Modulation of surface TNF expression by human leukaemic cells alters their sensitivity to exogenous TNF.

In this study, U937 leukaemic cells underwent apoptotic cell death following exposure to TNF. Pre-incubation of cells for 48 h with VitD(3) (10(-8)M) induced resistance to TNF, whereas incubation with tau-IFN or GM-CSF increased susceptibility to TNF. Resistance to exogenous TNF (exTNF) following culture with VitD(3) was associated with increased expression of endogenous TNF (enTNF). The TNF inhibitors pentoxifylline(PTF) and dichloroisocoumarin (DCI) inhibited TNF synthesis by U937 cells and abrogated the increase in resistance to TNF seen with VitD(3). The tau-IFN increased TNF expression, whereas GM-CSF had little effect. The data show that the sensitivity of leukaemic cells to exTNF can be modulated by cytokines. The protective effect of VitD(3) is mediated in part by directly upregulating enTNF synthesis.

The effects of islet cell surface antibodies on the metabolic function of mouse islet B cells in vitro.

Immunoglobulin (Ig) fractions from the plasma of a group of newly diagnosed insulin-dependent diabetes mellitus (type 1) patients and set of control subjects were assessed for their effects on isolated mouse islet function. It was found that Igs from type 1 patients caused a significant inhibitory effect on insulin secretion when incubated with mouse islets as compared with controls (25.6 +/- 2.9 pg islet-1 h-1 vs 44.7 +/- 7.7 pg islet-1 h-1, P < 0.05). The plasma samples from which the Igs were obtained were then tested for the presence of antibodies to the mouse islet cell surface (ICSA). Four of the nine patients were positive for ICSA, and plasma samples from eight control subjects were all negative. ICSA-positive samples appeared to have the greatest inhibitory effect on insulin secretion when compared with their respective controls (53.3 +/- 7.0 pg insulin islet -1 min-1 vs 30.9 +/- 3.7 pg insulin islet -1 min-1, (P < 0.05). In contrast, it was also found that ICSA-positive Ig fractions had no significant effect on glucose oxidation when co-incubated with mouse islets as compared with the controls (11.3 +/- 2.3 pmol islet-1 h-1 vs 11.2 +/- 2.9 pmol islet-1 h-1). These studies suggest that Igs from newly diagnosed type 1 patients containing ICSA may impair insulin secretion from isolated mouse islets by mechanisms which do not involve the inhibition of B-cell glucose metabolism.

Development of a rapid latex enhanced turbidimetric assay for retinol binding protein in urine.

We describe a simple, rapid and sensitive homogeneous immunoassay for urinary retinol-binding protein (RBP) using latex particle-enhanced turbidimetric immunoassay. Rabbit anti-human RBP is covalently coupled to 40 nm latex particles and the assay performed on the IL Monarch 2000 centrifugal analyser, with a 20 microL sample volume and the reaction monitored at 340 nm over an 8 min period. The assay range is 0-6 mg/L with a detection limit of 25 micrograms/L. The within and between assay coefficients of variation are less than 1.5% and less than 2.5%, respectively. Comparison with radioimmunoassay for RBP showed good agreement.

Coxsackie B4 viruses with the potential to damage beta cells of the islets are present in clinical isolates.

Infections with Coxsackie viruses (especially Coxsackie B4) are thought to be involved in the pathogenesis of diabetes. Many interdependent variables determine the outcome of an infection with a Coxsackie virus, one of them being the tropism of the virus for a specific tissue. The extent to which Beta cell tropic variants of Coxsackie B4 virus occur naturally was assessed. Human isolates of this virus were tested in an in vitro system in which elevated insulin release from infected islets incubated at a non-stimulatory (2 mmol/l) glucose concentration appears to be related to viral attack. Using this technique, 8/24 isolates tested, impaired secretory function in mouse islets. Some strains of Coxsackie B4 virus, therefore, will directly infect mouse islets in vitro leading to changes in islet cell function. In conclusion, these findings confirm that variants of Coxsackie B4 virus with the potential to damage Beta cells occur quite frequently in the natural population. In certain circumstances the damage they inflict on Beta cells may cause destruction of these cells, or precipitate overt diabetes.

In vivo infection of mice with Coxsackie B4 virus induces long-term functional changes in pancreatic islets with minimal alteration in blood glucose.

The long-term effects of Coxsackie B4 (CB4) infection of mice on pancreatic islet function were investigated. Mice were inoculated with various strains of CB4 virus and 2, 3 and 6 months later islet insulin synthesis and release from isolated islets were measured. Insulin release at basal glucose concentration (2 mmol l-1) was higher in islets from mice inoculated with pancreas-adapted CB4 strains than in control islets or those from mice inoculated with tissue culture-adapted CB4. Thus, two strains of pancreas-adapted virus (P11 and P12) increased basal insulin release by 72% compared with control islets (p less than 0.05) 1 month after inoculation. Another strain (P13) increased insulin release by 421% at 3 months post-inoculation (p less than 0.01) and by 192% at 6 months (p less than 0.05) compared with control islets. The rate of total protein synthesis in islets from P11-inoculated mice 1 month later was 61% lower than in control islets at basal glucose levels (p less than 0.001), and was 25% lower at 20 mmol l-1 glucose (p less than 0.01). There were no significant changes in protein synthesis in islets from infected mice at 3 or 6 months. The abnormal insulin release occurred with minimal changes in random blood glucose concentrations. Histologically the islets were unchanged and there were no detectable islet cell antibodies. These results show that CB4 infection may lead to a persistent metabolic dysfunction in islets with minimal changes in blood glucose levels.