Pathogenesis Of Chronic Allograft Nephropathy Research Articles

Identification of Hub Gene and Transcription Factor Related to Chronic Allograft Nephropathy Based on WGCNA Analysis

Introduction: Kidney transplantation (KT) has surpassed dialysis as the optimal therapy for end-stage kidney disease. Yet, most patients could suffer from a slow but continuous deterioration of kidney function leading to graft loss mostly due to chronic allograft nephropathy (CAN) after KT. The dysregulated gene expression for CAN is still poorly understood. Methods: To explore the pathogenesis of genomics in CAN, we analyzed the differentially expressed genes (DEGs) of kidney transcriptome between CAN and nonrejecting patients by downloading gene expression microarrays from the Gene Expression Omnibus database. Then, we used weighted gene coexpression network analysis (WGCNA) to analyze the coexpression of DEGs to explore key modules, hub genes, and transcription factors in CAN. Functional enrichment analysis of key modules was performed to explore pathogenesis. ROC curve analysis was used to validate hub genes. Results: As a result, 3 key modules and 15 hub genes were identified by WGCNA analysis. Three key modules had 21 mutual Gene Ontology term enrichment functions. Extracellular structure organization, extracellular matrix organization, and extracellular region were identified as significant functions in CAN. Furthermore, transcription factor 12 was identified as the key transcription factor regulating key modules. All 15 hub genes, Yip1 interacting factor homolog B, membrane trafficking protein, toll like receptor 8, neutrophil cytosolic factor 4, glutathione peroxidase 8, mesenteric estrogen dependent adipogenesis, decorin, serpin family F member 1, integrin subunit beta like 1, SRY-box transcription factor 15, trophinin associated protein, SRY-box transcription factor 1, metallothionein 3, lysosomal protein transmembrane, FERM domain containing kindlin 3, and cathepsin S, had a great diagnostic performance (AUC > 0.7). Conclusion: This study updates information and provides a new perspective for understanding the pathogenesis of CAN by bioinformatics means. More research is needed to validate and explore the results we have found to reveal the mechanisms underlying CAN.

Impact of immunosuppressive agents on the expression of indoleamine 2,3-dioxygenase, heme oxygenase-1 and interleukin-7 in mesangial cells.

Chronic allograft nephropathy (CAN) is a major cause of graft loss following kidney transplantation and may result from the interactions of various immune and non-immune factors. The aim of the present study was to establish an in vitro model of glomerular mesangial cell injury in order to examine the gene expression levels of indoleamine 2,3-dioxygenase (IDO), heme oxygenase-1 (HO-1) and interleukin-7 (IL-7) in mesangial cells during the healing process as well as to investigate the effects of various immunosuppressants on the expression of these genes. The HBZY-1 glomerular mesangial cell line was pre-treated in vitro with cytochalasin B for 2 h to induce reversible damage. Following the pre-treatment, the HBZY-1 cells were divided into five groups: Blank control group, cyclosporine A (CsA) group, tacrolimus (Tac) group, mycophenolate mofetil (MMF) group and rapamycin (RAPA) group. After treating the mesangial cells with each immunosuppressive drug for 6, 12 or 24 h, the mRNA and protein expression levels of IDO, HO-1 and IL-7 were examined using reverse transcription quantitative polymerase chain reaction (RT-qPCR), western blot and immunohistochemical analyses. The results showed that expression levels of HO-1 were significantly upregulated in response to treatment with CsA, FK506, RAPA and MMF, whereas the expression levels of IL-7 were markedly downregulated by treatment with the above immunosuppressants. CsA, FK506 and MMF significantly enhanced the expression levels of IDO, whereas RAPA exhibited no apparent effect on IDO. The present study may contribute to the understanding of the pathogenesis of CAN and provide novel strategies for the prevention and treatment of CAN.

Increased Expression of p-Akt correlates with Chronic Allograft Nephropathy in a Rat Kidney Model.

Chronic allograft nephropathy (CAN) is the most common cause of chronic graft dysfunction leading to graft failure, our study investigates the expression and significance of p-Akt in the pathogenesis of CAN in rats. Kidneys of Fisher (F344) rats were orthotopically transplanted into Lewis (LEW) rats. The animals were evaluated at 4, 8, 12, 16, and 24 weeks post-transplantation for renal function and histopathology. Phosphorate Akt (p-Akt) protein expression was determined by Western blot and immunohistological assays. Our data show that 24-h urinary protein excretion in CAN rats increased significantly at week 16 as compared with F344/LEW controls. Allografts got severe interstitial infiltration of mononuclear cells at week 4 and week 8, but it was degraded as the time went on after week 16. Allografts markedly presented with severe interstitial fibrosis (IF) and tubular atrophy at 16 and 24 weeks. p-Akt expression was upregulated in rat kidneys with CAN, and the increase became more significant over time after transplantation. p-Akt expression correlated significantly with 24-h urinary protein excretion, serum creatinine levels, tubulointerstitial mononuclear cells infiltration, smooth muscle cells (SMCs) migration in vascular wall, and IF. It was concluded that p-Akt overexpression might be the key event that involved mononuclear cells infiltration and vascular SMCs migration at early stage, and IF and allograft nephroangiosclerosis at the late stage of CAN pathogenesis in rats.

Experimental rat models of chronic allograft nephropathy: a review.

Chronic allograft nephropathy (CAN) is the leading cause of late allograft loss after renal transplantation (RT), which continues to remain an unresolved problem. A rat model of CAN was first described in 1969 by White et al. Although the rat model of RT can be technically challenging, it is attractive because the pathogenesis of CAN is similar to that following human RT and the pathological features of CAN develop within months as compared with years in human RT. The rat model of RT is considered as a useful investigational tool in the field of experimental transplantation research. We have reviewed the literature on studies of rat RT reporting the donor and recipient strain combinations that have investigated resultant survival and histological outcomes. Several different combinations of inbred and outbred rat combinations have been reported to investigate the multiple aspects of transplantation, including acute rejection, cellular and humoral rejection mechanisms and their treatments, CAN, and potential targets for its prevention.

High Nerve Growth Factor Blood Concentration In Renal Transplantation: A New Prognostic Marker?

Nerve growth factor (NGF) belongs to the family of neurotropic proteins NGF is markedly expressed in proteinuric renal diseases and in end-stage renal disease; it might be involved in kidney physiopathology. To date, little is known about NGF concentrations in kidney transplant recipients (KTRs). Because NGF exerts its action on cell survival and differentiation, tissue repair, and inflammatory responses, it may also be implicated in the pathogenesis of chronic allograft nephropathy. The aim of this study was to determine circulating NGF concentrations in KTRs and to ascertain their use as a prognostic marker for kidney transplant outcomes. Using enzyme-linked immunosorbent assay, we performed quantification of NGF in the serum of 40 prevalent KTRs at baseline and at 6 months. NGF concentrations in KTRs averaged 1.16 ± 0.67 ng/mL. They negative-linearly correlated with recipient age. Logistic multivariate regression analysis showed NGF to be independently associated with increased proteinuria over the 6-month follow-up. Our data demonstrated that serum concentrations of NGF in KTRs were elevated and that they could be considered to be a prognostic marker in kidney transplantation.

Chronic Allograft Nephropathy - an Indian Study

Introduction: Chronic allograft nephropathy(CAN) is a major cause of late graft loss. Multiple factors have been implicated in the pathogenesis of CAN thus making the treatment of this entity equally difficult. Recent evidence has suggested involvement of Aldosterone in progression of Renal fibrosis. Hence this study was carried out for Aldactone (Spironolactone) as a novel form of therapy. Material and methods: A total of 72 patients during the study period from Aug 2004 to May 2011 with biopsy proven Chronic Allograft Nephropathy were randomized to receive Aldactone(Spironolactone) -(group A) & placebo-(group B) in addition to immunosuppressive & other medication. Group A received 25 mg of Aldactone. Investigations in the form of biochemical parameters including serum creatinine, proteinuria, GFR, C-reactive protein were studied in both the groups at the beginning and after 6 months and yearly thereafter. Follow up allograft biopsy could not be done in all the cases because of various reasons. The patients were followed up (minimum 1 year and maximum 6 years) on continued treatment from Aug 2004 to May2011. Results: Serum creatinine remained stable in Group A patients(p=0.020). GFR as measured with DTPA renal allograft scintigraphy showed a significant improvement (p=0.003) as compared to Group B patients. Changes in GFR correlated positively with decline in proteinuria & albuminuria (p=0.04) in Group A patients. The graft function progressively deteriorated in Group B patients (p=0.04). There was no significant hyperkalemia noted in thegroup A patients. There was no significant correlation of CRP levels with level of GFR(p=0.65), noted. The base line characterstics of both the groups were similar. Conclusions: The present study reveals benefit of using Aldactone (Spironolactone) in patients having Chronic Allograft Nephropathy resulting in significant improvement of graft function. It was well tolerated without any significant hyperkalemia.It is a novel, easily acceptable & cost effective form of therapy for CAN. This is the first Double blind, placebo controlled, randomized, prospective study using Aldactone for CAN from India & is a part of ongoing study.

Expression of the Integrin-Linked Kinase in a Rat Kidney Model of Chronic Allograft Nephropathy

The purpose of this study was to investigate the expression and significance of integrin-linked kinase (ILK) in the pathogenesis of chronic allograft nephropathy (CAN) in rats. For this, kidneys of Fisher (F344) rats were orthotopically transplanted into Lewis (LEW) rats. The animals were evaluated at 4, 8, 12, 16, and 24weeks post-transplantation for renal function and histopathology. ILK protein expression was determined by Western-blot and immunohistological assays, and mRNA by RT-PCR. Our data show that 24-h urinary protein excretion in CAN rats increased significantly at week 16 as compared with F344/LEW controls. Allografts showed markedly increased mononuclear cells infiltration and presented with severe interstitial fibrosis and tubular atrophy at 16 and 24weeks. ILK expression (protein/mRNA) was upregulated in rat kidneys with CAN, and the increase became more significant over time after transplantation. ILK expression correlated significantly with 24-h urinary protein excretion, serum creatinine levels, tubulointerstitial mononuclear cells infiltration, smooth muscle cells (SMCs) migration in vascular wall, and interstitial fibrosis. Therefore, it was concluded that ILK overexpression was the key event that involved mononuclear cells infiltration and vascular SMCs migration at early stage, and interstitial fibrosis and allograft nephroangiosclerosis at later stage of CAN pathogenesis in rats.

Early short-term imatinib treatment is sufficient to prevent the development of chronic allograft nephropathy

Chronic allograft nephropathy (CAN), now defined as interstitial fibrosis and tubular atrophy not otherwise specified, is a near universal finding in kidney grafts by the end of the first decade posttransplantation. Platelet-derived growth factor (PDGF) is a major mitogen mediating mesenchymal cell proliferation in CAN. Here, we investigated whether early short-term PDGF inhibition with imatinib could prevent CAN. Kidney transplantations were performed from Dark-Agouti (DA) to Wistar-Furth (WF) rats and syngenic controls were done between DA rats. Allografts were immunosuppressed with cyclosporine. One group was also treated with imatinib for the first 30 days after transplantation. Serum creatinine levels were measured once a week. Grafts were harvested 90 days after transplantation. In control allografts, moderate to intense chronic changes were seen, whereas in syngenic grafts, no changes were seen. The early imatinib treatment prevented the development of CAN significantly compared to control allografts. Only few histological changes were seen. Fibrogenic growth factor ligand and receptor induction as well as inflammatory cell response was significantly inhibited by imatinib. Creatinine values of imatinib-treated allografts were also significantly lower compared to controls. We show that short-term imatinib treatment is sufficient to prevent CAN significantly, indicating that early PDGF induction has an important role in the pathogenesis of CAN. Here, we provide preclinical work that will need to be confirmed in patients with CAN.

Insertion/Deletion Polymorphism of Angiotensin-Converting Enzyme as a Risk Factor for Chronic Allograft Nephropathy

Angiotensin-converting enzyme (ACE) inhibitor therapy is widely used to treat chronic allograft nephropathy (CAN), which suggests a possible role of the renin-angiotensin system in the pathologic mechanism of the disease. The objective of this study was to investigate the possible link between CAN and ACE. The ACE insertion/deletion polymorphism and the amount and activity of ACE were determined in cadaver kidney recipients with CAN (n = 38) or normal renal function (n = 34). The DD genotype was observed significantly more frequently in the CAN group compared with the group with normal renal function. Moreover, the DD genotype was associated with a higher serum ACE concentration and greater serum ACE activity, compared with II genotype homozygotes. The insertion/deletion polymorphism of ACE affects ACE expression and activity in serum, and, therefore, may have an important role in the pathogenesis of CAN. These findings suggest that determination of the ACE genotype may be useful in identifying patients at high risk. In particular, the DD genotype may be considered an indication for ACE inhibitor therapy.

The pathogenesis and treatment of chronic allograft nephropathy

Despite improvements in immunosuppressive therapy, long-term allograft survival after kidney transplantation remains as low as 50%. Chronic allograft nephropathy (CAN) is a major cause of late graft loss in renal transplant recipients. The histopathologic signs of CAN-interstitial fibrosis, tubular atrophy, glomerulopathy and vasculopathy-are nonspecific; therefore, the 2007 Banff classification dispensed with the term CAN in favor of 'interstitial fibrosis and tubular atrophy without evidence of any specific etiology'. In this Review, however, the term CAN is used to describe a clinical syndrome that is characterized by progressive decline in renal function from 3 months after transplantation, accompanied by the development of proteinuria and hypertension. The pathogenesis of CAN is complex and incompletely understood, and involves several immunological and non-immunological factors. We discuss the contributory roles of acute rejection, donor age, anti-human-leukocyte-antigen antibodies, calcineurin inhibitor nephrotoxic effects, viral infection, hypertension and hyperlipidemia. The prevention and treatment of CAN needs multidisciplinary strategies. Early detection by means of protocol biopsy and calculation of glomerular filtration rate is the first step, followed by management of modifiable risk factors.

Clinical study of the risk factors of insulin resistance and metabolic syndrome after kidney transplantation

To investigate the risk factors of insulin resistance (IR) and the role of IR and metabolic syndrome in the pathogenesis of chronic allograft nephropathy (CAN). One hundred and twenty-seven kidney transplant recipients with normal renal function and no proteinuria at the 6th month after transplantation, and without the experience of acute rejection, calcinurine intoxication and severe infection, were involved in the study. Their primary disease of ESRF was chronic glomerulonephritis but not diabetes mellitus and hypertension. Half year and one year after transplantation, blood and urine biochemical determinations and physical examination were performed in the recipients, and HOMA calculated. 200 ordinary community residents were randomized selected as controls. The incidence of MS in the recipients was significantly higher than controls. The incidences of obesity and overweight between recipients and controls were no significant difference. While the insulin resistance level and urine albumin level, and the incidence of MS and microalbuminuria (MAU) were significantly higher in recipients with obesity or overweight than that in recipients without obesity or overweight. The insulin resistance level in tacrolimus-treated recipients was markedly higher than CsA-treated recipients, and there was a positive correlation between the blood concentration of tacrolimus and insulin resistance level. MAU positive recipients had higher insulin resistance levels than the recipients without MAU. The recipients with metabolic syndrome had higher insulin resistance levels compared to recipients without metabolic syndrome, and higher insulin resistance levels existed in recipients with hypertriglyceridemia or hypercholesterolemia, hypertension. It is shown in the study that obesity or overweight, tacrolimus (especially when its blood concentration was high) were risk factors resulting in insulin resistance in kidney transplant recipients. It is suggested in the study that insulin resistance often accompanied with hypertriglyceridemia, hypercholesterolemia and hypertension in kidney transplant recipients might be involved in the pathogenesis of the pathogenesis of CAN.

Identification of platelet-derived growth factor D in human chronic allograft nephropathy

Chronic allograft nephropathy (CAN), a descriptive term denoting chronic scarring injury of the renal parenchyma and vasculature in allograft kidneys arising from various etiologies including chronic rejection, is the most common cause of late allograft failure, but mediators of this progressive injury largely remain unknown. We hypothesized that platelet-derived growth factor D (PDGF-D) and its specific receptor PDGF-Rbeta may be an important mediator in the pathogenesis of CAN and, hence, sought to identify its expression in this setting. Allograft nephrectomies demonstrating CAN, obtained from patients with irreversible transplant kidney failure (n = 15), were compared with renal tissues without prominent histopathological abnormalities (n = 18) and a series of renal allograft biopsies demonstrating acute vascular rejection (AVR) (n = 12). Antibodies to PDGF-D and PDGF-Rbeta were used for immunohistochemistry. Double and triple immunohistochemistry was used to identify cell types expressing PDGF-D. PDGF-D was widely expressed in most neointimas in arteries exhibiting the chronic arteriopathy of CAN and only weakly expressed in a small proportion of sclerotic arteries in the other 2 groups. Double and triple immunolabeling demonstrated that the neointimal cells expressing PDGF-D were alpha-smooth muscle actin-expressing cells, but not infiltrating macrophages or endothelial cells. PDGF-Rbeta expression evaluated in serial sections was localized to the same sites where neointimal PDGF-D was expressed. PDGF-Rbeta was expressed in interstitial cells more abundantly in the CAN group compared with the normal and AVR groups, without demonstrable colocalization of PDGF-D. PDGF-D is present in the neointima of the arteriopathy of CAN, where it can engage PDGF-Rbeta to promote mesenchymal cell migration, proliferation, and neointima formation. PDGF-D may engage the PDGF-Rbeta to promote interstitial injury in chronic allograft injury, but its sources within the interstitium were unidentified.

Epithelial-to-Mesenchymal Transition of Human Proximal Tubular Epithelial Cells: Effects of Rapamycin, Mycophenolate, Cyclosporin, Azathioprine, and Methylprednisolone

Epithelial-to-mesenchymal transition (EMT) of renal proximal tubular epithelial cells (PTEC) into myofibroblasts is an important step in the pathogenesis of chronic allograft nephropathy. The effects of commonly used immunosuppressives in renal transplantation on EMT are not known. PTEC were cultured in transforming growth factor-beta to induce EMT. The effects of the immunosuppressives on cell morphology and alpha-smooth muscle actin were studied by phase contrast microscopy, immunocytochemistry, and western blotting. The effects on versican were studied by [S] labeling and polyacrylamide gel electrophoresis. Rapamycin and mycophenolate mofetil (MMF) prevented EMT and moreover returned myofibroblasts to PTEC morphology. These immunosuppressives also reduced versican production by both PTEC and myofibroblasts. Cyclosporine A, azathioprine, and methylprednisolone were less effective than rapamycin and MMF. Moreover, these immunosuppressives did not decrease versican. Rapamycin and MMF have a greater inhibitory effect on EMT in vitro than older immunosuppressives and may result in less fibrosis and a better long-term allograft survival.

Mechanisms of ageing in chronic allograft nephropathy

Background. Chronic allograft nephropathy (CAN) is the single most important cause of late graft loss. Histologic signs of CAN overlap with the histology of normally ageing kidneys. Clinical observations show an inverse relationship between either donor age or ischemia time and long-term graft survival. The mechanisms by which donor age or ischemia lead to reduced functional lifespan are currently unknown. Material and methods. A review of the current literature on renal transplantation, ageing mechanisms and advances in the molecular pathogenesis of CAN was performed. Results and conclusions. Ischemia time and donor age have been identified as major allo-antigen-independent risk factors. Following transplantation, reactive oxygen species cause oxidative DNA and telomeric damage. Evidence for a molecular connection between oxidative DNA damage and ageing has increased, and suggests that the mechanisms causing normal ageing and long-term organ dysfunction may be fundamentally the same. The accelerated appe...

Early Short-Term Platelet-Derived Growth Factor Inhibition Prevents the Development of Chronic Allograft Nephropathy in Experimental Rat Kidney Transplantation

Chronic allograft nephropathy (CAN) remains the primary reason for late allograft loss in kidney transplantation. Platelet-derived growth factor (PDGF) is a major mitogen mediating mesenchymal cell proliferation in CAN. When administered continuously the PDGF receptor tyrosine kinase inhibitor imatinib prevents the development of CAN and restores kidney function in experimental kidney transplantation. Herein we investigated whether early short-term imatinib treatment prevented CAN. Kidney transplantations were performed from DA to WF rats and syngenic controls were done between DA rats. Allograft recipients were immunosuppressed with cyclosporine (CsA; 1.5 mg/kg/d sc). One group of allografts was also treated with imatinib (10 mg/kg/d po). Serum creatinine levels were measured once a week. Grafts were harvested 90 days after transplantation for histology and immunohistochemistry (PDGF-AA, -BB, PDGFR-alpha, -beta). Histological changes were scored according to the Chronic Allograft Damage Index (CADI). Among syngenic grafts, no signs of CAN were observed, namely, CADI 0.3 +/- 0.2 (mean +/- SEM). Control allografts showed moderate to intense chronic changes, CADI 6.5 +/- 1.3. Early short-term imatinib treatment significantly prevented the development of CAN compared with control allografts. Only a few histological changes were seen, namely, CADI 3.3 +/- 1.4. Compared with control allografts PDGF ligand and receptor induction was significantly inhibited by imatinib to nearly the same level as in syngenic grafts. Creatinine values of imatinib-treated allografts were also lower than control allografts. Our results demonstrated that early short-term imatinib treatment significantly prevented CAN. This indicated that early PDGF induction has an important role in the pathogenesis of CAN.

Pioglitazone limits cyclosporine nephrotoxicity in rats

Chronic cyclosporine (CsA) nephrotoxicity is a relevant factor in the pathogenesis of chronic allograft nephropathy. Pioglitazone is an agonist of PPARgamma, capable of reducing chronic inflammation. We investigated the capacity of pioglitazone in preventing renal dysfunction. Adult male Wistar rats were assigned to: Vehicle (olive oil 1 ml/kg/day), CsA (10 mg/kg/day) alone and with pioglitazone (5 or 10 mg/kg/day). The animals were sacrificed at 28 days, where blood (serum creatinine ratio, CR) and kidney samples (arteriolopathy analyses) were collected. The mRNA transcripts of TGF-beta1, PAI-1, Smad3 and 7 were evaluated by real-time PCR. As expected, CsA treatment significantly decreased renal function that peaked at day 28, compared with vehicle (CR=1.29+/-0.03 vs. 0.95+/-0.14, p<0.05). In contrast, the administration of pioglitazone 5 or 10 mg/kg combined with CsA resulted in better renal function (CR=1.09+/-0.05 and 1.14+/-0.14, respectively, p<0.05). Animals treated with CSA showed relevant arteriolopathy (49.5+/-2.86%) and pioglitazone administration significantly limited it (37.0+/-3.59% and 36.6+/-1.72%, respectively, 5 or 10 mg/kg, p<0.05). In CsA-treated animals (alone and with pioglitazone), TGF-beta1 and Smad3 increased significantly. In animals treated with CsA and pioglitazone (5 mg/kg), PAI-1 was significantly lower than CsA alone (3.96+/-0.92 vs. 7.53+/-1.38, p<0.05). Interestingly, the administration of pioglitazone 5 or 10 mg/kg was associated with an increase in Smad7 (1.79+/-0.25 and 1.75+/-0.19, respectively), compared to vehicle and to CsA-treated groups (1.08+/-0.17 and 1.17+/-0.19, respectively, p<0.05). These data provide evidence that pioglitazone acts through down regulation of pro-fibrotic cytokine PAI-1 and overexpression of the regulatory Smad7.

The expression of monocyte chemoattractant protein-1 and C–C chemokine receptor 2 in post-kidney transplant patients and the influence of simvastatin treatment

Inflammation is involved in the pathogenesis of chronic allograft nephropathy (CAN) and cardiovascular disease (CVD) in post-kidney transplantation. Sixty patients, selected from 167 renal transplant patients, were divided into 2 groups: Group A (normal lipidemia group, n=30) and Group B (hyperlipidemia group, n=30). In addition, Control Group came from 30 healthy volunteers. In Group B, the patients were treated with simvastatin for 3 months. The mRNA expressions of MCP-1 and CCR2 were detected with reverse transcription-polymerase chain reaction (RT-PCR). The mRNA expressions of MCP-1 and CCR2 of all post-kidney transplant patients were significantly higher than controls. Compare Group A with Group B and the mRNA expressions of MCP-1 and CCR2 in Group B were much higher than Group A. After simvastatin treatment, the mRNA expressions of MCP-1 and CCR2 were significantly reduced in one and a half months and decreased to the lowest levels in three months. Simvastatin decreased the expressions of MCP-1 and CCR2 in post-kidney transplant patients with hyperlipidemia.

TGF-β1 mRNA upregulation influences chronic renal allograft dysfunction

Acute rejection (AR) is a dominant risk factor for developing chronic allograft nephropathy (CAN) after kidney transplantation. CAN is characterized by progressive interstitial fibrosis. It has been associated with increased transforming growth factor (TGF)-beta1 expression, however, kinetic studies are absent. We investigated whether intragraft TGF-beta1 expression in various causes of early graft dysfunction may influence late renal allograft dysfunction. A total of 174 human renal biopsies were quantified for TGF-beta1 mRNA expression using real-time reverse transcriptase-polymerase chain reaction. Expression levels were correlated with the Banff histopathological grades, TGF-beta1 immunohistology, and clinical follow-up. TGF-beta1 was most markedly upregulated in AR, CAN, and acute tubular necrosis - delayed graft function compared to non-rejecting controls (P < 0.001). TGF-beta1 expression was heightened in borderline changes (P < 0.01), recurrence of glomerulonephritis, and cyclosporine toxicity (P < 0.05). There was no correlation between intragraft TGF-beta1 expression during AR and short-term outcome of a rejection episode. TGF-beta1 gene overexpression during CAN has been shown to be associated with the increased risk for renal allograft dysfunction 18 months after biopsy (odds ratios 9.9 vs 3.2, respectively). Intragraft TGF-beta1 mRNA expression is significantly upregulated in both AR and CAN. Thus, our results support the hypothesis that TGF-beta1 might play a key role in chronic allograft dysfunction.

CD40L Proinflammatory and Profibrotic Effects on Proximal Tubular Epithelial Cells

Chronic allograft nephropathy (CAN) is the main cause of renal graft loss, but its pathogenic mechanisms are still unclear. Immune system activation has been suggested as a key event in the development of CAN. CD40 is a co-stimulatory protein whose expression is upregulated in proximal tubular epithelial cells (PTEC) in acute rejection. This receptor interacts with CD40L, expressed by activated T cells. CD40L induces the production by PTEC of different proinflammatory cytokines, but very little is known of its profibrotic effects. The aim of this study was to investigate the effect of CD40/CD40L interaction on PTEC expression of plasminogen activator inhibitor-1 (PAI-1), a powerful profibrotic mediator, and monocyte chemoattractant protein-1 (MCP-1), a proinflammatory cytokine, and to investigate the signaling pathways that lead to these effects. Soluble CD40L induced a time-dependent increase in both PAI-1 and MCP-1 gene expression and protein production in PTEC. CD40 cross-linking on PTEC caused TNF-R-associated factors 2 and 6 membrane translocation. This event led to NF-kappaB activation, through the NF-kappaB-inducing kinase, and to a significant increase in the phosphorylation of lyn, a src-related tyrosine kinase. Lyn, upon phosphorylation, became strictly associated with caveolin-1, a scaffolding protein enriched in caveolae. Lyn inhibition did not have any effect on CD40L-induced NF-kappaB activation and MCP-1 expression but abolished PAI-1 induction. On the contrary, NF-kappaB inhibition significantly reduced only MCP-1 expression. In conclusion, CD40L could play a key role in the pathogenesis of CAN through PAI-1 induction. CD40L profibrotic and proinflammatory effects are mediated by different signaling pathways, suggesting that drugs that inhibit inflammation may not be equally effective in reducing fibrosis.

Profibrotic cytokines and lymphocyte proliferation in stable renal allograft recipients treated with or without cyclosporine A

Profibrotic cytokines such as transforming growth factor-beta 1 (TGF-beta1) and endothelin-1 (ET-1) are involved in the pathogenesis of chronic allograft nephropathy. We assessed the effect of maintenance immunosuppression with or without cyclosporine A on TGF-beta1 and ET-1 expression as well as lymphocyte proliferation in renal allograft recipients. Patients were divided into groups according to their maintenance immunosuppression: A, azathioprine + methylprednisolone; B, cyclosporine A + azathioprine + methylprednisolone. TGF-beta1 and ET-1 plasma concentrations were not different between both groups. TGF-beta1 concentrations in cell culture supernatants and lymphocyte proliferation were higher in group B as compared to group A. No correlation was found between TGF-beta1 plasma concentrations and lymphocyte proliferation. Multiple linear regression analysis revealed that patient characteristics influence TGF-beta1 and ET-1 expression. In conclusion, plasma levels of profibrotic cytokines do not reflect the existing profibrotic potential of immunosuppressive drugs. Demographic factors and the employed co-medication confound the results.