Evidence for a sustained increase in clonal beta-cell basal intracellular Ca2+ levels after incubation in the presence of newly diagnosed Type-1 diabetic patient sera. Possible role in serum-induced inhibition of insulin secretion

Abstract

We have previously reported that newly diagnosed Type-1 diabetic patient sera potently suppressed insulin secretion from a clonal rat pancreatic -cell line (BRIN BD11) but did not alter cell viability. Here, we report that apoptosis in BRIN BD11 cells incubated in various sera types (fetal calf serum (FCS), normal human serum and Type-1 diabetic patient) was virtually undetectable. Although low levels of necrosis were detected, these were not significantly different between cells incubated in sera from different sources. ATP levels were reduced by approximately 30% while nitrite production increased twofold from BRIN BD11 cells incubated for 24 h in the presence of Type-1 diabetic patient sera compared with normal human sera. Additionally, ATP levels were reduced by approximately 40% and DNA fragmentation increased by more than 20-fold in BRIN BD11 cells incubated in FCS in the presence of a pro-inflammatory cytokine cocktail (interleukin-1 , tumour necrosis factor-and interferon-), compared with cells incubated in the absence of cytokines. Nitric oxide production from BRIN BD11 cells was markedly increased (up to 10-fold) irrespective of sera type when the cytokine cocktail was included in the incubation medium.Type-1 diabetic patient sera significantly (P<0·001) raised basal levels of intracellular free Ca 2+ concentration ([Ca 2+ ] i ) in BRIN BD11 cells after a 24-h incubation. The alteration in [Ca 2+ ] i concentration was complement dependent, as removal of the early complement components C1q and C3 resulted in a significant reduction (P<0·01) of sera-induced [Ca 2+ ] i changes. We propose that the mechanism of Type-1 diabetic patient sera-induced inhibition of insulin secretion from clonal -cells may involve complement-stimulated elevation of [Ca 2+ ] i which attenuates the nutrient-induced insulin secretory process possibly by desensitizing the cell to further changes in Ca 2+ .

Evidence that low-grade systemic inflammation can induce islet dysfunction as measured by impaired calcium handling

“…Calcium is a key component of the pathway that couples glucose stimulation with insulin secretion [25], but excess cytosolic calcium can lead to beta-cell dysfunction and cell death [15,16]. Exposure to serum from T1D patients has been shown to elevate basal calcium levels in clonal beta-cells [24] and to promote death in primary beta-cells in a calcium-dependent fashion [23]. Serum levels of proinflammatory cytokines are also elevated in the prediabetic stage of T1D and in first-degree relatives [45].…”

“…In addition, chronic treatment of beta cells from control mice with a combination of cytokines upregulated expression of this channel [22]. Also, exposing beta cells to serum isolated from T1D patients induced apoptosis, but not when the L-type channel blocker nifedipine was included in the media [23,24]. …”

The role of the complement system in metabolic organs and metabolic diseases

“…Interestingly, non-obese diabetic (NOD) mice, which are a model for T1DM, lack C5 because of a 2-base pair deletion in the coding region [40]. In vitro , treatment of a rat pancreatic β-cell line with serum from newly diagnosed T1DM patients inhibited their capacity for insulin secretion [41, 42], a phenomenon that was dependent on C1q and C3, since depletion of these complement components reversed the inhibitory effect of the serum of T1DM patients [41]. Furthermore, complement activation, as assessed by the presence of MAC in the serum, was higher in newly diagnosed patients with T1DM than in control individuals, whereas conditioned medium of isolated rat islet cells treated with sera from T1DM patients displayed increased terminal complement activation when compared to medium from cells treated with control serum [41, 43].…”

The central role of calcium in the effects of cytokines on beta-cell function: Implications for type 1 and type 2 diabetes

“…In addition, chronic treatment of beta-cells from control mice with a combination of cytokines upregulated expression of this channel [112]. Also, exposing beta-cells to serum isolated from T1D patients induced apoptosis, but not when the L-type channel blocker nifedipine was included in the media [113,114]. Hyperglycemia is a known contributing factor to beta-cell loss in the latter stages of both T1D and T2D, and evidence suggests that this pathway is also inhibited by blocking L-type calcium channels [115].…”