A major challenge in transplantation medicine is controlling the very strong immune responses to foreign antigens that are responsible for graft rejection. Although immunosuppressive drugs efficiently inhibit acute graft rejection, a substantial proportion of patients suffer chronic rejection that ultimately leads to functional loss of the graft. Induction of immunological tolerance to transplants would avoid rejection and the need for lifelong treatment with immunosuppressive drugs. Tolerance to self-antigens is ensured naturally by several mechanisms; one major mechanism depends on the activity of regulatory T lymphocytes. Here we show that in mice treated with clinically acceptable levels of irradiation, regulatory CD4+CD25+Foxp3+ T cells stimulated in vitro with alloantigens induced long-term tolerance to bone marrow and subsequent skin and cardiac allografts. Regulatory T cells specific for directly presented donor antigens prevented only acute rejection, despite hematopoietic chimerism. By contrast, regulatory T cells specific for both directly and indirectly presented alloantigens prevented both acute and chronic rejection. Our findings demonstrate the potential of appropriately stimulated regulatory T cells for future cell-based therapeutic approaches to induce lifelong immunological tolerance to allogeneic transplants.

+ T-cell subsets (CD8KO). In contrast to CD8KO mice, MHCIIKO mice did not develop ventricular dilation and dysfunction. MHCIIKO mice also displayed very low fibrosis, collagen accumulation, and cross-linking within cardiac tissue. Interestingly, mice with transgenic CD4 + T-cell receptor specific for ovalbumin failed to develop HF and adverse remodeling.

Recent studies showed that mesenchymal stem cells (MSCs) transplantation significantly decreased cardiac fibrosis; however, the mechanisms involved in these effects are still poorly understood. In this work, we investigated whether the antifibrotic properties of MSCs involve the regulation of matrix metalloproteinases (MMPs) and matrix metalloproteinase endogenous inhibitor (TIMP) production by cardiac fibroblasts. In vitro experiments showed that conditioned medium from MSCs decreased viability, a-smooth muscle actin expression, and collagen secretion of cardiac fibroblasts. These effects were concomitant with the stimulation of MMP-2/MMP-9 activities and membrane type 1 MMP expression. Experiments performed with fibroblasts from MMP2-knockout mice demonstrated that MMP-2 plays a preponderant role in preventing collagen accumulation upon incubation with conditioned medium from MSCs. We found that MSC-conditioned medium also decreased the expression of TIMP2 in cardiac fibroblasts. In vivo studies showed that intracardiac injection of MSCs in a rat model of postischemic heart failure induced a significant decrease in ventricular fibrosis. This effect was associated with the improvement of morphological and functional cardiac parameters. In conclusion, we showed that MSCs modulate the phenotype of cardiac fibroblasts and their ability to degrade extracellular matrix. These properties of MSCs open new perspectives for understanding the mechanisms of action of MSCs and anticipate their potential therapeutic or side effects. STEM

ABSTRACT

Tubulointerstitial fibrosis in chronic renal disease is strongly associated with progressive loss of renal function. We studied the potential involvement of lysophosphatidic acid (LPA), a growth factor-like phospholipid, and its receptors LPA 1-4 in the development of tubulointerstitial fibrosis (TIF). Renal fibrosis was induced in mice by unilateral ureteral obstruction (UUO) for up to 8 d, and kidney explants were prepared from the distal poles to measure LPA release into conditioned media. After obstruction, the extracellular release of LPA increased approximately 3-fold. Real-time reverse transcription PCR (RT-PCR) analysis demonstrated significant upregulation in the expression of the LPA 1 receptor subtype, downregulation of LPA 3 , and no change of LPA 2 or LPA 4 . TIF was significantly attenuated in LPA 1 (Ϫ/Ϫ) mice compared to wild-type littermates, as measured by expression of collagen III, ␣-smooth muscle actin (␣-SMA), and F4/80. Furthermore, treatment of wild-type mice with the LPA 1 antagonist Ki16425 similarly reduced fibrosis and significantly attenuated renal expression of the profibrotic cytokines connective tissue growth factor (CTGF) and transforming growth factor ␤ (TGF␤). In vitro, LPA induced a rapid, dose-dependent increase in CTGF expression that was inhibited by Ki16425. In conclusion, LPA, likely acting through LPA 1 , is involved in obstruction-induced TIF. Therefore, the LPA 1 receptor might be a pharmaceutical target to treat renal fibrosis.

Abstract-The mitochondrial enzyme monoamine oxidase (MAO), its isoform MAO-A, plays a major role in reactive oxygen species-dependent cardiomyocyte apoptosis and postischemic cardiac damage. In the current study, we investigated whether sphingolipid metabolism can account for mediating MAO-A-and reactive oxygen speciesdependent cardiomyocyte apoptosis. In H9c2 cardiomyoblasts, MAO-A-dependent reactive oxygen species generation led to mitochondria-mediated apoptosis, along with sphingosine kinase-1 (SphK1) inhibition. These phenomena were associated with generation of proapoptotic ceramide and decrease in prosurvival sphingosine 1-phosphate. These events were mimicked by inhibition of SphK1 with either pharmacological inhibitor or small interfering RNA, as well as by extracellular addition of C 2 -ceramide or H 2 O 2 . In contrast, enforced expression of SphK1 protected H9c2 cells from serotonin-or H 2 O 2 -induced apoptosis. Analysis of cardiac tissues from wild-type mice subjected to ischemia/reperfusion revealed significant upregulation of ceramide and inhibition of SphK1. It is noteworthy that SphK1 inhibition, ceramide accumulation, and concomitantly infarct size and cardiomyocyte apoptosis were significantly decreased in MAO-Adeficient animals. In conclusion, we show for the first time that the upregulation of ceramide/sphingosine 1-phosphate ratio is a critical event in MAO-A-mediated cardiac cell apoptosis. In addition, we provide the first evidence linking generation of reactive oxygen species with SphK1 inhibition. Key Words: sphingosine kinase-1 Ⅲ monoamine oxidases Ⅲ ischemia/reperfusion Ⅲ ROS Ⅲ serotonin T he ischemia/reperfusion (I/R) syndrome occurring during revascularization results in the progressive loss of cardiac myocytes, ultimately leading to heart failure. 1 A wealth of reports indicate a pivotal role for apoptosis in cardiac myocyte death after I/R. 2-4 Indeed, cardiomyocyte apoptosis was found during the early phase of reperfusion after myocardial ischemia in animal models 5,6 and after cardiac transplantation in humans. 7,8 In addition, prevention of apoptosis strongly reduced postischemic cardiac damage.Abundant evidence suggests that reactive oxygen species (ROS) play an important role in the development and progression of postischemic myocardial damage, leading to apoptosis. 9,10 The induction of oxidative stress is related to an activation of several enzymatic systems including monoamine oxidases (MAOs). 11-13 MAOs catalyze oxidative deamination of several monoamines (eg, serotonin [5-hydroxytryptamine{5-HT}], noradrenaline, dopamine), resulting in significant ROS production. 14 Based on their substrate preference and inhibitor specificity, 2 functional isoenzymes, MAO-A and MAO-B, have been identified. 15 In the heart, MAO-A is a predominant enzyme involved in the deamination of endogenous or exogenous amines. 16 Recently, we have shown that oxidative stress induced by MAO-A is responsible for receptor-independent, serotonin-mediated apoptosis during postischemic myocardial injury....

Our data provide the first evidence that apelin inhibits TGF-β-stimulated activation of cardiac fibroblasts through a SphK1-dependent mechanism. We also demonstrated that the administration of apelin during the phase of reactive fibrosis prevents structural remodelling of the myocardium and ventricular dysfunction. These findings may have important implications for designing future therapies for myocardial performance during fibrotic remodelling, affecting the clinical management of patients with progressive heart failure.

Adipose tissue secretes a variety of bioactive factors, which can regulate cardiomyocyte hypertrophy via reactive oxygen species (ROS). In the present study we investigated whether apelin affects ROS-dependent cardiac hypertrophy. In cardiomyocytes apelin inhibited the hypertrophic response to 5-HT and oxidative stress induced by 5-HT- or H(2)O(2) in a dose-dependent manner. These effects were concomitant to the increase in mRNA expression and activity of catalase. Chronic treatment of mice with apelin attenuated pressure-overload-induced left ventricular hypertrophy. The prevention of hypertrophy by apelin was associated with increased myocardial catalase activity and decreased plasma lipid hydroperoxide, as an index of oxidative stress. These results show that apelin behaves as a catalase activator and prevents cardiac ROS-dependent hypertrophy.