Role of Sphingosine-1-Phosphate in Transplant Vasculopathy Evoked by Anti-HLA Antibody.

Transplant vasculopathy (TV) represents the main cause of late graft failure and limits the long-term success of organ transplantation. Cellular and humoral immune responses contribute to the pathogenesis of the concentric and diffuse intimal hyperplasia of arteries of the grafted organ. We recently reported that the mitogenic signaling, evoked in human vascular smooth muscle cells (hmSMC) by the anti-HLA class I monoclonal antibody W6/32, implicates neutral sphingomyelinase-2, suggesting a role for sphingolipids in intimal hyperplasia of TV. Here, we investigated whether the mitogenic sphingolipid, sphingosine-1-phosphate (S1P), is involved in intimal hyperplasia elicited by W6/32. Studies were done on cultured hmSMC and on an in vivo model of TV, consisting of human mesenteric arteries grafted into SCID/beige mice, injected weekly with W6/32. hmSMC migration and DNA synthesis elicited by W6/32 were inhibited by the sphingosine kinase-1 (SK1) inhibitor dimethylsphingosine, the anti-S1P antibody Sphingomab and the S1PR1/R3 inhibitor VPC23019. W6/32 stimulated SK1 activity, while siRNA silencing SK1, S1PR1 and S1PR3 inhibited hmSMC migration. In vivo, Sphingomab significantly reduced the intimal thickening induced by W6/32. These data emphasize the role of S1P in intimal hyperplasia elicited by the humoral immune response, and open perspectives for preventing TV with S1P inhibitors.

Transition from metabolic adaptation to maladaptation of the heart in obesity: role of apelin.

BACKGROUND/OBJECTIVES: Impaired energy metabolism is the defining characteristic of obesity-related heart failure. The adipocyte-derived peptide apelin has a role in the regulation of cardiovascular and metabolic homeostasis and may contribute to the link between obesity, energy metabolism and cardiac function. Here we investigate the role of apelin in the transition from metabolic adaptation to maladaptation of the heart in obese state. METHODS: Adult male C57BL/6J, apelin knock-out (KO) or wild-type mice were fed a high-fat diet (HFD) for 18 weeks. To induce heart failure, mice were subjected to pressure overload after 18 weeks of HFD. Long-term effects of apelin on fatty acid (FA) oxidation, glucose metabolism, cardiac function and mitochondrial changes were evaluated in HFD-fed mice after 4 weeks of pressure overload. Cardiomyocytes from HFD-fed mice were isolated for analysis of metabolic responses. RESULTS: In HFD-fed mice, pressure overload-induced transition from hypertrophy to heart failure is associated with reduced FA utilization (P<0.05), accelerated glucose oxidation (P<0.05) and mitochondrial damage. Treatment of HFD-fed mice with apelin for 4 weeks prevented pressure overload-induced decline in FA metabolism (P<0.05) and mitochondrial defects. Furthermore, apelin treatment lowered fasting plasma glucose (P<0.01), improved glucose tolerance (P<0.05) and preserved cardiac function (P<0.05) in HFD-fed mice subjected to pressure overload. In apelin KO HFD-fed mice, spontaneous cardiac dysfunction is associated with reduced FA oxidation (P<0.001) and increased glucose oxidation (P<0.05). In isolated cardiomyocytes, apelin stimulated FA oxidation in a dose-dependent manner and this effect was prevented by small interfering RNA sirtuin 3 knockdown. CONCLUSIONS: These data suggest that obesity-related decline in cardiac function is associated with defective myocardial energy metabolism and mitochondrial abnormalities. Furthermore, our work points for therapeutic potential of apelin to prevent myocardial metabolic abnormalities in heart failure paired with obesity.

Role of serotonin 5-HT2A receptors in the development of cardiac hypertrophy in response to aortic constriction in mice.

Serotonin, in addition to its fundamental role as a neurotransmitter, plays a critical role in the cardiovascular system, where it is thought to be involved in the development of cardiac hypertrophy and failure. Indeed, we recently found that mice with deletion of monoamine oxidase A had enhanced levels of blood and cardiac 5-HT, which contributed to exacerbation of hypertrophy in a model of experimental pressure overload. 5-HT2A receptors are expressed in the heart and mediate a hypertrophic response to 5-HT in cardiac cells. However, their role in cardiac remodeling in vivo and the signaling pathways associated are not well understood. In the present study, we evaluated the effect of a selective 5-HT2A receptor antagonist, M100907, on the development of cardiac hypertrophy induced by transverse aortic constriction (TAC). Cardiac 5-HT2A receptor expression was transiently increased after TAC, and was recapitulated in cardiomyocytes, as observed with 5-HT2A in situ labeling by immunohistochemistry. Selective blockade of 5-HT2A receptors prevented the development of cardiac hypertrophy, as measured by echocardiography, cardiomyocyte area and heart weight-to-body weight ratio. Interestingly, activation of calmodulin kinase (CamKII), which is a core mechanism in cardiac hypertrophy, was reduced in cardiac samples from M100907-treated TAC mice compared to vehicle-treated mice. In addition, phosphorylation of histone deacetylase 4 (HDAC4), a downstream partner of CamKII was significantly diminished in M100907-treated TAC mice. Thus, our results show that selective blockade of 5-HT2A receptors has beneficial effect in the development of cardiac hypertrophy through inhibition of the CamKII/HDAC4 pathway.

Delayed blockade of the kinin B1 receptor reduces renal inflammation and fibrosis in obstructive nephropathy.

Renal fibrosis is the common histological feature of advanced glomerular and tubulointerstitial disease leading to end-stage renal disease (ESRD). However, specific antifibrotic therapies to slow down the evolution to ESRD are still absent. Because persistent inflammation is a key event in the development of fibrosis, we hypothesized that the proinflammatory kinin B1 receptor (B1R) could be such a new target. Here we show that, in the unilateral ureteral obstruction model of renal fibrosis, the B1R is overexpressed and that delayed treatment with an orally active nonpeptide B1R antagonist blocks macrophage infiltration, leading to a reversal of the level of renal fibrosis. In vivo bone marrow transplantation studies as well as in vitro studies on renal cells show that part of this antifibrotic mechanism of B1R blockade involves a direct effect on resident renal cells by inhibiting chemokine CCL2 and CCL7 expression. These findings suggest that blocking the B1R is a promising antifibrotic therapy.

Hypomyelination associated with bovine viral diarrhea virus type 2 infection in a longhorn calf.

A newborn Longhorn heifer calf presented with generalized tremors, muscle fasciculations, ataxia, and nystagmus. At necropsy, no gross central nervous system lesions were observed. Histologically, the brain and spinal cord had mild to moderate diffuse microgliosis and astrocytosis, minimal nonsuppurative encephalitis, and decreased myelin staining. Ultrastructural examination revealed thinning and absence of myelin sheaths. Various cell types were immunohistochemically positive for bovine viral diarrhea virus (BVDV). Noncytopathogenic BVDV was isolated from the brain and identified as BVDV type 2 by phylogenetic analysis. BVDV-induced hypomyelination is rare and analogous to lesions in neonates infected with border disease and classical swine fever viruses. This is the first documented case of hypomyelination in a calf specifically attributed to BVDV type 2 and the first description of the ultrastructural appearance of BVDV-induced hypomyelination.

HLA class I antibodies provoke graft arteriosclerosis in human arteries transplanted into SCID/beige mice.

Antibodies toward HLA class I and/or MICA are commonly observed in transplanted patients suffering from allograft arteriosclerosis, also called chronic vascular rejection (CVR). The relative importance of cellular versus humoral alloreactivity for CVR is still disputed. We demonstrate that antibodies toward HLA class I provoke lesions typical for CVR in human arteries in vivo in the absence of cellular immunity. To show this, we grafted segments of human mesenteric arteries from 8 deceased organ donors into 36 immunodeficient SCID/beige mice in the infrarenal aortic position. Three mice died postoperatively. The remaining 33 mice received weekly i.v. injections of either a monoclonal antibody toward HLA class I, toward MICA or an irrelevant monoclonal antibody. At sacrifice after 6 weeks, mice receiving the HLA antibody showed a significant neointimal thickening in the grafted artery due to smooth muscle cell (SMC) proliferation while control mice receiving anti-MICA or irrelevant antibody showed little or no thickening. Whereas antibodies toward HLA class I were mitogenic to SMC in vitro, those directed toward MICA did not have any effect. Humoral alloreactivity toward HLA may thus play a causal role for the development of CVR and this opens new possibilities for the treatment of CVR.

Lipid and oxidative stress disorders in a rat model of chronic rejection.

Chronic allograft dysfunction is the primary cause of graft loss after the first posttransplant year. Graft arteriosclerosis, a main component of this pathology, has oxidative stress and interactions with lipid disorders as part of the pathogenesis. The objective of our study was to determine whether oxidative stress was associated with the vascular lesions observed in a rodent model of graft arteriosclerosis. Using model of orthotopic aortic allograft in the rat, the allotransplantation (A) group included 12 Sprague-Dawley donors to 12 Lewis recipients, and the isotransplantation (B) group. 12 Lewis donors to 12 Lewis recipients. The rats received no immunosuppressants or antioxidants. After 12 weeks, the rats were humanely killed and the aorta cryopreserved until analysis. Blood samples were drawn for lipid assessment and oxidative stress analysis. Tissue expression of NADPH oxidase was quantified by Western blot, determining the constitutive membrane unit (p22phox) and the cytosolic regulating unit (p67phox). We observed a greater increase in the plasma markers of oxidative stress in group A than group B but without lipid abnormalities. The expression of NADPH subunits p22phox and p67phox were similar in both groups. These results showed that oxidative stress was associated with vascular lesions in our aortic graft model, but the origin of oxidative stress seemed to be independent of the NADPH oxidase.

Use of human mesenteric arteries to study chronic vascular rejection in SCID/beige mice reconstituted with human spleen cells.

We wanted to establish a preclinical model of chronic vascular rejection (CVR) by transplanting small arteries from the mesentery of cadaveric organ donors by the rapid "sleeve" technique into SCID/beige mice reconstituted with human allogeneic spleen cells. After institutional authorization and with informed consent from relatives, we obtained tissues and cells from cadaveric organ donors. A piece of mesentery was recovered from the donor and kept in buffered solution at 4 degrees C until use. After dissection of the mesentery, small arteries of suitable size were transplanted in place of the infrarenal aorta of the mice. Cells for the immunological reconstitution of the mice were spleen cells from the same or other organ donors. Twenty-three suitable arterial segments were obtained from the mesentery of three cadaveric donors. Ten of the mice received 3 x 10(7) human spleen cells intraperitoneally 1 week after the arterial graft and they all showed circulating human CD3+ and CD19+ cells 2 weeks after injection. The mice were sacrificed 5 weeks after the arterial graft. SCID/beige mice reconstituted with allogeneic spleen cells showed a typical CVR, whereas mice that received no cells had a normal vascular anatomy. We believe our model is well suited for the study of treatment of CVR under human allograft conditions.

Chronic vascular rejection: histologic comparison between two murine experimental models.

BACKGROUND: We previously developed an experimental model to study chronic vascular rejection (CVR) in mice, the orthotopic aortic allograft. More recently we performed human arterial grafts into SCID/Beige mice reconstituted with human spleen cells. We report herein the differences in CVR lesions. MATERIAL AND METHODS: In the first model, recipient mice were C57BL/6 (H-2b), and donor mice were DBA/2 (H-2d). In the second model, terminal branches of the human superior mesenteric artery were transplanted into SCID/Beige mice in the infrarenal aorta. Human immune reconstitution was achieved by a single intraperitoneal injection of 30 x 10(6) human spleen cells. The presence of human lymphocytes and IgG was verified weekly. In both models, the vascular grafts were inserted in the infrarenal aortic position using the sleeve technique. The transplanted mice were sacrificed at 35 days after the operation. The grafts were analyzed by histology and morphometry. The mean intimal thickening was calculated based on transverse sections at 0.1-mm intervals. RESULTS: Typical CVR lesions developed with neointimal thickening, T-cell infiltration, and smooth muscle cell (SMC) proliferation in both models. In the mouse aortic model, disappearance of SMC in the media was noted in contrast to human arterial transplants, where the media remained intact. CONCLUSION: Other groups have noted that arteries conserve their media in clinical organ transplants. From this point of view, the lesions in the second experimental model (human arteries) better reflect the pathology of CVR in clinical transplantation than the murine aortic transplant model.

Human immune reconstitution with spleen cells in SCID/Beige mice.

BACKGROUND: We developed an original experimental model to study chronic vascular rejection (CVR) consisting of a graft of human mesenteric artery followed by human immune reconstitution into CB.17 SCID/Beige mice. Human immune reconstitution achieved after human PBMC injection has often been variable and incomplete. The aim of this work was to develop an alternative method to achieve a complete, functional human immune reconstitution. METHOD: After institutional authorizations, spleen cells were recovered from cadaveric organ donors. Single intraperitoneal injections of various doses of spleen cells were made into 70 CB.17 SCID/Beige mice. Reconstitution of the human immune system was monitored by flow cytometry (circulating human cells) and ELISA (human IgG). Colonization of murine lymphoid organs by human cells was studied by immunohistochemistry and flow cytometry. Evaluation of the immune function consisted of examination of CVR lesions in human arterial grafts. The animals were humanely killed at day 28. RESULTS: After injection of 30 to 40 x 10(6) spleen cells, the mice showed significant human CD3(+), CD19(+), and CD56(+) populations in peripheral blood. The mean human cells levels were, respectively, 8.2% +/- 5.4%, 2.9% +/- 1.2%, and 5.3% +/- 5.1%. Murine spleen and mesenteric lymph nodes were colonized by human T and B cells, while the murine thymus was only colonized by human T cells. Human IgG was detected in murine serum (65.9 +/- 63.3 mg/L) and typical CVR lesions were observed within the allogeneic grafts. CONCLUSION: Intraperitoneal injection of 30 to 40 x 10(6) human spleen cells into CB.17 SCID/Beige mice induces complete and functional human immune reconstitution allowing the study of CVR under human allogeneic conditions.

Factors influencing retroviral-mediated gene transfer into hepatocytes in vivo.

Direct gene transfer into hepatocytes represents an attractive alternative to organ transplantation for the treatment of genetic liver diseases. This approach is hampered either by the difficulty to obtain, cultivate, and reimplant hepatocytes or by the poor stability of the expression of the transgene. In the present report, we show that direct in vivo infection of hepatocytes with a retroviral vector following partial hepatectomy results in a life-long expression of the transgene in adult rats and mice. We demonstrate that the kinetics of hepatocyte susceptibility to infection is closely associated with the kinetics of cell division. We also present evidence that a complete vascular exclusion of the organ allows better gene transfer as compared to simple portal infusion of the viral particles, presumably through a higher volume of retrovirus-containing medium delivered to the liver.

Prevention of bone marrow and cardiac graft rejection in an H-2 haplotype disparate mouse combination by an anti-LFA-1 antibody.

The aim of the present study was to compare the preventive effect of a nondepleting monoclonal antibody specific for the LFA-1 alpha chain (CD11a) on the rejection of bone marrow, vascularized cardiac, and nonvascularized skin grafts in the same haplotype-disparate mouse strain combination. A 7-day treatment with a total dose of 0.5 mg of anti-LFA-1 antibody (H-129) had no effect on the rejection of BDF1(H-2b/d) skin grafts by CDF1 (H-2k/d)-treated mice. In contrast, the same treatment regimen significantly prolonged the survival of BDF1 cardiac allografts in CDF1 mice: 7 out of 10 mice had a functional graft after 70 days, whereas all control mice had rejected their graft by 11 days. Nevertheless, cardiac allografts were ultimately rejected. In contrast, infusion of anti-LFA-1 antibody was able to promote definitive engraftment of T-depleted BDF1 marrow in 9 gray-irradiated CDF1 recipients: in surviving mice, engraftment increased from 10% in controls to 78% in antibody-treated recipients. In mice that tolerated their cardiac graft for more than 70 days, there was a slight delay in the rejection of donor skin graft but no in vitro evidence of tolerance. In contrast, mice with successful marrow engraftment did not reject donor skin graft and failed to mount proliferative and cytotoxic responses against donor alloantigens, whatever the percentage of engrafted donor leukocytes. These results indicate that a nondepleting anti-LFA-1 antibody can efficiently protect against rejection of MHC-incompatible heart and bone marrow. The protective effect of anti-LFA-1 antibody was associated with the induction of T cell tolerance toward donor antigens after bone marrow transplantation.

High level of alpha2-adrenoceptor in rat foetal liver and placenta is due to alpha2B-subtype expression in haematopoietic cells of the erythrocyte lineage.

1. Rat foetal liver contains large amounts of alpha2-adrenoceptors. The present work aimed to identify the receptor subtype and the cell type accounting for high expression and to clarify the mechanisms responsible for the sharp decrease in hepatic receptivity occurring during the late stage of foetal development. 2. Binding experiments indicated that the density of alpha2-adrenoceptors in the foetal liver (embryonic day 18; 615+/-155 fmol mg(-1) of protein) is 18 fold higher than in newborn or adult (35.2+/-4.3 fmol mg(-1)). A high amount of receptor is also found in the placenta (443+/-53 fmol mg(-1)). In both tissues, the rank order of antagonists to inhibit radioligand binding matched the pharmacological profile of the alpha2B-adrenoceptor and exclusively RNG transcripts were detected by RNase protection assays. 3. Isolation of cell fractions from foetal liver showed that alpha2B-adrenoceptor is primarily expressed by haematopoietic cells. Consistent with this view, the receptor is found to be abundant in foetal blood, carried by reticulocytes. The expression in blood gradually declines to zero at 3 weeks of age and it is not recovered following induction of reticulocytosis in adults. 4. In foetal reticulocytes, a low proportion of the receptor population is coupled to G-protein. The alpha2-agonist UK14304 has a marginal effect on cyclic AMP level but significantly increases arachidonic acid release. The function of the receptor remains to be elucidated. However, together with observations on alpha2B-knockout mice, the current finding strongly suggests a role for alpha2B-adrenoceptor during foetal haematopoiesis in rodents.

Orthotopic aortic transplantation in mice: a new model of allograft arteriosclerosis.

BACKGROUND: Graft arteriosclerosis is a major cause of death after allotransplantation of organs such as the heart or the kidney. Aortic allotransplantation in mice is a useful experimental model to study the mechanisms of this pathology. However, the conventional heterotopic aortic model is limited by a high morbidity and is technically difficult to perform. We developed a new simple method for aortic transplantation in mice. METHODS: The infrarenal aorta from the donor mouse was anastomosed to the recipient's aorta at the same position using a sleeve technique. Orthotopic aortic transplantation was performed in 45 mice, 5 isografts and 40 allografts. No immunosuppression was given, and the mice were killed at day 15 or 30. The graft was examined macroscopically, and several histologic sections were made. RESULTS: The overall survival rate was 78%. The incidence of thrombosis was low (4 cases) compared with previously published series. Histology of aortas revealed typical aspects of rejection in the allografts with a chronic picture at day 30. No significant lesion was observed in isografts. CONCLUSIONS: We have developed a model of orthotopic aortic transplantation in mice. This new model is easy to carry out and has a low incidence of thrombosis, probably because there is no size discrepancy between donor and recipient aortic segment.

In vivo retroviral-mediated transfer of a marker-gene in ornithine transcarbamylase-deficient Spf(ash) mice.

Gene therapy is a new therapeutic approach for inherited metabolic hepatopathies. The authors studied the potential application of such a strategy to the correction of ornithine transcarbamylase (OTC) deficiency by in vivo protocol of retroviral-mediated gene transfer to the liver. A partial hepatectomy was followed (24 to 48 hours later) by asanguinous perfusion of the regenerating liver with beta-galactosidase (beta-gal) recombinant retrovirus. This protocol allowed beta-gal gene transfer in normal C57B6 mice liver with 60 +/- 52 positive cells per square centimeter. This proportion never exceeded 20 cells per square centimeter in OTC-deficient spf(ash) mice. The high mortality rate for spf(ash) mice was explained by an important sensitivity of those mice to the protein catabolism rather than by technical difficulties during intraportal perfusion. This first in vivo retroviral-mediated gene transfer study in animals with a life-threatening metabolic inherited hepatopathy showed that, despite efficiency of gene therapy in normal animal models, several experimental difficulties should be overcome before human application of this protocol is considered.

Simultaneous orthotopic transplantation of carotid and aorta in the rat by the sleeve technique.

Graft vascular disease (GVD) remains the major limitation to long-term survival after solid organ transplantation. Aortic or carotid allografts in rats have been shown to be useful models because similar changes to those observed in man develop within weeks. Both immunological and non-immunological factors influence the process of GVD and a method that could permit rapid multiple arterial allotransplantation in the rat would be of great value. We performed simultaneous orthotopic aortic and carotid allotransplantations in 25 rats. The vessels were anastomosed using a sleeve technique. No immunosuppression was given. The animals were killed at 15, 30, or 60 days and histological analyses of the grafts were performed. The overall survival rate was 80% and the incidence of technical failure was very low. The histopathological aspect revealed typical progressive GVD. In conclusion, we have developed a new model of simultaneous aortic and carotid transplantation in rats. This model, which incorporates a modification of the sleeve anastomosis, is rapid and yields an easy tool to investigate immunological and non-immunological processes driving GVD.

[Anastomoses of the small intestine in a septic environment protected by polyglactin 910-collagen mesh. Experimental study in rats]. / Anastomoses de l'intestin grêle en milieu septique protégées par un treillis de polyglactine 910-collagène. Etude expérimentale chez le rat.

An experimental study was carried out in rats to evaluate the outcome of small bowel anastomoses in the presence of peritonitis with and without protection by a polyglactin 910 mesh. One hundred and thirty rats were operated. 1) Thirty small bowel anastomoses were performed in a sterile environment to evaluate the morbidity and mortality due to the surgical procedure itself. All these animals had an uneventful course. 2) Forty anastomoses were performed in a septic environment without protection. Twelve rats died immediately after the procedure. Six rats developed an anastomotic fistula and 22 had an uneventful course. 3) Sixty anastomoses were performed in a septic environment and protected by a polyglactin 910 mesh. Seventeen rats died immediately after the procedure. There were no anastomotic disruptions. Seventeen rats had an uneventful course, and 28 (65%) developed stenosis of the anastomosis. Protection of small bowel anastomoses by a polyglactin 910 mesh appears to effectively prevent disruptions (no anastomotic fistulae or locoregional infections were recorded). However, the mesh is responsible for an intense inflammatory reaction, that often results in intestinal obstruction.

Intraparenchymal injection of bone marrow mesenchymal stem cells reduces kidney fibrosis after ischemia-reperfusion in cyclosporine-immunosuppressed rats.

Ischemia-reperfusion and immunosuppressive therapy are a major cause of progressive renal failure after kidney transplantation. Recent studies have shown that administration of bone marrow mesenchymal stem cells (MSCs) improves kidney functional recovery in the acute phase of post ischemia-reperfusion injury. In the present study, we used an original model of renal ischemia-reperfusion in immunosuppressed rats (NIRC) to investigate the effects of bone marrow MSCs on progression of chronic renal failure and the mechanisms potentially involved. Left renal ischemia-reperfusion (IR) was induced in unilateral nephrectomized Lewis rats. After IR, rats were treated daily with cyclosporine (10 mg/kg SC) for 28 days. MSCs were injected into the kidney at day 7 after IR. At day 28 after IR, kidneys were removed for histomorphological, biochemical, and gene expression analysis. The effect of conditioned media from MSCs on epithelial-mesenchymal transition was studied in vitro on HK2 cells. Our results show that, as compared to untreated NIRC rats, rats treated by intrarenal injection of MSCs 7 days after IR displayed improvement in renal function, reduction of interstitial fibrosis, and decrease in chronic tubule injury. These effects were associated with a decrease in interstitial α-SMA accumulation and MMP2 activity, markers of fibroblast/fibroblast-like cell activation, and renal remodeling, respectively. Finally, experiments in vitro showed that MSC-conditioned medium prevented epithelial-mesenchymal transition induced by TGF-ß in HK2 cells. In conclusion, our results show that, in immunosuppressed animals, a single intrarenal administration of MSCs reduced renal fibrosis and promoted the recovery of renal function.

Evaluation of alginate microspheres for mesenchymal stem cell engraftment on solid organ.

Mesenchymal stem cells (MSCs) may be used as a cell source for cell therapy of solid organs due to their differentiation potential and paracrine effect. Nevertheless, optimization of MSC-based therapy needs to develop alternative strategies to improve cell administration and efficiency. One option is the use of alginate microencapsulation, which presents an excellent biocompatibility and an in vivo stability. As MSCs are hypoimmunogenic, it was conceivable to produce microparticles with [alginate-poly-L-lysine-alginate (APA) microcapsules] or without (alginate microspheres) a surrounding protective membrane. Therefore, the aim of this study was to determine the most suitable microparticles to encapsulate MSCs for engraftment on solid organ. First, we compared the two types of microparticles with 4 × 10(6) MSCs/ml of alginate. Results showed that each microparticle has distinct morphology and mechanical resistance but both remained stable over time. However, as MSCs exhibited a better viability in microspheres than in microcapsules, the study was pursued with microspheres. We demonstrated that viable MSCs were still able to produce the paracrine factor bFGF and did not present any chondrogenic or osteogenic differentiation, processes sometimes reported with the use of polymers. We then proved that microspheres could be implanted under the renal capsule without degradation with time or inducing impairment of renal function. In conclusion, these microspheres behave as an implantable scaffold whose biological and functional properties could be adapted to fit with clinical applications.