF B-chain in patients in group 1. Up-regulation of the PDGF B-chain and β receptor in the glomeruli, elevated plasma levels of PDGF B-chain, and increased expression of PDGF mRNA in PBMCs could be associated with the pathogenesis of IgAN. The plasma concentration of PDGF B-chain may be a useful marker for patients with IgAN who would be responsive to corticosteroid therapy.

Expression of PDGF and PDGF receptor mRNA in glomeruli in IgA nephropathy.

This study investigated the mRNA expression of the platelet-derived growth factor (PDGF) A-chain and B-chain and PDGF-beta receptor in glomeruli of 15 immunoglobulin A (IgA) nephropathy kidneys and those with minimal-change lesion (N = 7), membranous nephropathy (N = 3), and focal segmental glomerulonephritis (N = 5), by using competitive RT-PCR methods. The level of PDGF B-chain and beta receptor mRNA expression in IgA nephropathy was significantly higher than in the other forms of glomerulonephritis, but mRNA expressions of PDGF A-chain were not significantly different. Significant correlations were observed between the urinary protein level and the mRNA level of PDGF-beta receptor expression and PDGF B-chain expression, and between the serum creatinine level and the mRNA level of PDGF-beta receptor expression. The PDGF B-chain and beta-receptor may be upregulated and accelerate cell proliferation in a paracrine or autocrine manner and may play a role in the pathogenesis of IgA nephropathy.

Egr-1-induced endothelial gene expression: a common theme in vascular injury.

A number of pathophysiologically relevant genes, including platelet-derived growth factor B-chain (PDGF-B), are induced in the vasculature after acute mechanical injury. In rat aorta, the activated expression of these genes was preceded by a marked increase in the amount of the early-growth-response gene product Egr-1 at the endothelial wound edge. Egr-1 interacts with a novel element in the proximal PDGF-B promoter, as well as with consensus elements in the promoters of other genes induced by endothelial injury. This interaction is crucial for injury-induced PDGF-B promoter-dependent expression. Sp1, whose binding site in the PDGF-B promoter overlaps that of Egr-1, occupies this element in unstimulated cells and is displaced by increasing amounts of Egr-1. These findings implicate Egr-1 in the up-regulated expression of PDGF-B and other potent mediators in mechanically injured arterial endothelial cells.

Mechanisms involved in the pathogenesis of tubulointerstitial fibrosis in 5/6-nephrectomized rats

The 5/6 nephrectomy model is used to study pathogenetic mechanisms underlying chronic renal failure. We previously demonstrated that increased mesangial cell proliferation and glomerular PDGF B-chain expression precede glomerulosclerosis in this model. In the present study we have assessed the concomitant changes in the cortical tubulointerstitium. A wave of tubular and interstitial cell proliferation (as determined by immunostaining for PCNA) occurred at week 1 after 5/6 nephrectomy. This wave preceded the peak glomerular cell proliferation by one week. Tubulointerstitial cell proliferation decreased thereafter and reached control values by week 10. In situ hybridization and immunostaining for PDGF B-chain and beta-receptor in sham-operated controls showed labeling of distal tubules and collecting ducts, while no signal was present in the interstitium. PDGF B-chain mRNA and protein expression was markedly increased in tubules at weeks 2 and 4 after 5/6 nephrectomy and in the interstitium (particularly in areas of inflammatory infiltrates) at weeks 2 to 10. Similar changes occurred with PDGF receptor beta-subunit immunostaining. Interstitial expression of desmin and alpha-smooth muscle actin (markers of myofibroblasts) progressively increased after week 1. Interstitial influx of monocytes/macrophages with focal accentuation started at week 2. Counts of lymphocytes, neutrophils and platelets showed only minor changes. In parallel to the monocyte/macrophage influx, progressive interstitial accumulation of collagens I and IV, laminin, and fibronectin occurred. All of these changes were correlated with the increase in serum creatinine, proteinuria and an index of tubulointerstitial damage. We conclude that tubulointerstitial changes after 5/6 nephrectomy show similarities with those observed in the glomeruli. Tubular and interstitial overexpression of PDGF B-chain and its receptor may play a role in mediating fibroblast migration and/or proliferation in areas of tubulointerstitial injury.