Injury to a target organ can be sensed by bone marrow stem cells that migrate to the site of damage, undergo differentiation, and promote structural and functional repair. This remarkable stem cell capacity prompted an investigation of the potential of mesenchymal and hematopoietic stem cells to cure acute renal failure. The model of renal injury induced in mice by the anticancer agent cisplatin was chosen. Injection of mesenchymal stem cells of male bone marrow origin remarkably protected cisplatin-treated syngeneic female mice from renal function impairment and severe tubular injury. Y chromosome–containing cells localized in the context of the tubular epithelial lining and displayed binding sites for Lens culinaris lectin, indicating that mesenchymal stem cells engraft the damaged kidney and differentiate into tubular epithelial cells, thereby restoring renal structure and function. Mesenchymal stem cells markedly accelerated tubular proliferation in response to cisplatin-induced damage, as revealed by higher numbers of Ki-67–positive cells within the tubuli with respect to cisplatin-treated mice that were given saline. Hematopoietic stem cells failed to exert beneficial effects. These results offer a strong case for exploring the possibility that mesenchymal stem cells by virtue of their renotropic property and tubular regenerative potential may have a role in the treatment of acute renal failure in humans.

The renin-angiotensin system plays a role in the etiology of hypertension and the pathophysiology of cardiac and renal diseases in humans. Ang II is the central product of this system and is involved in regulating immune responses, inflammation, cell growth, and proliferation by acting through Ang II type 1 receptors (AT₁ and AT₂). Here, we show that targeted disruption of the Agtr1a gene that encodes AT_1A results in marked prolongation of life span in mice. Agtr1a^–/– mice developed less cardiac and vascular injury, and multiple organs from these mice displayed less oxidative damage than wild-type mice. The longevity phenotype was associated with an increased number of mitochondria and upregulation of the prosurvival genes nicotinamide phosphoribosyltransferase (Nampt) and sirtuin 3 (Sirt3) in the kidney. In cultured tubular epithelial cells, Ang II downregulated Sirt3 mRNA, and this effect was inhibited by an AT₁ antagonist. These results demonstrate that disruption of AT₁ promotes longevity in mice, possibly through the attenuation of oxidative stress and overexpression of prosurvival genes, and suggests that the Ang II/AT₁ pathway may be targeted to influence life span in mammals.

In the present study, we examined the hypothesis that dynamic characteristics of flow modulate the production of vasoactive mediators, namely nitric oxide (NO) and endothelin-1 (ET-1), by human umbilical vein endothelial cells (HUVECs). Cells were exposed for 6 hours in a cone-and-plate apparatus to different types of flow: steady laminar, with shear stresses of 2, 8, and 12 dyne/cm ² ; pulsatile laminar, with shear stress from 8.2 to 16.6 dyne/cm ² and a frequency of 2 Hz; periodic laminar, with square wave cycles of 15 minutes and shear stress from 2 to 8 dyne/cm ² ; and turbulent, with shear stress of 8 dyne/cm ² on average. A second culture dish was kept in a normal incubator as a static control for each experiment. Laminar flow induced synthesis of NO by HUVECs that was dependent on shear-stress magnitude. Laminar shear stress at 8 dyne/cm ² also upregulated the level of NO synthase mRNA. As observed with steady laminar flow, pulsatile flow also induced an increase in NO release by endothelial cells. When HUVECs were subjected to step-change increases of laminar shear, a further increase of NO synthesis was observed, compared with steady laminar shear of the same magnitude. Turbulent flow did not upregulate NO synthase mRNA or increase NO release. Both laminar and turbulent shear stress reduced, although not significantly, ET-1 mRNA and ET-1 production compared with the static condition. These results indicate that local blood flow conditions modulate the production of vasoactive substances by endothelial cells. This may affect vascular cell functions such as nonthrombogenicity, regulation of blood flow, and vascular tone.

Acute kidney injury (AKI) is a public health concern with an annual mortality rate that exceeds those of breast and prostate cancer, heart failure, and diabetes combined. Oxidative stress and mitochondrial damage are drivers of AKI-associated pathology; however, the pathways that mediate these events are poorly defined. Here, using a murine cisplatin-induced AKI model, we determined that both oxidative stress and mitochondrial damage are associated with reduced levels of renal sirtuin 3 (SIRT3). Treatment with the AMPK agonist AICAR or the antioxidant agent acetyl-l-carnitine (ALCAR) restored SIRT3 expression and activity, improved renal function, and decreased tubular injury in WT animals, but had no effect in Sirt3^–/– mice. Moreover, Sirt3-deficient mice given cisplatin experienced more severe AKI than WT animals and died, and neither AICAR nor ALCAR treatment prevented death in Sirt3^–/– AKI mice. In cultured human tubular cells, cisplatin reduced SIRT3, resulting in mitochondrial fragmentation, while restoration of SIRT3 with AICAR and ALCAR improved cisplatin-induced mitochondrial dysfunction. Together, our results indicate that SIRT3 is protective against AKI and suggest that enhancing SIRT3 to improve mitochondrial dynamics has potential as a strategy for improving outcomes of renal injury.

In this study, we investigated whether mesenchymal stem cells (MSC) had immunomodulatory properties in solid organ allotransplantation, using a semiallogeneic heart transplant mouse model, and studied the mechanism(s) underlying MSC tolerogenic effects. Either single (portal vein, day −7) or double (portal vein, day −7 and tail vein, day −1) pretransplant infusions of donor-derived B6C3 MSC in B6 recipients induced a profound T cell hyporesponsiveness and prolonged B6C3 cardiac allograft survival. The protolerogenic effect was abrogated when donor-derived MSC were injected together with B6C3 hematopoietic stem cells (HSC), suggesting that HSC negatively impact MSC immunomodulatory properties. Both the induction (pretransplant) and the maintenance phase (>100 days posttransplant) of donor-derived MSC-induced tolerance were associated with CD4⁺CD25⁺Foxp3⁺ Treg expansion and impaired anti-donor Th1 activity. MSC-induced regulatory T cells (Treg) were donor-specific since adoptive transfer of splenocytes from tolerant mice prevented the rejection of fully MHC-mismatched donor-specific secondary allografts but not of third-party grafts. In addition, infusion of recipient-derived B6 MSC tolerized a semiallogeneic B6C3 cardiac allograft, but not a fully MHC-mismatched BALB/c graft, and expanded Treg. A double i.v. pretransplant infusion of recipient-derived MSC had the same tolerogenic effect as the combined intraportal/i.v. MSC infusions, which makes the tolerogenic protocol applicable in a clinical setting. In contrast, single MSC infusions given either peritransplant or 1 day after transplant were less effective. Altogether these findings indicate that MSC immunomodulatory properties require HSC removal, partial sharing of MHC Ags between the donor and the recipient and pretransplant infusion, and are associated with expansion of donor-specific Treg.

We addressed the role of hyperglycemia in leukocyte-endothelium interaction under flow conditions by exposing human umbilical vein endothelial cells for 24 h to normal (5 mM), high concentration of glucose (30 mM), advanced glycosylation end product-albumin (100 microg/ml), or hyperglycemic (174-316 mg/dl) sera from patients with diabetes and abnormal hemoglobin A1c (8.1+/-1.4%). At the end of incubation endothelial cells were perfused with total leukocyte suspension in a parallel plate flow chamber under laminar flow (1.5 dyn/cm2). Rolling and adherent cells were evaluated by digital image processing. Results showed that 30 mM glucose significantly (P < 0. 01) increased the number of adherent leukocytes to endothelial cells in respect to control (5 mM glucose; 151+/-19 versus 33+/-8 cells/mm2). A similar response was induced by endothelial stimulation with IL-1beta, here used as positive control (195+/-20 cells/mm2). The number of rolling cells on endothelial surface was not affected by high glucose level. Stable adhesion of leukocytes to glucose-treated as well as to IL-1beta-stimulated endothelial cells was preceded by short interaction of leukocytes with the endothelial surface. The distance travelled by leukocytes before arrest on 30 mM glucose, or on IL-1beta-treated endothelial cells, was significantly (P < 0.01) higher than that observed for leukocytes adhering on control endothelium (30 mM glucose: 76.7+/-3.5; IL1beta: 69.7+/-4 versus 5 mM glucose: 21.5+/-5 microm). Functional blocking of E-selectin, intercellular cell adhesion molecule-1, and vascular cell adhesion molecule-1 on endothelial cells with the corresponding mouse mAb significantly inhibited glucose-induced increase in leukocyte adhesion (67+/-16, 83+/-12, 62+/-8 versus 144+/-21 cells/ mm2). Confocal fluorescence microscopy studies showed that 30 mM glucose induced an increase in endothelial surface expression of E-selectin, intercellular cell adhesion molecule-1, and vascular cell adhesion molecule-1. Electrophoretic mobility shift assay of nuclear extracts of human umbilical vein endothelial cells (HUVEC) exposed for 1 h to 30 mM glucose revealed an intense NF-kB activation. Treatment of HUVEC exposed to high glucose with the NF-kB inhibitors pyrrolidinedithiocarbamate (100 microM) and tosyl-phe-chloromethylketone (25 microM) significantly reduced (P < 0.05) leukocyte adhesion in respect to HUVEC treated with glucose alone. A significant (P < 0.01) inhibitory effect on glucose-induced leukocyte adhesion was observed after blocking protein kinase C activity with staurosporine (5 nM). When HUVEC were treated with specific antisense oligodesoxynucleotides against PKCalpha and PKCepsilon isoforms before the addition of 30 mM glucose, a significant (P < 0.05) reduction in the adhesion was also seen. Advanced glycosylation end product-albumin significantly increased the number of adhering leukocytes in respect to native albumin used as control (110+/-16 versus 66+/-7, P < 0.01). Sera from diabetic patients significantly (P < 0.01) enhanced leukocyte adhesion as compared with controls, despite normal levels of IL-1beta and TNFalpha in these sera. These data indicate that high glucose concentration and hyperglycemia promote leukocyte adhesion to the endothelium through upregulation of cell surface expression of adhesive proteins, possibly depending on NF-kB activation.