Protective effects of human amniotic fluid stem cells in a model of aorta allograft vasculopathy in rats.

BACKGROUND: Chronic allograft vasculopathy (CAV) is an important cause of graft loss. Considering the immune inflammatory events involved in the development of CAV, therapeutic approaches to target this process are of relevance. Human amniotic fluid-derived stem cells (hAFSCs), a class of fetal, pluripotent stem cells with intermediate characteristics between embryonic and adult stem cells, display immunomodulatory properties. hAFSCs express mesenchymal and embryonic markers, show high proliferation rates; however, they do not induce tumor formation, and their use does not raise ethical issues. Thus, we sought to investigate the effect of hAFSC on CAV in a model of aorta transplantation. METHODS: Orthotopic aorta transplantation was performed using Fisher (F344) rats as donors and Lewis rats as recipients. Rats were divided into three groups: syngeneic (SYNG), untreated F344 receiving aorta from F344 (n = 8); allogeneic (ALLO), Lewis rats receiving allogeneic aorta from F344 (n = 8); and ALLO + hAFSC, ALLO rats treated with hAFSC (10(6) cells; n = 8). Histological analysis and immunohistochemistry were performed 30 days posttransplantation. RESULTS: The ALLO group developed a robust aortic neointimal formation (208.7 ± 25.4 µm) accompanied by a significant high number of ED1+ (4845 ± 841 cells/mm2) and CD43+ cells (4064 ± 563 cells/mm2), and enhanced expression of α-smooth muscle actin in the neointima (25 ± 6%). Treatment with hAFSC diminished neointimal thickness (180.7 ± 23.7 µm) and induced a significant decrease of ED1+ (1100 ± 276 cells/mm2), CD43+ cells (1080 ± 309 cells/µm2), and α-smooth muscle actin expression 8 ± 3% in the neointima. CONCLUSIONS: These preliminary results showed that hAFSC suppressed inflammation and myofibroblast migration to the intima, which may contribute to ameliorate vascular changes in CAV.

Induction of indoleamine 2,3-dioxygenase by gene delivery in allogeneic islets prolongs allograft survival.

Indoleamine 2,3-dioxygenase (IDO), an enzyme that plays a critical role in fetomaternal tolerance, exerts immunoregulatory functions suppressing T-cell responses. The aims of this study were to promote IDO expression in rat islets using a nonviral gene transfer approach, and to analyze the effect of the in vivo induction of IDO in a model of allogeneic islet transplantation. The IDO cDNA was isolated from rat placenta, subcloned into a plasmid and transfected into rat islets using Lipofectamine. The efficiency of transfection was confirmed by qRT-PCR and functional analysis. The in vivo effect of IDO expression was analyzed in streptozotocin-induced diabetic Lewis rats transplanted with allogeneic islets under the renal capsule. Transplantation of IDO-allogeneic islets reversed diabetes and maintained metabolic control, in contrast to transplantation of allogeneic nontransfected islets, which failed shortly after transplantation in all animals. Graft survival of allograft islets transfected with IDO transplanted without any immunosuppression was superior to that observed in diabetic rats receiving nontransfected islets. These data demonstrated that IDO expression induced in islets by lipofection improved metabolic control of streptozotocin-diabetic rats and prolonged allograft survival.

Effectiveness of thalidomide and tamoxifen in preventing neointimal hyperplasia in experimental vascular injury in rats.

INTRODUCTION: Chronic allograft vasculopathy is an important cause of graft loss. Considering the inflammatory response in the development of chronic vascular lesions, therapeutic approaches to target the inflammatory process may be useful. We sought to investigate the possible protective effects on balloon catheter-induced vascular injury of thalidomide and tamoxifen, 2 drugs with powerful anti-inflammatory, immunomodulatory, and antifibrotic effects, using an animal model that mimics the morphologic features of chronic allograft vasculopathy. METHODS: Male Wistar rats subjected to balloon catheter carotid injury (INJ) were treated with thalidomide (100 mg/kg), or tamoxifen (10 mg/kg), or vehicle. Contralateral right carotid arteries were used as uninjured controls. Morphometric and immunohistochemical analyses were performed at 14 days postinjury. RESULTS: Injured carotid arteries showed marked neointimal hyperplasia, which was significantly inhibited among animals treated with thalidomide or tamoxifen: neointimal/media ratios of 1.4 +/- 0.4 versus 0.2 +/- 0.1 versus 0.4 +/- 0.2, for INJ, INJ + Thalid, and INJ + Tamox; respectively (P < .001). The endothelial cell loss was significantly less pronounced among animals subjected to carotid balloon injury that were treated with thalidomide (24 +/- 14 vs 1 +/- 1 cells per section in INJ, respectively (P < .05). Therapy with either thalidomide or tamoxifen effectively maintained alpha-smooth muscle actin expression in the media, similar to uninjured arteries. In this setting, tamoxifen was additionally effective to prevent the migration of myofibroblasts in to the intima. CONCLUSION: Thalidomide and tamoxifen were effective to reduce neointimal hyperplasia secondary to vascular damage. The vasculoprotective effects of thalidomide were more pronounced to preserve endothelial cells, whereas tamoxifen inhibited smooth muscle cell migration and proliferation. A possible beneficial effect of combined therapy with thalidomide plus tamoxifen should be addressed in future studies.

A feasibility study of a web-based performance improvement system for substance abuse treatment providers.

We report here on the feasibility of implementing a semiautomated performance improvement system-Patient Feedback (PF)-that enables real-time monitoring of patient ratings of therapeutic alliance, treatment satisfaction, and drug/alcohol use in outpatient substance abuse treatment clinics. The study was conducted in six clinics within the National Institute on Drug Abuse Clinical Trials Network. It involved a total of 39 clinicians and 6 clinic supervisors. Throughout the course of the study (consisting of five phases: training period [4 weeks], baseline [4 weeks], intervention [12 weeks], postintervention assessment [4 weeks], sustainability [1 year]), there was an overall collection rate of 75.5% of the clinic patient census. In general, the clinicians in these clinics had very positive treatment satisfaction and alliance ratings throughout the study. However, one clinic had worse drug use scores at baseline than other participating clinics and showed a decrease in self-reported drug use at postintervention. Although the implementation of the PF system proved to be feasible in actual clinical settings, further modifications of the PF system are needed to enhance any potential clinical usefulness.

The use of iodixanol for the purification of rat pancreatic islets.

Transplantation of pancreatic islets is a promising therapeutic treatment for type 1 diabetes mellitus. For clinical and experimental transplantation, a large number of pure pancreatic islets are required for transplantation. Thus, the improvement of islet isolation and purification techniques are crucial. In this context, iodixanol-based solution, successfully used for the purification of porcine islets, seems to be a possible alternative to Ficoll for purification of islets. The aim of this study was to test the efficacy of iodixanol compared with Ficoll density gradients for the purification of rat pancreatic islets. Twelve Wistar rats were used for isolation and purification of pancreatic islets. Pancreata were digested with Liberase R1 and islets purified by two gradients: Ficoll or iodixanol gradient. The number and the purity of the pancreatic islets were assessed. To analyze the response of isolated pancreatic islet to glucose challenge, in vitro experiments were performed by measuring the insulin concentration in the Supernatant. The results demonstrated that the iodixanol gradient provided a higher purity of pancreatic islets compared to the Ficoll gradient. In addition, the rat islet yield by iodixanol gradient was significantly higher compared to a Ficoll gradient (751 +/- 16 versus 464 +/- 19 pancreatic islets, respectively; P < .001). The viability of pancreatic islets isolated by an iodixanol gradient was confirmed by high glucose challenge, with more than twofold higher increase in insulin secretion. The present study demonstrated that iodixanol density gradient overcomes Ficoll density gradient, providing a greater number of pure and functional rat pancreatic islets.

Statin monotherapy attenuates renal injury in a salt-sensitive hypertension model of renal disease.

BACKGROUND: Several salutary biological effects of statins have been described. We sought to investigate more closely the anti-inflammatory and antiproliferative effects of simvastatin (SIMV) in a model of hypertension and progressive renal disease, as well as its effects on the cyclin-cdk inhibitors p21 and p27. METHODS: Munich-Wistar rats received the nitric oxide (NO) synthase inhibitor L-NAME (25 mg/kg/day p.o.) for 20 days accompanied by a high-salt diet (HS, 3% Na) and then were kept on HS for 60 days. Animals were then divided into two groups: vehicle (VH) or SIMV 2 mg/kg/day p.o. Albuminuria and tail-cuff pressure were determined at 30 and 60 days. RT-PCR was done to assess renal expression of TGF-beta1, collagen I and III, fibronectin, p27, p21 and monocyte chemoattractant protein-1 (MCP-1). Renal protein expression was assessed by Western blot (proliferating cell nuclear antigen (PCNA)) and immunostaining (macrophage, lymphocyte, PCNA). RESULTS: SIMV did not prevent the development of severe hypertension or albuminuria. SIMV-treated animals had less severe renal interstitial inflammation and cell proliferation. MCP-1 expression was significantly diminished in the SIMV-treated animals (55.4 +/- 7.3 vs. 84.4 +/- 8.2 OD, p = 0.02). mRNA renal expression for p27 and TGF-beta did not change between groups, but p21 mRNA renal expression, highly induced in this model, significantly decreased with SIMV treatment (31.6 +/- 6.6 vs. 50.2 +/- 5.8 OD, p < 0.05). The interstitial fibrosis score significantly decreased with SIMV (2.46 +/- 0.40 vs. 4.07 +/- 0.38%, p < 0.01), which was confirmed by a decrease in renal collagen I and fibronectin expression. Serum cholesterol level did not change with SIMV. CONCLUSION: SIMV attenuated interstitial fibrosis associated with this model of hypertensive renal disease. The mechanism involved MCP-1 downregulation. SIMV treatment was also associated with a p21 downregulation in the kidney, which might be involved in the protection of renal scarring.