Long-Term Type 1 Diabetes Enhances In-Stent Restenosis after Aortic Stenting in Diabetes-Prone BB Rats

Type 1 diabetic patients have increased risk of developing in-stent restenosis following endovascular stenting. Underlying pathogenetic mechanisms are not fully understood partly due to the lack of a relevant animal model to study the effect(s) of long-term autoimmune diabetes on development of in-stent restenosis. We here describe the development of in-stent restenosis in long-term (~7 months) spontaneously diabetic and age-matched, thymectomized, nondiabetic Diabetes Prone BioBreeding (BBDP) rats (n = 6-7 in each group). Diabetes was suboptimally treated with insulin and was characterized by significant hyperglycaemia, polyuria, proteinuria, and increased HbA1c levels. Stented abdominal aortas were harvested 28 days after stenting. Computerized morphometric analysis revealed significantly increased neointima formation in long-term diabetic rats compared with nondiabetic controls. In conclusion, long-term autoimmune diabetes in BBDP rats enhances in-stent restenosis. This model can be used to study the underlying pathogenetic mechanisms of diabetes-enhanced in-stent restenosis as well as to test new therapeutic modalities

Spironolactone ameliorates transplant vasculopathy in renal chronic transplant dysfunction in rats

Waanders F, Rienstra H, Walther Boer M, Zandvoort A, Rozing J, Navis G, van Goor H, Hillebrands JL. Spironolactone ameliorates transplant vasculopathy in renal chronic transplant dysfunction in rats. Am J Physiol Renal Physiol 296: F1072-F1079, 2009. First published February 25, 2009; doi:10.1152/ajprenal.90643.2008.-Chronic transplant dysfunction (CTD) is the leading cause of long-term renal allograft loss and is characterized by specific histological lesions including transplant vasculopathy, interstitial fibrosis, and focal glomerulosclerosis. Increasing evidence indicates that aldosterone is a direct mediator of renal damage via the mineralocorticoid receptor (MR). The MR antagonist spironolactone is renoprotective in native chronic kidney disease, but its effects on CTD are unknown. We studied the effects of spironolactone treatment on CTD development in the Dark Agouti-to-Wistar-Furth renal allograft transplant model, by treatment with 20 mg/kg spironolactone or vehicle daily by oral gavage from 2 days before transplantation (donors and recipients) throughout the experiment (12 wk, recipients). Dark Agouti-to-Dark Agouti isografts served as negative controls. Spironolactone significantly ameliorated the development of transplant vasculopathy in allografts by reducing the number of affected intrarenal arteries. In addition, spironolactone treatment showed a trend toward reduced proteinuria and focal glomerulosclerosis, and significantly reduced glomerular macrophage influx. However, spironolactone treatment did not affect interstitial fibrosis, interstitial macrophage influx, creatinine clearance, and systolic blood pressure. We conclude that spironolactone selectively ameliorates transplant vasculopathy and glomerular lesions in renal CTD in rats. These results suggest that spironolactone may have renoprotective potential as an adjunct treatment in renal transplantation to ameliorate CTD

Rosiglitazone attenuates transplant arteriosclerosis after allogeneic aorta transplantation in rats

BACKGROUND. Transplant arteriosclerosis is a leading cause of chronic transplant dysfunction and is characterized by occlusive neointima formation in intragraft arteries. Development of transplant arteriosclerosis is refractory to conventional immunosuppressive drugs and adequate therapy is not available. In this study, we determined the efficacy of the synthetic peroxisome proliferator-activated receptor (PPAR)-γ agonist rosiglitazone to attenuate the development of transplant arteriosclerosis in rat aortic allografts. METHODS. Lewis aortic allografts were transplanted into Brown Norway recipient rats. Recipient rats received either ∼5 mg rosiglitazone/day (starting 1 week before transplantation until the end of the experiment) or were left untreated. Transplant arteriosclerosis was quantified using morphometric analysis. Alloreactivity was measured in vitro using mixed lymphocyte reactions. Regulatory T cell frequency and function were analyzed using flow cytometry and in vitro suppression assays, respectively. Intragraft gene expression was analyzed using real-time polymerase chain reaction. Finally, medial and neointimal vascular smooth muscle cell proliferation was analyzed in vitro. RESULTS. Rosiglitazone significantly reduced transplant arteriosclerosis development 8 weeks after transplantation (P&lt;0.01 vs. nontreated). Rosiglitazone reduced T cell alloreactivity which was not mediated through modulation of CD4CD25FoxP3 regulatory T cells. Reduced development of transplant arteriosclerosis coincided with reduced intragraft expression of stromal-derived factor-1α and platelet-derived growth factor receptor-β. Finally, rosiglitazone reduced growth-factor-driven proliferation of both medial and neointimal vascular smooth muscle cells in vitro, which was not mediated through PPARγ. CONCLUSION. PPARγ agonists may offer a new therapeutic strategy in clinical transplantation to attenuate the development of transplant arteriosclerosis and thereby chronic transplant dysfunction.</p

Rosiglitazone attenuates transplant arteriosclerosis after allogeneic aorta transplantation in rats

BACKGROUND. Transplant arteriosclerosis is a leading cause of chronic transplant dysfunction and is characterized by occlusive neointima formation in intragraft arteries. Development of transplant arteriosclerosis is refractory to conventional immunosuppressive drugs and adequate therapy is not available. In this study, we determined the efficacy of the synthetic peroxisome proliferator-activated receptor (PPAR)-γ agonist rosiglitazone to attenuate the development of transplant arteriosclerosis in rat aortic allografts. METHODS. Lewis aortic allografts were transplanted into Brown Norway recipient rats. Recipient rats received either ∼5 mg rosiglitazone/day (starting 1 week before transplantation until the end of the experiment) or were left untreated. Transplant arteriosclerosis was quantified using morphometric analysis. Alloreactivity was measured in vitro using mixed lymphocyte reactions. Regulatory T cell frequency and function were analyzed using flow cytometry and in vitro suppression assays, respectively. Intragraft gene expression was analyzed using real-time polymerase chain reaction. Finally, medial and neointimal vascular smooth muscle cell proliferation was analyzed in vitro. RESULTS. Rosiglitazone significantly reduced transplant arteriosclerosis development 8 weeks after transplantation (P&lt;0.01 vs. nontreated). Rosiglitazone reduced T cell alloreactivity which was not mediated through modulation of CD4CD25FoxP3 regulatory T cells. Reduced development of transplant arteriosclerosis coincided with reduced intragraft expression of stromal-derived factor-1α and platelet-derived growth factor receptor-β. Finally, rosiglitazone reduced growth-factor-driven proliferation of both medial and neointimal vascular smooth muscle cells in vitro, which was not mediated through PPARγ. CONCLUSION. PPARγ agonists may offer a new therapeutic strategy in clinical transplantation to attenuate the development of transplant arteriosclerosis and thereby chronic transplant dysfunction.</p

Dichotomous effects of rosiglitazone in transplantation-induced systemic vasodilator dysfunction in rats

Background. Transplantation-induced systemic endothelial dysfunction causes severe cardiovascular morbidity and mortality after transplantation. Interventions that improve systemic endothelial function after transplantation and furthermore reduce intragraft vascular dysfunction might improve graft and patient survival. Treatment with the PPAR gamma agonist rosiglitazone is an intervention that potentially fulfills these criteria. In this study, we determined the effect of rosiglitazone treatment on transplantation-induced endothelial dysfunction and vasomotor activity in an experimental model for chronic transplant dysfunction in rats. Methods. Lewis abdominal aortic allografts were orthotopically transplanted into Brown Norway recipients that received either regular chow or chow containing rosiglitazone (similar to 4.2 mg/day). Endothelium-dependent (response to metacholine) and total (response to sodium nitrite) vasodilatory responses were determined in autologous thoracic aortic rings using an ex vivo organ bath setup. Measurements were performed 8 weeks after transplantation. Results. Aortic allografting induced systemic endothelial dysfunction as measured by reduced endothelium-dependent vasodilation in the recipient's vascular system. Rosiglitazone treatment restored endothelium-dependent vasodilatory responses to pretransplantation levels. However, rosiglitazone treatment reduced the total dilatory response despite normalized endothelial function, indicating impairment of vascular smooth muscle cell vasomotor activity. Conclusions. Rosiglitazone treatment after allogeneic transplantation restores endothelial function but impairs vascular smooth muscle cell vasomotor activity. This dichotomous effect of rosiglitazone might impede use of rosiglitazone after organ transplantation since this potentially increases cardiovascular risk despite improved endothelial cell function

Development of transplant vasculopathy in aortic allografts correlates with neointimal smooth muscle cell proliferative capacity and fibrocyte frequency

Objective: Transplant vasculopathy consists of neointima formation in graft vasculature resulting from vascular smooth muscle cell recruitment and proliferation. Variation in the severity of vasculopathy has been demonstrated. Genetic predisposition is suggested as a putative cause of this variation, although cellular mechanisms are still unknown. Using a rat aorta transplant model we tested the hypothesis that kinetics of development of transplant vasculopathy are related to neointimal smooth muscle cell proliferative capacity and fibrocyte frequency, the latter being putative neointimal smooth muscle ancestral cells. Methods: Aortic allografts were transplanted in Lewis and Brown Norway, as well as MHC-congenic Lewis. 1N and Brown Norway. 1L recipients. Severity of transplant vasculopathy was quantified 4, 8, 12 and 24 weeks after transplantation. Host-endothelial chimerism, as a reflection of vascular injury, was determined by specific immunofluorescence. Neointimal smooth muscle cell proliferative capacity was determined in vitro and in situ. Fibrocyte frequency and phenotype were determined after in vitro culture by cell counting, immunofluorescence and in situ zymography. Results: Compared to Lewis, Brown Norway recipients developed accelerated transplant vasculopathy which is dependent on the presence of Brown Norway non-MHC-encoded determinants. Accelerated transplant vasculopathy was associated with increased levels of host-endothelial chimerism and increased neointimal smooth muscle cell proliferation, the latter being accompanied by increased endothelial and smooth muscle cell-derived neuropilin-like protein mRNA expression. Moreover, accelerated transplant vasculopathy was associated with increased frequency of circulating gelatinase-expressing CD45(+) vimentin(+) fibrocytes. Conclusion: Susceptibility for transplant vasculopathy appears to be genetically controlled and correlates with neointimal smooth muscle cell proliferative capacity and circulating fibrocyte frequency. (C) 2009 Elsevier Ireland Ltd. All rights reserved

Rosiglitazone attenuates transplant arteriosclerosis after allogeneic aorta transplantation in rats

Background. Transplant arteriosclerosis is a leading cause of chronic transplant dysfunction and is characterized by occlusive neointima formation in intragraft arteries. Development of transplant arteriosclerosis is refractory to conventional immunosuppressive drugs and adequate therapy is not available. In this study, we determined the efficacy of the synthetic peroxisome proliferator-activated receptor (PPAR)-gamma agonist rosiglitazone to attenuate the development of transplant arteriosclerosis in rat aortic allografts. Methods. Lewis aortic allografts were transplanted into Brown Norway recipient rats. Recipient rats received either similar to 5 mg rosiglitazone/day (starting 1 week before transplantation until the end of the experiment) or were left untreated. Transplant arteriosclerosis was quantified using morphometric analysis. Alloreactivity was measured in vitro using mixed lymphocyte reactions. Regulatory T cell frequency and function were analyzed using flow cytometry and in vitro suppression assays, respectively. Intragraft gene expression was analyzed using real-time polymerase chain reaction. Finally, medial and neointimal vascular smooth muscle cell proliferation was analyzed in vitro. Results. Rosiglitazone significantly reduced transplant arteriosclerosis development 8 weeks after transplantation (P <0.01 vs. nontreated). Rosiglitazone reduced T cell alloreactivity which was not mediated through modulation of CD4(+)CD25(+)FoxP3(+) regulatory T cells. Reduced development of transplant arteriosclerosis coincided with reduced intragraft expression of stromal-derived factor-1 alpha and platelet-derived growth factor receptor-beta. Finally, rosiglitazone reduced growth-factor-driven proliferation of both medial and neointimal vascular smooth muscle cells in vitro, which was not mediated through PPAR-gamma. Conclusion. PPAR gamma agonists may offer a new therapeutic strategy in clinical transplantation to attenuate the development of transplant arteriosclerosis and thereby chronic transplant dysfunction

Clonal diversity predicts adverse outcome in chronic lymphocytic leukemia

Genomic analyses of chronic lymphocytic leukemia (CLL) identified somatic mutations and associations of clonal diversity with adverse outcomes. Clonal evolution likely has therapeutic implications but its dynamic is less well studied. We studied clonal composition and prognostic value of seven recurrently mutated driver genes using targeted next-generation sequencing in 643 CLL patients and found higher frequencies of mutations in TP53 (35 vs. 12%, p < 0.001) and SF3B1 (20 vs. 11%, p < 0.05) and increased number of (sub)clonal (p < 0.0001) mutations in treated patients. We next performed an in-depth evaluation of clonal evolution on untreated CLL patients (50 “progressors” and 17 matched “non-progressors”) using a 404 gene-sequencing panel and identified novel mutated genes such as AXIN1, SDHA, SUZ12, and FOXO3. Progressors carried more mutations at initial presentation (2.5 vs. 1, p < 0.0001). Mutations in specific genes were associated with increased (SF3B1, ATM, and FBXW7) or decreased progression risk (AXIN1 and MYD88). Mutations affecting specific signaling pathways, such as Notch and MAP kinase pathway were enriched in progressive relative to non-progressive patients. These data extend earlier findings that specific genomic alterations and diversity of subclones are associated with disease progression and persistence of disease in CLL and identify novel recurrently mutated genes and associated outcomes