Leukocyte extravasation into the pancreatic tissue in transgenic mice expressing interleukin 10 in the islets of Langerhans.

Transgenic expression of interleukin 10 (IL-10) in the islets of Langerhans leads to a pronounced pancreatic inflammation, without inflammation of the islets of Langerhans and without diabetes. A scattered infiltration of macrophages (M pi) precedes localized accumulations of CD4+ and CD8+ T lymphocytes, B lymphocytes, and M pi. This recruitment of inflammatory cells to the pancreas is somewhat surprising, since the biological activities of IL-10 in vitro indicate that IL-10 is a powerful immunosuppressive cytokine. Since endothelial cells play a major role in leukocyte extravasation, we examined if vascular changes and extralymphoid induction of peripheral and mucosal type vascular addressins contributed to IL-10-induced homing of mononuclear cells to the pancreas. The endothelium lining small vessels was highly activated in areas of inflammation, as the endothelial cells became cuboidal, and exhibited increased expression of major histocompatibility complex class II (Ia), intercellular adhesion molecule 1, and von Willebrand Factor. Furthermore, induction of vascular addressins simultaneously with accumulation of mononuclear cells around islets and vessels indicated that the endothelial cells take on the phenotype of differentiated endothelium specialized for leukocyte extravasation. In conclusion, pancreatic inflammation and vascular changes are prominent in IL-10 transgenic mice. We hypothesize that IL-10, in addition to its immuno-inhibitory properties, is a potent recruitment signal for leukocyte migration in vivo. These effects are relevant for in vivo therapeutic applications of IL-10

Production of interleukin 10 by islet cells accelerates immune-mediated destruction of beta cells in nonobese diabetic mice.

The T helper type 2 (Th2) cell product interleukin 10 (IL-10) inhibits the proliferation and function of Th1 lymphocytes and macrophages (M phi). The nonobese diabetic mouse strain (NOD/Shi) develops a M phi and T cell-dependent autoimmune diabetes that closely resembles human insulin-dependent diabetes mellitus (IDDM). The objective of the present study was to explore the consequences of localized production of IL-10 on diabetes development in NOD/Shi mice. Surprisingly, local production of IL-10 accelerated the onset and increased the prevalence of diabetes, since diabetes developed at 5-10 wk of age in 92% of IL-10 positive I-A beta g7/g7, I-E- mice in first (N2) and second (N3) generation backcrosses between IL-10 transgenic BALB/c mice and (NOD/Shi) mice. None of the IL-10 negative major histocompatibility complex-identical littermates were diabetic at this age. Furthermore, diabetes developed in 33% of I-A beta g7/d, I-E+ N3 mice in the presence of IL-10 before the mice were 10 wk old. Our findings support the notion that IL-10 should not simply be regarded as an immunoinhibitory cytokine, since it possesses powerful, immunostimulatory properties as well. Furthermore, our observations suggest that beta cell destruction in NOD mice may be a Th2-mediated event

Myasthenia gravis-like syndrome induced by expression of interferon gamma in the neuromuscular junction.

Abnormal humoral responses toward motor end plate constituents in muscle induce myasthenia gravis (MG). To study the etiology of this disease, and whether it could be induced by host defense molecules, we examined the consequences of interferon (IFN) gamma production within the neuromuscular junction of transgenic mice. The transgenic mice exhibited gradually increasing muscular weakness, flaccid paralysis, and functional disruption of the neuromuscular junction that was reversed after administration of an inhibitor of acetylcholinesterase, features which are strikingly similar to human MG. Furthermore, histological examination revealed infiltration of mononuclear cells and autoantibody deposition at motor end plates. Immunoprecipitation analysis indicated that a previously unidentified 87-kD target antigen was recognized by sera from transgenic mice and also by sera from the majority of human MG patients studied. These results suggest that expression of IFN-gamma at motor end plates provokes an autoimmune humoral response, similar to human MG, thus linking the expression of this factor with development of this disease

SLC30A3 Responds to Glucose- and Zinc Variations in ß-Cells and Is Critical for Insulin Production and In Vivo Glucose-Metabolism During ß-Cell Stress

BACKGROUND:Ion transporters of the Slc30A- (ZnT-) family regulate zinc fluxes into sub-cellular compartments. beta-cells depend on zinc for both insulin crystallization and regulation of cell mass. METHODOLOGY/PRINCIPAL FINDINGS:This study examined: the effect of glucose and zinc chelation on ZnT gene and protein levels and apoptosis in beta-cells and pancreatic islets, the effects of ZnT-3 knock-down on insulin secretion in a beta-cell line and ZnT-3 knock-out on glucose metabolism in mice during streptozotocin-induced beta-cell stress. In INS-1E cells 2 mM glucose down-regulated ZnT-3 and up-regulated ZnT-5 expression relative to 5 mM. 16 mM glucose increased ZnT-3 and decreased ZnT-8 expression. Zinc chelation by DEDTC lowered INS-1E insulin content and insulin expression. Furthermore, zinc depletion increased ZnT-3- and decreased ZnT-8 gene expression whereas the amount of ZnT-3 protein in the cells was decreased. Zinc depletion and high glucose induced apoptosis and necrosis in INS-1E cells. The most responsive zinc transporter, ZnT-3, was investigated further; by immunohistochemistry and western blotting ZnT-3 was demonstrated in INS-1E cells. 44% knock-down of ZnT-3 by siRNA transfection in INS-1E cells decreased insulin expression and secretion. Streptozotocin-treated mice had higher glucose levels after ZnT-3 knock-out, particularly in overt diabetic animals. CONCLUSION/SIGNIFICANCE:Zinc transporting proteins in beta-cells respond to variations in glucose and zinc levels. ZnT-3, which is pivotal in the development of cellular changes as also seen in type 2 diabetes (e.g. amyloidosis in Alzheimer's disease) but not previously described in beta-cells, is present in this cell type, up-regulated by glucose in a concentration dependent manner and up-regulated by zinc depletion which by contrast decreased ZnT-3 protein levels. Knock-down of the ZnT-3 gene lowers insulin secretion in vitro and affects in vivo glucose metabolism after streptozotocin treatment

Severe lung fibrosis requires an invasive fibroblast phenotype regulated by hyaluronan and CD44

Hyaluronan synthase 2 and CD44 are required for severe lung fibrosis in response to bleomycin

Restoration of Haemoglobin Level Using Hydrodynamic Gene Therapy with Erythropoietin Does Not Alleviate the Disease Progression in an Anaemic Mouse Model for TGFβ1-Induced Chronic Kidney Disease

<div><p>Erythropoietin, Epo, is a 30.4 kDa glycoprotein hormone produced primarily by the fetal liver and the adult kidney. Epo exerts its haematopoietic effects by stimulating the proliferation and differentiation of erythrocytes with subsequent improved tissue oxygenation. Epo receptors are furthermore expressed in non-haematopoietic tissue and today, Epo is recognised as a cytokine with many pleiotropic effects. We hypothesize that hydrodynamic gene therapy with Epo can restore haemoglobin levels in anaemic transgenic mice and that this will attenuate the extracellular matrix accumulation in the kidneys. The experiment is conducted by hydrodynamic gene transfer of a plasmid encoding murine Epo in a transgenic mouse model that overexpresses TGF-β1 locally in the kidneys. This model develops anaemia due to chronic kidney disease characterised by thickening of the glomerular basement membrane, deposition of mesangial matrix and mild interstitial fibrosis. A group of age matched wildtype littermates are treated accordingly. After a single hydrodynamic administration of plasmid DNA containing murine EPO gene, sustained high haemoglobin levels are observed in both transgenic and wildtype mice from 7.5 ± 0.6 mmol/L to 9.4 ± 1.2 mmol/L and 10.7 ± 0.3 mmol/L to 15.5 ± 0.5 mmol/L, respectively. We did not observe any effects in the thickness of glomerular or tubular basement membrane, on the expression of different collagen types in the kidneys or in kidney function after prolonged treatment with Epo. Thus, Epo treatment in this model of chronic kidney disease normalises haemoglobin levels but has no effect on kidney fibrosis or function.</p></div

Resuscitation with adenocaine and magnesium reduces fluid requirement and improves cardiac function following 72% blood loss in the pig

Introduction: Fluid therapy following severe hemorrhagic shock triggers ischemia-reperfusion (I/R) injury and multi-organ dysfunction. The combination of adenocaine (adenosine and lidocaine), and Mg2+ (ALM) has been shown to stabilize cardiac function and hemodynamics after myocardial I/R. Hypothesis: ALM in 7.5%NaCl at resuscitation: 1) reduces fluid requirements during permissive hypotension and 2) improves post-resuscitation cardiac function. Methods: Pigs (38kg) were randomized to: Sham (n=5), Sham + ALM (n=5), hemorrhage (n=11), and hemorrhage + ALM (n=9). Animals were subjected to pressure controlled hemorrhage at a target MAP of 35mmHg. After 90 min Ringers acetate and 20ml 7.5% NaCl ± ALM (Adenosine 0.23 mg/kg; Lidocaine 0.64 mg/kg; Mg2+ 0.4 mg/kg) was infused to maintain a target MAP of 50 mmHg (permissive hypotension). After 30min 75% of the shed blood was re-infused ± ALM (Adenosine 0.82 mg/kg; Lidocaine 1.66 mg/kg) and the pigs were observed for 6 hrs. Cardiac function was evaluated by pressure-volume (P-V) analysis. Results: In the ALM group 40% less fluid was required to maintain a MAP of 50mmHg for 30min (hemorrhage: 41.5ml/kg CI: 27.7-61.8 vs. ALM: 24.7ml/kg CI:19.4-31.5*). During permissive hypotension 7.5% NaCl ALM significantly increased end-systolic pressure by 27% (hemorrhage: 70 mmHg ± 3 vs. ALM: 89 mmHg ± 9*) and cardiac contractility by 64% (dP/dtmax) (hemorrhage: 2622 mmHg/sec ± 331 vs. ALM: 4301 mmHg/sec ± 262*). Treatment with ALM at blood infusion reduced whole body O2 consumption by 27% lasting 30 min into infusion (group difference p<0.05). In addition, systolic function after blood reinfusion, evaluated by the end-systolic P-V relationship and preload recruitable stroke work, was significantly improved in ALM vs. hemorrhage. Furthermore, ALM enhanced relaxation (dP/dtmin) during reperfusion (hemorrhage: -1578 mmHg/sec ± 103 vs. ALM -2178 mmHg/sec ± 189*). Conclusion: 7.5% NaCl ALM reduced fluid requirements by 40% during permissive hypotension, improved cardiac function and reduced total body O2 consumption following severe hemorrhage. Administration of ALM is effective in minimizing fluid requirements and I/R injury following hemorrhage. (* p<0.05