Extensive study of human insulin immunoassays: promises and pitfalls for insulin analogue detection and quantification:

BACKGROUND: Over the last few decades, new synthetic insulin analogues have been developed. Their measurement is of prime importance in the investigation of hypoglycaemia, but their quantification is hampered by variable cross-reactivity with many insulin assays. For clinical analysis, it has now become essential to know the potential cross-reactivity of analogues of interest. METHODS: In this work, we performed an extensive study of insulin analogue cross-reactivity using numerous human insulin immunoassays. We investigated the cross-reactivity of five analogues (lispro, aspart, glulisine, glargine, detemir) and two glargine metabolites (M1 and M2) with 16 commercial human insulin immunoassays as a function of concentration. RESULTS: The cross-reactivity values for insulin analogues or glargine metabolites ranged from 0% to 264%. Four assays were more specific to human insulin, resulting in negligible cross-reactivity with the analogues. However, none of the 16 assays was completely free of cross-reactivity with analogues or metabolites. The results show that analogue cross-reactivity, which varies to a large degree, is far from negligible, and should not be overlooked in clinical investigations. CONCLUSIONS: This study has established the cross-reactivity of five insulin analogues and two glargine metabolites using 16 immunoassays to facilitate the choice of the immunoassay(s) and to provide sensitive and specific analyses in clinical routine or investigation

Improvement of rat islet viability during transplantation: validation of pharmacological approach to induce VEGF overexpression:

Delayed and insufficient revascularization during islet transplantation deprives islets of oxygen and nutrients, resulting in graft failure. Vascular endothelial growth factor (VEGF) could play a critical role in islet revascularization. We aimed to develop pharmacological strategies for VEGF overexpression in pancreatic islets using the iron chelator deferoxamine (DFO), thus avoiding obstacles or safety risks associated with gene therapy. Rat pancreatic islets were infected in vivo using an adenovirus (ADE) encoding human VEGF gene (4.10(8) pfu/pancreas) or were incubated in the presence of DFO (10 mumol/L). In vitro viability, functionality, and the secretion of VEGF were evaluated in islets 1 and 3 days after treatment. Infected islets or islets incubated with DFO were transplanted into the liver of syngenic diabetic rats and the graft efficiency was estimated in vivo by measuring body weight, glycemia, C-peptide secretion, and animal survival over a period of 2 months. DFO induced transient VEGF overexpression over 3 days, whereas infection with ADE resulted in prolonged VEGF overexpression lasting 14 days; however, this was toxic and decreased islet viability and functionality. The in vivo study showed a decrease in rat deaths after the transplantation of islets treated with DFO or ADE compared with the sham and control group. ADE treatment improved body weight and C-peptide levels. Gene therapy and DFO improved metabolic control in diabetic rats after transplantation, but this effect was limited in the presence of DFO. The pharmacological approach is an interesting strategy for improving graft efficiency during transplantation, but this approach needs to be improved with drugs that are more specific

Instant blood-mediated inflammatory reaction during islet transplantation: the role of Toll-like receptors signaling pathways:

The instant blood-mediated inflammatory reaction (IBMIR) leads to massive destruction of transplanted islets. Islet isolation and time of culture may elicit the release of potent activators of Toll-like receptors (TLRs) signaling pathways during IBMIR. This work sought to evaluate the role of TLR signaling pathways to mediate inflammatory reactions. Isolated rat pancreatic islets were cultured for 12, 24, or 48 hours. Their viability was assessed by fluorescein diacetate/propidium iodide and their functionality, by glucose stimulation tests. Endotoxin levels were quantified using the Limulus Amebocyte Lysate assays. After RNA extraction and reverse transcription, we performed polymerase chain reaction (PCR) arrays. Samples obtained immediately after isolation were defined as controls. Eighty-four genes belonging to the TLR signaling pathways, were compared with control samples. After culture, islets were viable and functional with low endotoxin levels (< 0.1 endotoxin units/mL) showed TLR activation not due to exogenous contamination. Analysis of PCR arrays highlighted significant up-regulation of TLR-2. After 24 hours of culture, TLR-2 was up-regulated to 6.8 +/- 0.6-fold (P < .001) compared with controls but decreased to 4.3 +/- 1.4-fold after 48 hours. In the same way, expression of myeloid differentiation primary response gene 88 (Myd88) was significantly up-regulated (3.2 +/- 0.4-fold [P < .001]) compared with controls. After 12 hours of culture, interleukin-10 gene expression was significantly up-regulated at 11.6 +/- 3.7- fold (P < .05), reaching 17.5 +/- 8.3 after 24 hours. Finally, the cyclo-oxygenase-2 gene expression was up-regulated to 509 +/- 67.1-fold (P < .05) after 12 hours of culture. These data confirmed the implication of TLR signaling pathways in early inflammatory events

Gastric Electrical Stimulation Reduces Refractory Vomiting in a Randomized Crossover Trial

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