Glutaminolysis is a well-known source of energy for effector T cells but its contribution to each T cell subset and the mechanisms which are responsible for the control of involved metabolic enzymes are not fully understood. We report that Th17 but not Th1, Th2, or Treg cell induction in vitro depends on glutaminolysis and the up-regulation of glutaminase 1 (Gls1), the first enzyme in the glutaminolysis pathway. Both pharmacological and siRNA-based selective inhibition of Gls1 reduced in vitro Th17 differentiation and reduced the CD3/TCR-mediated increase of the mammalian target of rapamycin complex 1 activity. Treatment of mice with a Gls1 inhibitor ameliorated experimental autoimmune encephalomyelitis. Furthermore, -deficient mice that received Gls1-shRNA-transfected 2D2 T cells had reduced experimental autoimmune encephalomyelitis scores compared with those that received control-shRNA-treated cells. Next we found that T cells deficient in inducible cAMP early repressor (ICER), a transcriptional factor known to promote Th17 differentiation, display reduced activity of oxidative phosphorylation rates in the presence of glutamine and reduced Gls1 expression, both of which could be restored by ICER overexpression. Finally, we demonstrate that ICER binds to the promoter directly and increases its activity. These findings demonstrate the importance of glutaminolysis in the generation of Th17 and the direct control of Gls1 activity by the IL-17-promoting transcription factor ICER. Pharmaceutical modulation of the glutaminolysis pathway should be considered to control Th17-mediated pathology.

Abstract-Matrix metalloproteinases (MMPs) have been implicated in the process of neovascularization. However, the exact roles of individual MMPs in vessel formation are poorly understood. To study the putative role of MMP-2 in ischemia-induced neovascularization, a hindlimb ischemia model was applied to MMP-2 ϩ/ϩ and MMP-2 Ϫ/Ϫ mice. Serial laser Doppler blood-flow analysis revealed that the recovery of the ischemic/normal blood-flow ratio in MMP-2 Ϫ/Ϫ young and old mice remained impaired throughout the follow-up period. At day 35, microangiography and anti-L-lectin immunohistochemical staining revealed lesser developed collateral vessels and capillary formation in both old and young MMP-2 Ϫ/Ϫ mice compared with the age-matched MMP-2 ϩ/ϩ mice. An aortic-ring culture assay showed a markedly impaired angiogenic response in MMP-2 Ϫ/Ϫ mice, which was partially recovered by supplementation of the culture medium with recombinant MMP-2. Aorta-derived endothelial cells or bone marrow-derived endothelial progenitor cell (EPC)-like c-Kit ϩ cells from MMP-2 Ϫ/Ϫ showed marked impairment of invasive or/and proliferative abilities. At day 7, plasma and ischemic tissues of vascular endothelial growth factor protein were reduced in MMP-2 Ϫ/Ϫ . Flow cytometry showed that the numbers of EPC-like CD31 ϩ c-Kit ϩ cells in peripheral blood markedly decreased in MMP-2-deficient mice. Transplantation of bone marrow-derived mononuclear cells from MMP-2 ϩ/ϩ mice restored neovascularization in MMP-2 Ϫ/Ϫ young mice. These data suggest that MMP-2 deficiency impairs ischemia-induced neovascularization through a reduction of endothelial cell and EPC invasive and/or proliferative activities and EPC mobilization. (Circ Res. 2007;100:904-913.) Key Words: ischemia Ⅲ angiogenesis Ⅲ matrix metalloproteinase Ⅲ endothelium Ⅲ mobilization Ⅲ migration I t is well known that the process of new blood vessel formation is associated with extracellular matrix (ECM) remodeling involving various proteolytic systems. Among such systems, matrix metalloproteinases (MMPs) are a family of zincdependent endopeptidases comprising at least 20 members that are collectively capable of degrading all known ECM components. 1,2 A number of studies have shown that various kinds of MMPs were upregulated in ischemia-induced angiogenesis. 3 Although MMP activity is commonly thought to be involved in the process of angiogenesis, this notion has been challenged by recent studies using genetic or biological target methods. It has been reported that MMP-9 deficiency reduced neovascularization and tumor growth. 4 Study of membrane-type1 (MT1)-MMP knockout mice revealed that the deficiency impaired neovascularization in a mouse corneal micropocket model. 5 Whereas MMP-1 and MMP-10 appear to control the process of vascular regression rather than morphogenesis. 6 On the other hand, certain MMPs, including MMP-12 and MMP-7, are capable of converting plasminogen into angiostatin to inhibit endothelial cell (EC) tubulogenesis in vitro. 7 Interestingly, it has been reported that tiss...

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Podocyte malfunction occurs in autoimmune and nonautoimmune kidney disease. Calcium signaling is essential for podocyte injury, but the role of Ca2+/calmodulin-dependent kinase (CaMK) signaling in podocytes has not been fully explored. We report that podocytes from patients with lupus nephritis and focal segmental glomerulosclerosis and lupus-prone and lipopolysaccharide- or adriamycin-treated mice display increased expression of CaMK IV (CaMK4), but not CaMK2. Mechanistically, CaMK4 modulated podocyte motility by altering the expression of the GTPases Rac1 and RhoA and suppressed the expression of nephrin, synaptopodin, and actin fibers in podocytes. In addition, it phosphorylated the scaffold protein 14-3-3β, which resulted in the release and degradation of synaptopodin. Targeted delivery of a CaMK4 inhibitor to podocytes preserved their ultrastructure, averted immune complex deposition and crescent formation, and suppressed proteinuria in lupus-prone mice and proteinuria in mice exposed to lipopolysaccharide-induced podocyte injury by preserving nephrin/synaptopodin expression. In animals exposed to adriamycin, podocyte-specific delivery of a CaMK4 inhibitor prevented and reversed podocyte injury and renal disease. We conclude that CaMK4 is pivotal in immune and nonimmune podocyte injury and that its targeted cell-specific inhibition preserves podocyte structure and function and should have therapeutic value in lupus nephritis and podocytopathies, including focal segmental glomerulosclerosis.

Aim: Polypharmacy, which is often observed in elderly patients, has been associated with several unfavorable outcomes, including an increased risk of potentially inappropriate medications, medication non-adherence, drug duplication, drugdrug interactions, higher healthcare costs and adverse drug reactions. A significant association between polypharmacy and adverse outcomes among older people living in the community has also been confirmed. A reduction in the number of medications should thus be pursued for many older individuals. Nevertheless, the factors associated with polypharmacy in elderly home-care patients have not been reported. Here, we investigated those factors in elderly home-care patients in Japan.Methods: We used the data of the participants in the Observational Study of Nagoya Elderly with Home Medical investigation. Polypharmacy was defined as the current use of six or more different medications. We carried out univariate and multivariate logistic regression analyses to assess the associations between polypharmacy and each of several factors.Results: A total of 153 home-care patients were registered. The mean number of medications used per patient was 5.9, and 51.5% of the patients belonged to the polypharmacy group. The multivariate model showed that the patients' scores on the Charlson Comorbidity Index and the Mini-Nutrition Assessment Short Form were inversely associated with polypharmacy, and potentially inappropriate medication was most strongly associated with polypharmacy (odds ratio 4.992). Conclusions:The present findings showed that polypharmacy was quite common among the elderly home-care patients, and they suggest that home-care physicians should prescribe fewer medications in accord with the deterioration of home-care patients' general condition. Geriatr Gerontol Int 2017; ••: ••-••.

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Th17 cells favor glycolytic metabolism, and pyruvate dehydrogenase (PDH) is the key bifurcation enzyme, which in its active dephosphorylated form advances the oxidative phosphorylation from glycolytic pathway. The transcriptional factor, inducible cAMP early repressor/cAMP response element modulator (ICER/CREM), has been shown to be induced in Th17 cells and to be overexpressed in CD4 T cells from the patients with systemic lupus erythematosus (SLE). We found that glycolysis and lactate production in in vitro Th17-polarized T cells was reduced and that the expression of pyruvate dehydrogenase phosphatase catalytic subunit 2 (PDP2), an enzyme that converts the inactive PDH to its active form, and PDH enzyme activity were increased in Th17 cells from ICER/CREM-deficient animals. ICER was found to bind to the promoter and suppress its expression. Furthermore, forced expression of PDP2 in CD4 cells reduced the in vitro Th17 differentiation, whereas shRNA-based suppression of PDP2 expression increased in vitro Th17 differentiation and augmented experimental autoimmune encephalomyelitis. At the translational level, PDP2 expression was decreased in memory Th17 cells from patients with SLE and forced expression of PDP2 in CD4 T cells from lupus-prone MRL/ mice and patients with SLE suppressed Th17 differentiation. These data demonstrate the direct control of energy production during Th17 differentiation in health and disease by the transcription factor ICER/CREM at the PDH metabolism bifurcation level.

Objective Glutaminase 1 (Gls1) is the first enzyme in glutaminolysis. The selective Gls1 inhibitor bis‐2‐(5‐phenylacetamido‐1,3,4‐thiadiazol‐2‐yl)ethyl sulfide (BPTES) suppresses Th17 development and ameliorates experimental autoimmune encephalomyelitis (EAE). The present study was undertaken to investigate whether inhibition of glutaminolysis is beneficial for the treatment of systemic lupus erythematosus (SLE), and the involved mechanisms. Methods MRL/lpr mice were treated with BPTES or vehicle control, and disease activity was examined. Then naive CD4+ T cells from patients with SLE were cultured under Th17‐polarizing conditions with BPTES or vehicle. Furthermore, using newly generated Gls1 conditional‐knockout mice, in vitro Th17 differentiation was examined, and EAE was induced in the mice. Glutaminolysis and glycolysis were measured with an extracellular flux analyzer. The expression of hypoxia‐inducible factor 1α (HIF‐1α) was examined by Western blotting. Results Treatment of MRL/lpr mice with BPTES improved autoimmune pathology in a Th17‐dependent manner. T cells from patients with SLE treated with BPTES displayed decreased Th17 differentiation (P < 0.05). Using the conditional‐knockout mice, we demonstrated that both in vitro Th17 differentiation (P < 0.05) and the development of EAE were dependent on Gls1. Gls1 inhibition reduced glycolysis and the expression of HIF‐1α protein, which induces glycolysis. Conclusion We demonstrated that inhibition of glutaminolysis represents a potential new treatment strategy for patients with SLE and Th17‐related autoimmune diseases. Mechanistically, we have shown that inhibition of glutaminolysis affects the glycolysis pathway by reducing HIF‐1α protein in Th17 cells.