Mesenchymal stem cells and a vitamin D receptor agonist additively suppress T helper 17 cells and the related inflammatory response in the kidney.

Mesenchymal stem cells (MSCs) suppress T helper (Th)17 cell differentiation and are being clinically pursued for conditions associated with aberrant Th17 responses. Whether such immunomodulatory effects are enhanced by coadministration of MSCs with other agents is not well known. In the present study, individual and combined effects of MSCs and the vitamin D receptor (VDR) agonist paricalcitol on Th17 induction were investigated in vitro and in a mouse model of sterile kidney inflammation (unilateral ureteral obstruction). In vitro, MSCs and paricalcitol additively suppressed Th17 differentiation, although only MSCs suppressed expression of Th17-associated transcriptions factors. Combined administration of MSCs and paricalcitol resulted in an early (day 3) reduction of intrarenal CD4(+) and CD8(+) T cells, CD11b(+)/lymphocyte antigen 6G(+) neutrophils, and inflammatory (lymphocyte antigen 6C(hi)) monocytes as well as reduced transcript for IL-17 compared with untreated animals. Later (day 8), obstructed kidneys of MSC/paricalcitol double-treated mice, but not mice treated with either intervention alone, had reduced tubular injury and interstitial fibrosis as well as lower numbers of neutrophils and inflammatory monocytes and an increase in the ratio between M2 (CD206(+)) and M1 (CD206(-)) macrophages compared with control mice. Adjunctive therapy with VDR agonists may enhance the immunosuppressive properties of MSCs in the setting of pathogenic Th17-type immune responses and related inflammatory responses.

The influence of hypoxia on the differentiation capacities and immunosuppressive properties of clonal mouse mesenchymal stromal cell lines.

Multipotent mesenchymal stromal cells are multipotent cells capable of differentiating into different mesodermal cell types. Enigmatically, mesenchymal stromal cells present in the bone marrow support early lymphopoiesis yet can inhibit mature lymphocyte growth. Critical features of the bone marrow microenvironment, such as the level of oxygen, play an important role in mesenchymal stromal cell biology. Herein, we show that a panel of continuously growing mouse mesenchymal stromal cell lines, namely OP9, MS5, PA6, ST2 and B16-14, exhibit mesenchymal stromal cell characteristic phenotypes and respond physiologically to oxygen deprivation. Culturing freshly isolated bone marrow-derived mesenchymal stromal cells or cell lines at 5% O2 resulted in a dramatic increase in expression of hypoxia-inducible factor family members and of key genes involved in their differentiation. Phenotypically, their osteogenic and adipogenic differentiation capacity was generally improved in hypoxia, whereas their inhibitory effects on in vitro T-cell proliferation were preserved. Taken together, we conclude that these continuously growing mouse cell lines behave as canonical mesenchymal stromal cells and respond physiologically to hypoxia, thereby providing a potent tool for the study of different aspects of mesenchymal stromal cell biology.

Back from the brink: a mesenchymal stem cell infusion rescues kidney function in acute experimental rhabdomyolysis.

Systemic administration of mesenchymal stem (stromal) cells (MSCs) has shown benefit in a range of experimental models of acute kidney injury, although the reported mechanisms of action and requirement for MSC localization to the kidney have varied. Geng and colleagues now demonstrate that a single intravenous infusion of MSCs given 6 hours after induction of acute muscle necrosis (rhabdomyolysis) robustly ameliorates the resulting acute kidney injury and promotes early intra-renal accumulation of CD206+ (M2) macrophages. The benefit occurred in the absence of MSC localization to the kidney and could be reproduced by adoptive transfer of ex vivo-programmed M2 macrophages.

IL-10 mediates the immunoregulatory response in conjugated linoleic acid-induced regression of atherosclerosis.

Conjugated linoleic acid (CLA) induces regression of preestablished atherosclerosis in the ApoE(-/-) mouse. Understanding the mechanisms involved may help in identifying novel pathways associated with the regression of human disease. Animals were administered a 1% cholesterol diet for 12 wk, with 1% CLA supplementation from wk 8 to 12. ApoE(-/-) mice fed only the 1% cholesterol diet for 12 wk were employed as controls. Transcriptomic analysis of mouse aorta showed that many of the components of the IL-10 signaling pathway were modified during CLA-induced regression. Real-time PCR and Western blot analysis showed increased IL-10 receptor expression, phosphorylation of STAT3, and downstream target gene expression in the aorta, alongside an increase in serum IL-10 (79.8 ± 22.4 vs. 41.9 ± 5.5 pg/ml, n = 10; P < 0.01). CLA -supplementation also increased IL-10 production in bone marrow-derived macrophages (143.6 ± 28.6 vs. 94 ± 5.6 pg/ml, n = 5; P < 0.05). To explore the mechanisms for altered IL-10 production, we examined the profile of monocyte/macrophage phenotype in the vessel wall, bone marrow, and spleen. CLA increased macrophage polarization toward an anti-inflammatory M2 phenotype in vivo, increasing the population of Ly6C(lo) monocytes (29 vs. 77 ± 14, n=5, P < 0.05) in the aorta. CLA had similar effects on monocytes/macrophages differentiated from marrow-derived progenitor cells and on splenocytes. The induction of IL-10 on CLA supplementation in this model may reflect a systemic alteration toward an anti-inflammatory phenotype, which, in turn promotes increased vascular infiltration by Ly6C(lo) monocytes. These cells may contribute to CLA-induced disease regression.

Interleukin-1 accounts for intrarenal Th17 cell activation during ureteral obstruction.

Interleukin 17A-secreting T-helper 17 (Th17) cells are pathogenic in inflammatory kidney diseases, but their intrarenal regulation is poorly understood. In order to better define Th17 cell dynamics during interstitial inflammation, we utilized the mouse unilateral ureteral obstruction model to analyze inflammatory cell subtypes by multicolor flow cytometry and cell sorting and by effects on in vitro-generated Th17 cells. Interleukin 17A expression localized to CCR6(+)CCR4(+/-)CD4(+) T-cells and progressively increased in obstructed kidneys. The number of CCR6(+)CD4(+) T-cells increased over 10-fold by 72 h, were enriched for interleukin 17A production, and were highly proliferative based on in vivo bromodeoxyuridine incorporation. Secreted products of leukocytes isolated from obstructed kidneys enhanced the interleukin 17A production of in vitro-generated Th17 cells. This Th17-enhancing activity was identified as interleukin-1 produced by renal dendritic cells and monocytes. The in vivo validity of these findings was confirmed in mice lacking the interleulin-1 receptor and in mice treated with a recombinant interleukin-1 receptor antagonist, each of which exhibited reduced intrarenal Th17 activity compared with control mice. Thus, the inflamed kidney accumulates CCR6(+) Th17 cells that undergo activation and proliferation. Production of interleukin 1 family cytokines by resident dendritic cells and infiltrating monocytes enhances intrarenal Th17 activation in acute kidney injury.

Mesenchymal stem cell effects on T-cell effector pathways.

Mesenchymal stem (stromal) cells (MSCs) are rare, multipotent progenitor cells that can be isolated and expanded from bone marrow and other tissues. Strikingly, MSCs modulate the functions of immune cells, including T cells, B cells, natural killer cells, monocyte/macrophages, dendritic cells, and neutrophils. T cells, activated to perform a range of different effector functions, are the primary mediators of many autoimmune and inflammatory diseases as well as of transplant rejection and graft-versus-host disease. Well-defined T-cell effector phenotypes include the CD4+ (T helper cell) subsets Th1, Th2, and Th17 cells and cytotoxic T lymphocytes derived from antigen-specific activation of naïve CD8+ precursors. In addition, naturally occurring and induced regulatory T cells (Treg) represent CD4+ and CD8+ T-cell phenotypes that potently suppress effector T cells to prevent autoimmunity, maintain self-tolerance, and limit inflammatory tissue injury. Many immune-mediated diseases entail an imbalance between Treg and effector T cells of one or more phenotypes. MSCs broadly suppress T-cell activation and proliferation in vitro via a plethora of soluble and cell contact-dependent mediators. These mediators may act directly upon T cells or indirectly via modulation of antigen-presenting cells and other accessory cells. MSC administration has also been shown to be variably associated with beneficial effects in autoimmune and transplant models as well as in several human clinical trials. In a small number of studies, however, MSC administration has been found to aggravate T cell-mediated tissue injury. The multiple effects of MSCs on cellular immunity may reflect their diverse influences on the different T-cell effector subpopulations and their capacity to specifically protect or induce Treg populations. In this review, we focus on findings from the recent literature in which specific modulatory effects of MSCs on one or more individual effector T-cell subsets and Treg phenotypes have been examined in vitro, in relevant animal models of in vivo immunological disease, and in human subjects. We conclude that MSCs have the potential to directly or indirectly inhibit disease-associated Th1, Th2, and Th17 cells as well as cytotoxic T lymphocytes but that many key questions regarding the potency, specificity, mechanistic basis, and predictable therapeutic value of these modulatory effects remain unanswered.

Mesenchymal stem cell inhibition of T-helper 17 cell- differentiation is triggered by cell-cell contact and mediated by prostaglandin E2 via the EP4 receptor.

Mesenchymal stem cells (MSCs) inhibit T-cell activation and proliferation but their effects on individual T-cell-effector pathways and on memory versus naïve T cells remain unclear. MSC influence on the differentiation of naïve and memory CD4(+) T cells toward the Th17 phenotype was examined. CD4(+) T cells exposed to Th17-skewing conditions exhibited reduced CD25 and IL-17A expression following MSC co-culture. Inhibition of IL-17A production persisted upon re-stimulation in the absence of MSCs. These effects were attenuated when cell-cell contact was prevented. Th17 cultures from highly purified naïve- and memory-phenotype responders were similarly inhibited. Th17 inhibition by MSCs was reversed by indomethacin and a selective COX-2 inhibitor. Media from MSC/Th17 co-cultures contained increased prostaglandin E2 (PGE2) levels and potently suppressed Th17 differentiation in fresh cultures. MSC-mediated Th17 inhibition was reversed by a selective EP4 antagonist and was mimicked by synthetic PGE2 and a selective EP4 agonist. Activation-induced IL-17A secretion by naturally occurring, effector-memory Th17 cells from a urinary obstruction model was also inhibited by MSC co-culture in a COX-dependent manner. Overall, MSCs potently inhibit Th17 differentiation from naïve and memory T-cell precursors and inhibit naturally-occurring Th17 cells derived from a site of inflammation. Suppression entails cell-contact-dependent COX-2 induction resulting in direct Th17 inhibition by PGE2 via EP4.