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Milk fat globule-epidermal growth factor-factor VIII downregulates interleukin-17 expression in sepsis by modulating STAT3 activation.

ABSTRACT: Milk fat globule-epidermal growth factor-factor VIII (MFG-E8) is a secretory glycoprotein with a known role in inflammation. In sepsis, interleukin (IL)-17 acts as a proinflammatory cytokine to exaggerate systemic inflammation. We hypothesize that MFG-E8 downregulates IL-17 expression in sepsis.Sepsis was induced in 8-week-old male C57BL/6 mice by cecal ligation and puncture (CLP). Recombinant mouse MFG-E8 (rmMFG-E8) at a dosage of 20 ?g/kg body weight or phosphate-buffered saline was concurrently injected. After 10 hours, blood and spleen samples were harvested for analysis. For in vitro studies, splenocytes isolated from healthy mice pretreated with rmMFG-E8 and splenocytes from MFG-E8 knockout (mfge8(-/-)) mice were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin, followed by measurement of IL-17 expression with either quantitative PCR or enzyme-linked immunosorbent assay.At 10 hours after CLP, rmMFG-E8 inhibited the elevated levels of IL-17 protein in serum by 31%, compared with the vehicle. In the spleen, rmMFG-E8 reduced the upregulated IL-17 mRNA and protein levels by 81% and 51%, respectively. This correlated with a significant reduction in organ injury markers AST and ALT in sepsis after administration of rmMFG-E8. In vitro treatment of splenocytes isolated from healthy mice with rmMFG-E8 showed significant downregulation in PMA/ionomycin-induced IL-17 expression. In contrast, CD4 T-cells from mfge8(-/-) mice showed significant upregulation of IL-17 compared with wild-type mice. The phosphorylated level of signal transducer and activator of transcription 3 (STAT3) was downregulated in spleen tissue of septic mice treated with rmMFG-E8. Conversely, mfge8(-/-) mice showed increased phosphorylated STAT3 compared with wild-type mice after sepsis.Our findings demonstrate MFG-E8-mediated downregulation of IL-17 expression, implicating its potential as a novel therapeutic agent against sepsis.

SUBMITTER: Cen C

PROVIDER: S-EPMC4707120 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Milk fat globule-epidermal growth factor-factor VIII downregulates interleukin-17 expression in sepsis by modulating STAT3 activation.

Cen Cindy C Aziz Monowar M Yang Weng-Lang WL Nicastro Jeffrey J Coppa Gene F GF Wang Ping P

Surgery 20150914 2

<h4>Background</h4>Milk fat globule-epidermal growth factor-factor VIII (MFG-E8) is a secretory glycoprotein with a known role in inflammation. In sepsis, interleukin (IL)-17 acts as a proinflammatory cytokine to exaggerate systemic inflammation. We hypothesize that MFG-E8 downregulates IL-17 expression in sepsis.<h4>Methods</h4>Sepsis was induced in 8-week-old male C57BL/6 mice by cecal ligation and puncture (CLP). Recombinant mouse MFG-E8 (rmMFG-E8) at a dosage of 20 μg/kg body weight or phosp ...[more]

PMID: 26376757

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Project description:BACKGROUND:Acute kidney injury (AKI) is most commonly caused by sepsis in critically ill patients, and it is associated with high morbidity and mortality. The pathophysiology of sepsis-induced AKI is generally accepted to include direct inflammatory injury, endothelial cell dysfunction, and apoptosis. Milk fat globule-epidermal growth factor-factor VIII (MFG-E8) is a secretory glycoprotein with a known role in the enhancement of apoptotic cell clearance and regulation of inflammation. We hypothesize that administration of recombinant mouse MFG-E8 (rmMFG-E8) can protect mice from kidney injuries caused by sepsis. METHODS:Sepsis was induced in 8-wk-old male C57BL/6 mice by cecal ligation and puncture (CLP). rmMFG-E8 or phosphate-buffered saline (vehicle) was injected intravenously at a dosage of 20 ?g/kg body weight at time of CLP (n = 5-8 mice per group). After 20 h, serum and renal tissue were harvested for various analyses. The renal injury markers blood urea nitrogen (BUN) and creatinine were determined by enzymatic and chemical reactions, respectively. The gene expression analysis was carried out by real-time quantitative polymerase chain reaction. RESULTS:At 20 h after CLP, serum levels of BUN and creatinine were both significantly increased in the vehicle group compared with the sham group, whereas the mice treated with rmMFG-E8 had a significant reduction in BUN and creatinine levels by 28% and 24.1%, respectively (BUN: 197.7 ± 23.6 versus 142.3 ± 20.7 mg/dL; creatinine: 0.83 ± 0.12 versus 0.63 ± 0.06 mg/dL; P < 0.05). Expressions of novel biomarkers of renal tissue injury neutrophil gelatinase-associated lipocalin and kidney injury molecule-1 were also significantly downregulated by 58.2% and 95%, respectively, after treatment with rmMFG-E8. Proinflammatory cytokine interleukin-6 and tumor necrosis factor-? messenger RNA (mRNA) were significantly reduced by 50.8% and 50.3%, respectively, in rmMFG-E8-treated mice compared with vehicle-treated mice. The mRNA levels of the chemokines keratinocyte chemoattractant and macrophage inhibitory protein-2 were reduced by 85.1% and 78%, respectively, in mice treated with rmMFG-E8 compared with the vehicle mice. In addition, the expression of intercellular cell adhesion molecule-1 and platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) mRNA was downregulated by 35.6% and 77.8%, respectively, in rmMFG-E8-treated mice compared with the vehicle animals (P < 0.05). CONCLUSIONS:Treatment with rmMFG-E8 reduces renal tissue injury induced by sepsis through inhibiting the production of proinflammatory cytokines and chemokine, as well as through the activation of endothelial cells. Thus, MFG-E8 may have a therapeutic potential for treating AKI induced by sepsis.

Project description:ObjectivesRenal ischemia-reperfusion injury causes acute renal failure, and the hallmarks of renal ischemia-reperfusion injury are inflammation, apoptosis, necrosis, and capillary dysfunction. Milk fat globule-epidermal growth factor-factor VIII (MFG-E8), a membrane-associated secretory glycoprotein, is produced by immune cells and reported to participate in multiple physiologic processes associated with tissue remodeling. We have recently shown that MFG-E8 treatment attenuates organ injury, inflammatory responses, and survival after sepsis through the enhancement of phagocytosis of apoptotic cells. The purpose of this study was to determine whether administration of MFG-E8 attenuates renal ischemia-reperfusion injury.DesignProspective, controlled, and randomized animal study.Setting: A research institute laboratory.SubjectsMale C57BL/6J mice (20-25 g).Interventions: Renal ischemia-reperfusion injury with bilateral renal pedicle clamping for 45 mins, followed by reperfusion. A recombinant murine MFG-E8 (0.4 μg/20 g) was given intraperitoneally at the beginning of reperfusion.Measurements and main resultsMFG-E8 levels, organ injury variables, inflammatory responses, histology, apoptosis, and capillary functions were assessed at 1.5 and 20 hrs after reperfusion. A 60-hr survival study was conducted in MFG-E8 and recombinant murine MFG-E8-treated wild-type mice. After renal ischemia-reperfusion injury, MFG-E8 mRNA and protein expressions were significantly decreased in the kidneys and spleen. Treatment with recombinant murine MFG-E8 recovered renal dysfunction, significantly suppressed inflammatory responses, apoptosis, necrosis, and improved capillary functions in the kidneys. In the survival study, MFG-E8 mice showed a significant deterioration and, in contrast, recombinant murine MFG-E8-treated wild-type mice showed a significant improvement of survival compared with vehicle-treated wild-type mice.ConclusionsMFG-E8 can be developed as novel treatment for renal ischemia-reperfusion injury. This protective effect appears to be mediated through the enhancement of apoptotic cell clearance and improvement of capillary functions in the kidneys.

Project description:Aims/introductionMilk fat globule-epidermal growth factor 8 (MFG-E8) is the key mediator in anti-inflammatory responses that facilitate phagocytosis of apoptotic cells, and play an essential role in type 2 diabetes and pregnancy, both of which are under a low-grade inflammatory state. However, the action of MFG-E8 in gestational diabetes mellitus (GDM) is unclear. We measured plasma MFG-E8 levels in pregnancy and GDM for the first time, and elucidated possible relationships between its plasma levels and various metabolic parameters.Materials and methodsPlasma MFG-E8 levels were quantified by enzyme-linked immunosorbent assay in 66 women with GDM, 70 with normal pregnancy (p-NGT) and 44 healthy non-pregnant controls (CON), who were matched for age and body mass index. Inflammatory factors tumor necrosis factor-? (TNF-?) and C-reactive protein levels were measured, oral glucose tolerance test was carried out and ?-cell function was evaluated.ResultsPlasma MFG-E8 levels were remarkably higher in p-NGT than in CON (P = 0.024), and were further elevated in GDM vs p-NGT (P = 0.016). MFG-E8 concentrations correlated positively with hemoglobin A1c, glucose levels and insulin resistance (homeostasis model assessment for insulin resistance), and correlated inversely with TNF-? and insulin secretion evaluated by disposition indices in pregnancies. Fasting glucose levels, disposition index of first phase insulin secretion and TNF-? were independent predictors of MFG-E8 levels in pregnancies. Logistic regression analyses showed that women in the third tertile of MFG-E8 levels had a markedly elevated risk of GDM.ConclusionsCirculating MFG-E8 levels are dramatically elevated in pregnancy, and are significantly higher in GDM vs p-NGT. MFG-E8 concentrations are significantly associated with TNF-?, fasting glucose levels, homeostasis model assessment for insulin resistance and disposition indices. However, further studies are required to elucidate the regulation mechanism of MFG-E8 during pregnancy and GDM.

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Milk fat globule-epidermal growth factor-factor VIII downregulates interleukin-17 expression in sepsis by modulating STAT3 activation.

Publications

Milk fat globule-epidermal growth factor-factor VIII downregulates interleukin-17 expression in sepsis by modulating STAT3 activation.

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