Deficiency of macrophage migration inhibitory factor (MIF) inhibits cytokine-induced IL-1β generation in murine pancreatic islet cells

In diabetes, pancreatic islets are subjected to high levels of inflammatory mediators that can lead to beta cell destruction. We recently showed that pancreatic islet cells derived from MIF-deficient (MIF-KO) mice are resistant to apoptosis induction by the cytotoxic stimuli of cytokines. Here we show that MIF-KO islets under cytokine (IFN-γ+TNF- α+IL-1β) stimulation express and secrete significantly lower amounts of IL-1β, while the expression of caspase-1 mRNA is not influenced by MIF deficiency. These data suggest that MIF-KO islets possess an innate defect in the process of IL-1 β synthesis and secretion.nul

Effector Mechanisms in Low-Dose Streptozotocin-induced Diabetes

The cellular and molecular requirements for β-cell damages in an immune-mediated toxininduced insulin-dependent diabetes mellitus have been studied in the model of multiple low-dose streptozotocin-induced diabetes in rats and mice. It was found that strain-related susceptibility to diabetes induction correlated with a higher level of IL-2, IFN-γ, and TNF-α production, whereas such differences were not observed when IL-1 and NO production by macrophages were analyzed; elimination of immunoregulatory RT6+T cells that increases IFN-γ production, enhances susceptibility to MLD-STZ-induced diabetes; mercury-induced Th-2 cells downregulated the disease; IFN-γ-mediated macrophage activation to produce proinflammatory cytokines rather than NO is an important event in early diabetogenic effects of invading macrophages; inhibition of IL-1 activity downregulates diabetes induction; and generation of NO in β cells appears to be important for diabetogenic effects. Taken together, data indicate that MLD-STZ diabetes is induced by Th-1 lymphocytes that secrete soluble effector molecules that activate macrophages and promote destruction of β cells possibly by both nitric oxide and nonnitric oxide-mediated mechanisms

The role of macrophage migration inhibitory factor in obesity-associated type 2 diabetes in mice

Macrophage migration inhibitory factor (MIF) is implicated in the pathogenesis of several inflammationrelated diseases, including obesity and type 2 diabetes (T2D). However, MIF deficiency itself promotes obesity and glucose intolerance in mice. Here we show that the introduction of a high-fat diet (HFD) further aggravates the parameters of obesity-associated T2D: weight gain and glucose intolerance. Furthermore, in contrast to MIF-KO mice on standard chow, HFD-fed MIF-KO mice develop insulin resistance. Although the clinical signs of obesity-associated T2D are upgraded, inflammation in MIF-deficient mice on HFD is significantly lower. These results imply that MIF possesses a complex role in glucose metabolism and the development of obesity-related T2D. However, the downregulation of inflammation upon MIF inhibition could be a useful tool in short-term T2D therapy for preventing pancreatic islet deterioration.Projekat ministarstva br. 17301

Deficiency of macrophage migration inhibitory factor (MIF) inhibits cytokine-induced IL-1β generation in murine pancreatic islet cells

In diabetes, pancreatic islets are subjected to high levels of inflammatory mediators that can lead to beta cell destruction. We recently showed that pancreatic islet cells derived from MIF-deficient (MIF-KO) mice are resistant to apoptosis induction by the cytotoxic stimuli of cytokines. Here we show that MIF-KO islets under cytokine (IFN-γ+TNF- α+IL-1β) stimulation express and secrete significantly lower amounts of IL-1β, while the expression of caspase-1 mRNA is not influenced by MIF deficiency. These data suggest that MIF-KO islets possess an innate defect in the process of IL-1 β synthesis and secretion.nul

T cells cooperate with palmitic acid in induction of beta cell apoptosis

<p>Abstract</p> <p>Background</p> <p>Diabetes is characterized by progressive failure of insulin producing beta cells. It is well known that both saturated fatty acids and various products of immune cells can contribute to the reduction of beta cell viability and functionality during diabetes pathogenesis. However, their joint action on beta cells has not been investigated, so far. Therefore, we explored the possibility that leukocytes and saturated fatty acids cooperate in beta cell destruction.</p> <p>Results</p> <p>Rat pancreatic islets or insulinoma cells (RIN) were co-cultivated with concanavalin A (ConA)-stimulated rat lymph node cells (LNC), or they were treated with cell-free supernatants (Sn) obtained from ConA-stimulated spleen cells or from activated CD3⁺cells, in the absence or presence of palmitic acid (PA). ConA-stimulated LNC or Sn and PA cooperated in inducing caspase-3-dependent RIN cell apoptosis. The observed effect of PA and Sn on RIN cell viability was mediated by p38 mitogen-activated protein kinase (MAPK)-signaling and was achieved through auto-destructive nitric oxide (NO) production. The cooperative effect of Sn was mimicked with the combination of interleukin-1β, interleukin-2, interleukin-6, interleukin-17, interferon-γ and tumor necrosis factor-α.</p> <p>Conclusion</p> <p>These results imply that stimulated T cells produce cytokines that cooperate with saturated free fatty acids in beta cell destruction during diabetes pathogenesis.</p

Control of the of the final stage of immune-mediated diabetes by ISO-1, an antagonist of macrophage migration inhibitory factor

We recently showed that attenuation of inflammatory cytokine MIF with pharmacological inhibitor ISO-1 down-regulates the immune-mediated diabetes in mice. Here we explore the effects of MIF neutralization by ISO-1 on the local inflammatory pathway of the disease. In vivo treatment of mice with ISO-1 inhibited the expression of pro-inflammatory cytokines and iNOS in the pancreatic islets. Moreover, ISO-1 affected in vitro cytokine-induced NO pro­duction by fibroblasts, endothelial cells, insulinoma cells, and pancreatic islets, and rescued β cells from NO-dependent damage. These results suggest regulatory potential of ISO-1 at the level of the pancreas which can preserve the target tissue from autoimmune attack.Nedavno smo pokazali da neutralizacija inflamatornog citokina MIF-a farmakološkim inhibitorom ISO-1 negativno reguliše imunski posredovan dijabetes miševa. U ovom radu ispitivali smo efekte neutralizacije MIF-a pomoću ISO-1 na lokalne inflamatorne puteve bolesti. In vivo tretman miševa pomoću ISO-1 je inhibirao ekspresiju proinflamatornih citokina i inducibilne sintaze azot monoksida u ostrvcima pankreasa. Štaviše, ISO-1 je remetio in vitro produkciju azot monoksida indukovan u citokinima u fibroblastima, ćelijama endotela, insulinomama i pankreasnih ostrvaca i tako sačuvao beta ćelije od oštećenja izazvanih azot monoksidom. Ovi rezultati ukazuju na regulatorni potencijal ISO-1 na nivou ciljnog tkiva koji štiti ciljno tkivo od autoimunog ataka.Projekat ministarstva br. 143029

Pentoxifylline Prevents Autoimmune Mediated Inflammation in Low Dose Streptozotocin Induced Diabetes

Xanthine derivative, pentoxifylline (PTX), has been recently shown to exert a protective effects in certain animal models of autoimmunity, including diabetes in NOD mice. In the present study, the immunomodulatory potential of PTX was investigated in autoimmune diabetes induced by multiple low doses of streptozotocin (MLD-SZ) in genetically susceptible CBA/H mice (tested with 40 mg SZ/kg b.w. for 5 days) and DA rats (tested with 20 mg/kg b.w. for 5 days). In both species, 2 – 3 weeks following the MLD-SZ treatment, sustained hyperglycemia developed, as an outcome of inflammatory reaction with endothelial cell activation and accumulation of mononuclear cells. Although there was no evidence of typical insulitis in early disease development (day 10), in both rats and mice, macrophages, CD4+ and CD8+ cells were present in the islets of Langerhans as diffuse mononuclear infiltrates with the expression of IFN-γ and inducible NO synthase (iNOS). Administration of PTX (200 mg/kg/day for 10 days) in combination with MLD-SZ reduced insulitis and the production of mediators tested, and prevented the development of hyperglycemia. These results suggest that beneficial effects of PTX involve down-regulation of local proinflammatory cytokine-mediated NO synthase pathway. They also demonstrate that in addition to ameliorating spontaneous autoimmunity in NOD mice, PTX may be effective in downregulating an inflammatory autoimmune process triggered in susceptible host by an external agents, such as streptozotocin

Effect of interleukin-17 on nitric oxide production in murine fibroblast-like cell line L929

The ability of interleukin-17 (IL-17) to induce nitric oxide (NO) synthesis in murine L929 fibroblasts was investigated. L929 cells were incubated with IL-17 and/or LPS, interferon-g (IFN-gama), interleukin-1 (IL-1), or dibutyryl-cAMP. NO production was assessed indirectly, by measuring nitrite accumulation in 24 h culture supernatants. L929 fibroblasts did not produce NO constitutively, nor IL-17 alone induced NO production. Of all other stimuli tested, only IFN-gama significantly up regulated nitrite level in L929 cell cultures. However, when IL-17 was applied simultaneously with each of the tested stimuli, NO synthesis was markedly elevated, thus indicating involvement of distinct signaling pathways of NO induction by IL-17 and other tested agents. Production of NO by IL-17+LPS-treated L929 cells was dependent on synthesis and activity of inducible NO synthase (iNOS), as indicated by abrogation of nitrite accumulation with protein synthesis inhibitor cycloheximide or selective inhibitor of iNOS, aminoguanidine. A role for protein tyrosine kinase (PTK) and transcription factor NF-kB in iNOS activation is suggested by reduction of NO synthesis with PTK inhibitor genistein and an inhibitor of NF-kB activation, pyrrolidine dithiocarbamate (PDTC). In contrast, protein kinase C inhibitor staurosporine was ineffective in blocking IL-17+LPS-induced NO production in L929 cells. Taken together, our results for the first time showed an active participation of IL-17 in co-induction of fibroblast NO synthesis with LPS, cytokines (IL-1, IFN-gama), or cAMP analogue, and suggested that IL-17 up-regulated NO synthesis in fibroblasts through mechanisms involving PTK and NF-kB activation.Ispitan je uticaj interleukina-17 (IL-17) na produkciju azot monoksida (NO) od strane L929 fibroblasta. L929 ćelije su inkubirane sa IL-17 i/ili LPS-om, interferonom-gama (IFN-gama), interleukinom-1 (IL-1), ili dibutiril-cAMP-om. Produkcija NO ispitana je indirektno, merenjem koncentracije nitrita akumuliranih nakon 24 h u supernatantu ćelijskih kultura. Kultivisani L929 fibroblasti nisu produkovali NO konstitutivno, niti je sam IL-17 indukovao NO produkciju. Od ostalih ispitanih agenasa samo je IFN-gama značajno povećao nivo nitrita u kulturama L929 ćelija. Međutim, kada je IL-17 primenjen sa bilo kojim od testiranih agenasa, došlo je do snažnog povećanja NO sinteze, što je ukazalo da su signalni putevi indukcije NO sa IL-17 i ostalim navedenim agensima različiti. Sprečavanje akumulacije nitrita inhibitorom proteinske sinteze cikloheksimidom ili selektivnim inhibitorom iNOS aminoguanidinom u ćelijama stimulisanim sa IL-17+LPS ukazalo je da NO produkcija zavisi od sinteze i aktivnosti enzima inducibilne NO sinteze (iNOS). Inhibitor protein tirozin kinaze (PTK), genistein i inhibitor aktivacije transkripcionog faktora NF-kB, pirolidin ditiokarbamat (PDTC), takođe su redukovali sintezu NO. Nasuprot tome, inhibitor protein kinaze C, staurosporin, nije blokirao NO produkciju L929 ćelija stimulisanih sa IL-17+LPS. U celini, naši rezultati su po prvi put pokazali aktivno učešće IL-17 u koindukciji NO sinteze sa LPS-om, citokinima (IL-1, IFN-gama), ili cAMP analogom u fibroblastima i ukazali da stimulacija NO sinteze sa IL-17 uključuje mehanizme zavisne od PTK i NF-kB.nul

Control of the of the final stage of immune-mediated diabetes by ISO-1, an antagonist of macrophage migration inhibitory factor

We recently showed that attenuation of inflammatory cytokine MIF with pharmacological inhibitor ISO-1 down-regulates the immune-mediated diabetes in mice. Here we explore the effects of MIF neutralization by ISO-1 on the local inflammatory pathway of the disease. In vivo treatment of mice with ISO-1 inhibited the expression of pro-inflammatory cytokines and iNOS in the pancreatic islets. Moreover, ISO-1 affected in vitro cytokine-induced NO pro­duction by fibroblasts, endothelial cells, insulinoma cells, and pancreatic islets, and rescued β cells from NO-dependent damage. These results suggest regulatory potential of ISO-1 at the level of the pancreas which can preserve the target tissue from autoimmune attack.Nedavno smo pokazali da neutralizacija inflamatornog citokina MIF-a farmakološkim inhibitorom ISO-1 negativno reguliše imunski posredovan dijabetes miševa. U ovom radu ispitivali smo efekte neutralizacije MIF-a pomoću ISO-1 na lokalne inflamatorne puteve bolesti. In vivo tretman miševa pomoću ISO-1 je inhibirao ekspresiju proinflamatornih citokina i inducibilne sintaze azot monoksida u ostrvcima pankreasa. Štaviše, ISO-1 je remetio in vitro produkciju azot monoksida indukovan u citokinima u fibroblastima, ćelijama endotela, insulinomama i pankreasnih ostrvaca i tako sačuvao beta ćelije od oštećenja izazvanih azot monoksidom. Ovi rezultati ukazuju na regulatorni potencijal ISO-1 na nivou ciljnog tkiva koji štiti ciljno tkivo od autoimunog ataka.Projekat ministarstva br. 143029

Protective effects of carbonyl iron against multiple low-dose streptozotocin-induced diabetes in rodents.

Particulate adjuvants have shown increasing promise as effective, safe, and durable agents for the stimulation of immunity, or alternatively, the suppression of autoimmunity. Here we examined the potential of the adjuvant carbonyl iron (CI) for the modulation of organ-specific autoimmune disease-type 1 diabetes (T1D). T1D was induced by multiple low doses of streptozotocin (MLDS) that initiates beta cell death and triggers immune cell infiltration into the pancreatic islets. The results of this study indicate that the single in vivo application of CI to MLDS-treated DA rats, CBA/H mice, or C57BL/6 mice successfully counteracted the development of insulitis and hyperglycemia. The protective action was obtained either when CI was applied 7 days before, simultaneously with the first dose of streptozotocin, or 1 day after MLDS treatment. Ex vivo cell analysis of C57BL/6 mice showed that CI treatment reduced the proportion of proinflammatory F4/80+ CD40+ M1 macrophages and activated T lymphocytes in the spleen. Moreover, the treatment down-regulated the number of inflammatory CD4+ IFN-γ+ cells in pancreatic lymph nodes, Peyer's patches, and pancreas-infiltrating mononuclear cells, while simultaneously potentiating proportion of CD4+ IL17+ cells. The regulatory arm of the immune system represented by CD3+ NK1.1+ (NKT) and CD4+ CD25+ FoxP3+ regulatory T cells was potentiated after CI treatment. In vitro analysis showed that CI down-regulated CD40 and CD80 expression on dendritic cells thus probably interfering with their antigen-presenting ability. In conclusion, particulate adjuvant CI seems to suppress the activation of the innate immune response, which further affects the adaptive immune response directed toward pancreatic beta cells