Hans-Ulrich Demuth (Probiodrug, Halle/Saale, Germany). splenic, but not thymic or lymph node CD4+ T-cells, from nondiabetic NOD mice with soluble (s) DPP-IV increased migration. Sitagliptin abolished sDPP-IV effects on splenic CD4+ T-cell migration, whereas incretins decreased migration of lymph node, but not splenic, CD4+ T-cells. Splenic CD4+ T-cells demonstrating increased in vitro migration in response to sDPP-IV and lymph node CD4+ T-cells that were nonresponsive to incretins selectively infiltrated islets of NOD mice, after injection. Sitagliptin decreases migration of splenic CD4+ T-cells through a pathway involving Rac1/vasodilator-stimulated phosphoprotein, whereas its inhibitory effects around the migration of lymph Rabbit Polyclonal to DDX55 node CD4+ T-cells involve incretin-activation of the NF-B pathway. CONCLUSIONS Benefits of sitagliptin treatment in diabetic NOD mice may be mediated through selective effects on subpopulations of T-cells that are related to autoimmunity. The incretin hormones, glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide (GLP)-1, potentiate glucose-stimulated insulin secretion during a meal and exert additional actions, including promotion of -cell survival and proliferation (1C6). GIP and GLP-1 are primarily metabolized by the endopeptidase dipeptidyl peptidase IV (DPP-IV) (CD26), and both inhibitors of DPP-IV activity and RHPS4 DPP-IV-resistant incretin analogs have been targeted as type 2 diabetes therapeutic drugs, with the incretin mimetic exenatide (Byetta) and the DPP-IV inhibitors sitagliptin (Januvia) and saxagliptin (Onglyza) receiving U.S. Food and Drug Administration approval. Although the actions of DPP-IV inhibitors have been extensively studied for treatment of type 2 diabetes, considerably less RHPS4 is known about their potential in type 1 diabetes. In earlier studies, the DPP-IV inhibitor isoleucine thiazolidide was shown to improve glucose tolerance in type 1 diabetic animal models by increasing -cell survival and, possibly, neogenesis (7,8). Additionally, sitagliptin (MK0431) was demonstrated to prolong islet graft survival in streptozotocin-induced (9) and NOD (10) mice. In the latter study, sitagliptin guarded the islet graft through a mechanism that included modulation of splenic CD4+ T-cell migration (10). This response appeared to involve inhibition of direct DPP-IV effects on CD4+ T-cells, rather than through increasing levels of active incretins by preventing their degradation. However, the GLP-1 receptor (GLP-1R) is usually expressed in lymphoid tissue, and exendin-4 treatment was shown to increase numbers of CD4+ and CD8+ T-cells in lymph nodes and reduce the number of CD4+CD25+Foxp3+ regulatory T-cells in the thymus, but not the spleen, suggesting specific effects on different subpopulations of cells (11). One objective of the current studies was to examine responses to sitagliptin in additional subsets of CD4+ T-cell, including those from the thymus and lymph nodes. Using a double-labeling technique, we also examined whether in vitro treatment of splenic CD4+ T-cells with soluble (s) DPP-IV, or treatment of those from the lymph node with incretins, altered their ability to infiltrate islets of diabetic NOD mice. Previously sDPP-IV was shown to increase migration of splenic CD4+ T-cells via a pathway involving cAMP/protein kinase A (PKA)/Rac1 GTP binding activity, with DPP-IV inhibition abolishing these effects (10). Active, GTP-bound Rac1 plays an important role in regulating cell migration through modulation of actin-rich lamellipodial protrusions, crucial components for generating the driving pressure of cell movement (12). In several systems, inhibition of Rac resulted in complete prevention of cell movement (13C15), thus demonstrating its crucial role. In the current study, we examined whether a protein involved in actin reorganization, vasodilator-stimulated phosphoprotein (VASP), contributes to effects of sDPP-IV on CD4+ T-cell migration. We demonstrate that administration of sitagliptin in vivo reduces lymph node and splenic CD4+ T-cell migration, measured in vitro, via incretin- and nonincretin-mediated effects, respectively, and splenic sDPP-IV-responsive CD4+ T-cells and lymph node incretin nonresponsive CD4+ T-cells selectively infiltrated islets of diabetic NOD mice, after tail vein injection. We also identified a downstream role for VASP in sDPP-IV-stimulated CD4+ T-cell migration and for nuclear factor-B (NF-B) in GIP and GLP-1 stimulation of lymph node CD4+ T-cell migration. RESEARCH DESIGN AND METHODS Mice. NOD/LtJ RHPS4 mice (NOD, RHPS4 H2g7) were purchased from The Jackson Laboratory (Bar Harbor, ME). Mice (8C10 weeks aged) were fed either a normal chow diet (NCD) (Purina Rodent Chow 5015) or a diet made up of sitagliptin (16) (Purina Rodent Chow 5015 plus 4 g MK0431/kg; Research Diets, RHPS4 New Brunswick, NJ). All animal experiments were conducted in accordance with guidelines put forth by the University of British Columbia Committee on Animal Care and Canadian Council on Animal Care. CD4+ T-cell isolation. T-cells were prepared from.