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

Abstract

The appropriate regulation of intracellular calcium is a requirement for proper cell function and survival. This review focuses on the effects of proinflammatory cytokines on calcium regulation in the insulin-producing pancreatic beta-cell and how normal stimulus-secretion coupling, organelle function, and overall beta-cell viability are impacted. Proinflammatory cytokines are increasingly thought to contribute to beta-cell dysfunction not only in type 1 diabetes (T1D), but also in the progression of type 2 diabetes (T2D). Cytokine-induced disruptions in calcium handling result in reduced insulin release in response to glucose stimulation. Cytokines can alter intracellular calcium levels by depleting calcium from the endoplasmic reticulum (ER) and by increasing calcium influx from the extracellular space. Depleting ER calcium leads to protein misfolding and activation of the ER stress response. Disrupting intracellular calcium may also affect organelles, including the mitochondria and the nucleus. As a chronic condition, cytokine-induced calcium disruptions may lead to beta-cell death in T1D and T2D, although possible protective effects are also discussed. Calcium is thus central to both normal and pathological cell processes. Because the tight regulation of intracellular calcium is crucial to homeostasis, measuring the dynamics of calcium may serve as a good indicator of overall beta-cell function.

1. Overview

In the pancreatic beta-cell, cytokine-induced disruptions in calcium handling can impair insulin release in response to glucose stimulation, and more severe calcium disruptions can lead to cell death. The focus of this review is on calcium’s role in cytokine-mediated dysfunction and death of pancreatic islets and the potential role of key calcium handling organelles: the endoplasmic reticulum, mitochondria, nucleus and cytosolic spaces [3,4]. Detailed descriptions of the molecular mechanisms of cytokine action and their signaling pathways are not included but can be found in several excellent reviews [5-10].

2. Normal islet function in response to glucose stimulation

Islets of Langerhans are micro-organs within the pancreas that are responsible for regulating blood glucose and body energy metabolism [11]. The islet is composed of insulin-producing beta-cells (~60-80% of the total islet mass in rodents), glucagon-secreting alpha-cells (10-30%), somatostatin-secreting delta-cells (5-10%), and others [15,16]. At the level of the individual beta-cell, the ‘Consensus Model’ provides a detailed description of the cellular response to glucose stimulation [17-19]. As shown in Figure 1A (a-e), glucose is taken up through glucose transporters [20,21] and then metabolized in the beta-cell through glycolysis and the tricarboxylic acid (TCA) cycle to increase ATP production (for a more detailed description of these processes, see [22,23]). During this time, calcium levels in the mitochondria and ER increase in response to glucose stimulation, which causes an overall dip in cytosolic calcium as shown in Figure 1B. The resulting increase in the ATP-to-ADP ratio closes ATP-sensitive potassium channels (K_ATP-channels), which initiates a large spike in calcium influx and the first phase of insulin release. Following this initial burst, calcium and insulin release rates remain elevated throughout the second phase response, which continues as long as glucose remains elevated.

Figure 1.

Stimulus-secretion coupling in the pancreatic beta-cell. (A) The processes involved with glucose uptake to insulin release are described in steps a-e. Note that green arrows denote the redistribution of intracellular calcium, and red arrows indicate the targets of calcium influx. (B-C) This process can be recorded in real-time by the changes in calcium (B) and insulin release (C). Panel (C) was modified with permission from the Am Physiol Soc (Nunemaker et al, Am J Physiol-Endo and Metab, 2006 ([197]).

Because calcium is a strong trigger of exocytosis, both glucose-stimulated calcium (GSCa, Figure 1B) and glucose-stimulated insulin secretion (GSIS, Figure 1C) show similar trajectories under these conditions. GSCa can thus be used to assess the physiological response of islets to glucose stimulation. Calcium imaging is advantageous because it provides high temporal precision of real-time changes in response to stimuli at the level of the individual beta-cell. Changes in the latency, trajectory, and amplitude of the triphasic GSCa response may indicate specific defects in stimulus-secretion coupling or other aspects of islet dysfunction. To be complete, amplifying processes also operate in parallel with the pathways of the Consensus Model to couple glucose uptake and metabolism with insulin exocytosis [24-27]. These processes allow additional insulin release to occur independently of changes in calcium. GSCa thus provides a reasonable first approximation of GSIS and can provide a good estimate of overall islet health and function.

3. Cytokines, calcium, and islet dysfunction

Pro-inflammatory cytokines are broadly considered immunomodulators that consist of several families of signaling molecules, including interleukins and interferons. Cytokines play a prominent role in the pathophysiology of type 1 diabetes (T1D) [28-32], but increasing evidence suggests a significant role for cytokines in the loss of beta-cell mass in T2D as well [34,35]. There are notable similarities and differences in the action of cytokines on the beta-cell between T1D versus T2D [35-37]. First, in T1D, beta-cells are the direct target of an autoimmune invasion that results in insulitis and beta-cell death [37]. Whereas, in T2D metabolic stress is thought to activate the innate immune system, resulting in a chronic inflammatory state marked by increased cytokines, increased islet-associated macrophages, and beta-cell apoptosis [37-39]. Second, a number of factors contribute to beta-cell decline and destruction in T2D including glucotoxicity, lipotoxicity, ER stress, oxidative stress, and amyloid deposition [40-46], any of which can additionally trigger an inflammatory response. In fact, evidence is emerging that all of these factors may even be centrally linked with inflammasome activation [47-49]. Third, the most dominant pro-inflammatory cytokines in T1D are tumor necrosis factor alpha (TNF-a), interleukin (IL)-1B, and interferon-gamma (IFN-g). These cytokines are secreted by infiltrating immune cells at high concentrations in close proximity to the beta-cell and act synergistically to inflict direct inhibitory and cytotoxic effects [32,33,50,51]. In T2D, a chronic low-grade inflammatory state is thought to involve lower levels of cytokines that are produced by adipose tissue [52,53], islet-associated macrophages [54,55], and the islets themselves [7].

These differences in disease progression between T1D and T2D result in islet exposure to different extracellular cytokine concentrations, durations of exposure, and cytokine milieu, which can markedly affect cytokine signaling and net effects on the beta-cell. For example, short-term treatment with low-dose IL-1B may improve the function of rodent islets, whereas long-term, high-dose treatment impairs islet function [56-58] and accelerates the development of T1D [59]. Low-dose TNF-a treatment has been shown to inhibit the autoimmune response in models of T1D to slow the disease process [60-62], whereas TNF-a and IFN-g produce dose- and duration-dependent islet dysfunction in vitro when acting in concert [63,64]. In the context of T2D, the duration of low-grade inflammation may be a key factor, as cytokines could have compensatory/protective effects initially that become deleterious under chronic conditions. Differences among duration and dose of cytokine treatment [58,65], species being tested [66-68], and combination of cytokines [32,66,69,70] are thus all important factors that contribute to both positive or negative effects on the beta-cell. In addition, synergistic activity among multiple cytokines can alter or amplify signaling pathways [32,70-72], adding an additional layer of complexity to cytokine action.

A number of proinflammatory cytokines are found at serum levels 2-5x higher in obese individuals compared to lean individuals, and these increased levels are associated with increased risk of developing T2D [73-75]. This low-grade systemic inflammation may also play a direct role in triggering beta-cell dysfunction, particularly in T2D. We have previously reported that cytokines can directly affect calcium handling in rodent islets at circulating concentrations in vitro [76,77]. As shown in Figure 2, we present similar findings comparing islets treated overnight with a ‘low-dose’ cytokine treatment to mimic circulating cytokine levels in rodents (slightly higher than found in human serum) or a ‘high-dose’ cytokine treatment to mimic concentrations associated with direct immune assault in T1D [67,70,78], which is thought to be ~100-1000 fold higher than found in serum. First note that treatment with cytokines dose-dependently elevated the basal level of calcium in low glucose prior to stimulation. Prolonged excess calcium is a well-established trigger of beta-cell death [79]. Following glucose stimulation, the phase 0 response associated with increased calcium uptake by the endoplasmic reticulum was dose-dependently reduced by cytokines. The low-dose cytokine treatment also attenuated phase 1 response without affecting the phase 2 response as measured relative to basal calcium levels. Of interest, loss of first phase insulin secretion without the loss of second phase secretion is an early sign of islet dysfunction in both T1D and T2D [80]. The high-dose cytokine treatment substantially impaired all phases of the calcium response, as well as producing significant cell death in overnight culture (see also [76]). These data suggest that cytokines can harm beta-cells in a graded manner by impairing ER calcium handling and calcium flux across the plasma membrane. We note, however, inasmuch as stimulatory and protective effects have been observed with lower concentrations as discussed above, cytokine action in the beta-cell results in more complex and multi-faceted interactions than understood at present. The remaining sections of this review focus on four major calcium handling organelles (the plasma membrane, the ER, mitochondria, and nucleus) and how each may be involved with cytokine-induced changes in [Ca²⁺]_i.

Figure 2.

(A) Glucose-stimulated [Ca2+]i responses from islets after overnight treatment with “high” (0.5 ng/ml IL1-B + 1 ng/ml TNF-a + 10 ng/ml IFN-g, n=10), “low” (5 pg/ml IL1-B + 10 pg/ml TNF-a + 100 pg/ml IFN-g, n=11), or left untreated (n=18). (B) Dose-dependent cytokine effects were observed in phases 0, 1, and 2. *P<0.01, **P<0.001. These data were collected from islets that were isolated by our published protocol [198] and are in agreement with our published findings as described previously [76,77].

4. Cytokines and cell membrane sources of [Ca²⁺]_i

The influx of calcium through various ion channels and exchangers in the cell membrane provides a key source of [Ca²⁺]_i for the signaling cascades of proinflammatory cytokines [46,79,81,82]. Although the L-type calcium channel is dominant and considered critical to normal islet function, several other subtypes may also be physiologically relevant including the N-type and T-type calcium-channels [83,84]. The subunits that form these various channels can also vary, thus affecting voltage-sensing, Ca2+-conducting channel activation, inactivation, and current amplitude [85,86]. In addition, changes in the expression, regulation, and function of these calcium-channels, as well as the insertion and removal of the channels from the membrane, and the association of the channel with protein complexes in the membrane (the exocytotic machinery for example [87]) can all affect calcium flux into the beta-cell. Further, modulation of other voltage-gated ion channels, including the K_ATP-channel and Kv-channels, will alter the membrane potential to make calcium influx more or less favorable. A number of K+-channels, including BK [88,89], SK [90] and the K_Slow current [91,92] are also calcium-sensitive, and their activity can affect calcium flux via their effects on the membrane potential. Finally, the distribution and movement of calcium upon entering the beta-cell is intricate and crucial to normal islet function [93]. Thus, cytokines and other factors could indirectly regulate calcium flux in many ways. This is particularly pertinent to cytokine action since effects on calcium occur over a period of hours as opposed to the comparatively rapid effects of glucose-stimulated calcium changes on the order of seconds to minutes. No acute cytokine effects on the order of seconds have been observed on calcium-channels to our knowledge.

With regard to cytokines, several studies in the late-1980s provided evidence that cytokine action was linked with calcium influx, suggesting that the cytokine IL-1 may alter calcium flux across the beta-cell membrane to produce beta-cell dysfunction or toxicity [94,95]. Subsequent studies have suggested an important permissive role of calcium influx in cytokine-mediated cell death. For example, blocking L-type calcium channels prevents cell death caused by IL-1B/IFN-g treatment [64] in insulin cell lines and prevents IL-1B-induced apoptosis in rodent [97,98] and human islets [99]. Several signaling pathways related to IL-1B are also dependent on calcium influx through L-type calcium channels, including ERK1/2, p38, and JNK [97,99,100].

Sodium/calcium exchangers (NCXs), plasma-membrane Ca²⁺ATPases (PMCAs), and other calcium regulators may also contribute to cytokine effects. Under normal conditions, NCXs play an active role in calcium homeostasis, accounting for up to 70% of calcium extrusion and also participating in calcium influx in response to membrane depolarization [101]. Overexpression of NCX can deplete ER calcium and induce apoptosis in insulin-secreting BRIN-BD11 cells [101]. Similarly, PMCAs play a key role in calcium homeostasis, with over expression resulting in apoptosis by calcium depletion from both the ER and mitochondria [102]. Cytokines appear to downregulate PMCA expression in RINm5F cells [103], and reduced PMCA expression decreases TNF-induced cell death in L929 cells [104]. Insulin resistance also appears to downregulate PMCA expression and increase [Ca²⁺]_i in rat islets [105]. Collectively, these data present a complex picture regarding the role of NCXs and PMCAs warranting further study. Other ion channels may also influence beta-cell [Ca²⁺]_i [84], although little is known about cytokine sensitivity.

Even without inducing cell death, cytokine-mediated changes in calcium flux can impact beta-cell function in numerous ways. Early studies showed that IL-1 inhibits glucose-stimulated calcium influx [94], but others have shown stimulatory effects of IL-1 on insulin secretion that may depend on diacylglycerol production and protein kinase C activation [106,107] rather than changes in [Ca²⁺]_i [107]. Interleukin-6 (IL-6) has been shown to increase insulin secretion and preproinsulin mRNA expression by mechanisms that rely on calcium influx [108]. A recent study suggests that IL-6 enhances insulin secretion through involvement of the PLC-IP3 pathway [109]. In contrast, numerous studies have also shown inhibitory effects of IL-6 in combination with other inflammatory cytokines [110,111]. We recently reported that the combination of IL-6 and IL-1B at relatively low concentrations can elevate calcium levels in low glucose and attenuate glucose-stimulated calcium responses in mouse islets [77]. Our previous work suggests that calcium influx through L-type calcium channels is at least partly responsible for these cytokine-mediated effects[76]. As mentioned previously, differing effects on islet function may depend upon a variety of factors, chiefly the dose and combination of cytokines used [32,66,67,70].

Calcium influx also appears to be central to beta-cell dysfunction in several models of diabetes. In the NOD mouse model of T1D, increased expression of the low-voltage activated (LVA) T-type calcium channel was observed in beta-cells, resulting in elevated basal [Ca²⁺]_i [112]. 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]. These findings collectively suggest that numerous cytokine-mediated signals require calcium influx for their full effects.

5. Cytokines, ER Calcium homeostasis, and ER Stress

The ER plays an important role in calcium storage and signaling. The resting intra-ER calcium concentration is three to four orders of magnitude higher than cytosolic calcium [46]. When cytoplasmic calcium is high, the ER will sequester calcium, while when it is low, the ER will release calcium [116-118]. ER dysfunction is manifested by deficiencies in normal ER calcium handling due primarily to the changes in the sarco(endo)plasmic reticulum calcium ATP-ase (SERCA2b) pump in insulin-secreting cells [119]. Because ER calcium is required for protein binding and chaperone activity, severe ER calcium depletion will impair the quality of ER protein folding and assembly and cause ER stress [120-123]. Cells attempt to alleviate ER stress by means of the unfolded protein response (UPR), which includes increasing ER chaperone proteins to increase protein folding activity, degrading misfolded proteins, and decreasing protein translation to prevent accumulation of unfolded protein [124,125]. Failure to relieve ER stress or a prolonged UPR may lead to cell death through the activation of apoptotic signals by c-Jun N-terminal kinase (JNK) [124-127], CCAAT/enhancer binding protein homologous protein (CHOP)[124,125,128], Bcl-2-associated X protein (Bax) [125], and nuclear factor kappaB (NFkB) [74,124,126].

There is conflicting evidence as to how proinflammatory cytokines induce ER stress. One hypothesis is that proinflammatory cytokines activate inducible nitric oxide synthase (iNOS) leading to excessive nitric oxide (NO) production [129]. NO then acts as a signal to various intracellular components including the ER. By down-regulation of SERCA2b pump activity, NO causes a decrease or even depletion of ER calcium [130], leading to increased cytosolic calcium concentrations and ER stress [123,131,132]. Conversely, there is also evidence suggesting that cytokines induce ER stress through an undetermined pathway independent of nitric oxide production [133,134].

Non-cytokine induced ER stress and cytokine treatment share many of the same features in terms of disruptions in calcium handling. Similarly to cytokines, thapsigargin, a drug that inhibits SERCA activity, has been shown to deplete ER calcium and lead to ER stress [135]. Not surprisingly, cytokine and thapsigargin treatment both produce similar effects on calcium handling in response to glucose stimulation: (a) elevated basal calcium prior to stimulation, (b) attenuated phase 1 response to glucose stimulation, (c) disruptions in phase 2 oscillations [76,116,136]. However, unlike the SERCA inhibitor, cytokines do not appear to trigger key UPR genes, such as activating transcription factor 6 (ATF-6) and ER chaperone BiP, thereby depriving beta-cells of a mechanism for cell survival during ER stress [132] and promoting apoptosis. Two ER membrane calcium-releasing channels that should be noted as possible regulators of ER stress are inositol triphosphate receptors and ryanodine receptors. Inhibition of these receptors has been shown to alleviate ER stress and apoptosis resulting from impaired SERCA function, whereas activation of the receptors while inhibiting the SERCA pump has been shown to increase ER stress and cell death [137].

With regard to diabetes, emerging evidence suggests that ER stress linked to ER calcium depletion may play a key role in beta-cell failure, at least in T2D. The db/db mouse, a leptin-receptor-deficient model of (T2D) and obesity, has an increased activation of genes related to ER stress [138], and in the db/db mouse, it is evident that ER calcium mishandling plays a role in the onset of T2D. Compared to their controls, db/db islets lack the phase 0 dip in [Ca²⁺]_i and the subsequent [Ca²⁺]_i oscillations following glucose stimulation [76,139,140]. When control islets are treated with thapsigargin, they show basal and glucose-stimulated insulin secretion levels similar to those in db/db islets, suggesting that aberrant ER calcium sequestration underlies the impaired glucose responses in the db/db mouse and may play a role in defective insulin secretion [139]. While there is evidence that human islet ER stress exists for T2D [138], there is little evidence to date for ER stress in T1D in humans [141]. Two pieces of evidence, that NO is less important for cytokine-induced human β-cell death [67], and that islets from mice lacking an inducible iNOS (iNOS −/−) are only partially protected against cytokines [142,143], suggest a non-NO/ER stress cell death pathway in T1D [134,144-146].

6. Mitochondrial calcium and cytokine action

Mitochondrial metabolism and cytosolic calcium have a dynamic and often reciprocal relationship. Nutrient-stimulated changes in metabolism initially hyperpolarize the mitochondrial membrane potential (ΔΨ_m), but increases in [Ca²⁺]_i tend to increase mitochondrial calcium, which depolarizes the ΔΨ_m and can reduce ATP production [147-150]. Mitochondria can thus act as calcium buffers that compensate for temporary increases in [Ca²⁺]_i levels; however, an overload in mitochondrial calcium can lead to dysfunction or downstream activation of cell death pathways [151-153]. Mitochondrial-mediated cell death pathways are initiated by depolarization of the ΔΨ_m, onset of the mitochondrial permeability transition via opening of mitochondrial permeability transition pores causing osmotic changes that lead to mitochondrial swelling, and increased reactive oxygen species (ROS) production [137,151,152,154].

In the beta-cell, damaged or missing portions of mitochondrial DNA have been shown to disrupt expression of mitochondrial respiratory enzymes, leading to decreased insulin release, reduced calcium response to glucose stimulation, and reduced overall function [155-158]. This suggests that mitochondrial mutations could increase susceptibility to beta-cell dysfunction, the activation of mitochondrial-mediated cell death pathways, and to the development of diabetes. In fact, mutations in mitochondrial DNA are the direct cause of certain rare forms of diabetes [157]. Further, chronic conditions of high glucose, free fatty acids, proinflammatory cytokines, or other diabetes-associated triggers can impair [Ca²⁺]_i signaling and mitochondrial function in several tissues including islets [159-161] and may lead to mitochondrial calcium overload and cell death [162,163].

Exposure to proinflammatory cytokines is known to cause beta-cell dysfunction, and changes in mitochondrial calcium handling may play a role [64,164-167]. Grunnet et al. 2009 demonstrated that the proinflammatory cytokines IL-1β, IFN-γ, and TNF-α led to a calcium-activated, calcineurin-dependent dephosphorylation of the pro-apoptotic protein Bad, which caused beta-cells to undergo apoptosis [164]. Parkash et al 2005 demonstrated that TNF-α-treated RIN beta-cells expressed significantly less calbindin-D and a greater [Ca²⁺]_i response to increasing concentrations of ionomycin when compared to controls [168]. Further, TNF-α induced localization of the proapoptotic protein, Bax, to the perinuclear regions of RIN cells that contain the highest density of mitochondria, implying a calcium-dependent link between TNF-α and activation of the mitochondrial apoptotic pathway [168]. The uncoupling protein (UCP2) is another established effector of mitochondrial-mediated beta-cell death in several models of immune-mediated diabetes [169-171], and UCP2 is also known to regulate mitochondrial calcium sequestration [172]. Finally, the mitochondrial sodium-calcium exchanger, calcium uniporter or other anion channels can also affect islet function and could potentially play a role in cytokine action [173-176], although little is known at present. Cytokine action thus appears to be linked with mitochondrial dysfunction, at least in part, via calcium-dependent mechanisms.

7. Nuclear calcium and the effects of cytokines on the nucleus

Calcium’s most important role is in the nucleus due to its involvement in the transcription of genes regulating proliferation, apoptosis, and other key components of basic cellular life [1,2,177]. In the beta-cell, nutrients such as glucose are known to modulate the expression of many immediate early genes involved in these key processes in a [Ca²⁺]_i dependent manner [178]. In addition, nutrient-driven increases in calcium are thought to be the greatest in the nucleus. As demonstrated by Quesada et al., both cytoplasmic and nuclear calcium responses to depolarization by KCl are similar, however, the nuclear [Ca²⁺]_i response is significantly greater than that of the cytoplasm following glucose or oleate stimulation [179,180]. This effect is likely due to nuclear K_ATP channels and suggests that nutrients could produce nuclear-specific signals by generating transitory [Ca²⁺]_i levels within the nucleoplasm in excess of what is required for activation of cytosolic functions [180]. Raising nuclear calcium may indirectly regulate gene transcription as well as activate certain kinases, phosphatases and [Ca²⁺]_i binding transcription factors [2,181]. Also, the role of nuclear calcium may vary amongst particular cell types [2,181]. For instance, deep nuclear envelope invaginations have been observed in the nucleoplasm of GH3 pituitary cells and may even contain cytoplasmic organelles [150]. Thus, cell-specific morphological differences in the nucleoplasmic reticulum could potentially increase calcium release and aid in calcium homeostasis within the nucleoplasm [182]. In addition, recent studies indicate the possibility that [Ca²⁺]_i release from microstores into subnuclear microdomains could regulate the expression of restricted areas of the genome [181].

While cytokines are known to have adverse effects on calcium signaling [132], very little is known about cytokine effects on nuclear calcium signaling. Various proinflammatory cytokines affect many signal transduction pathways such as NF-kB, p38MAPK, JNK, and JAK-STAT, which are heavily involved in nuclear signaling and can induce apoptosis [183-186]. However, the extent to which changes in nuclear calcium can influence these pathways is not known. Numerous channels and receptors regulate nuclear calcium including ryanodine receptors, IP3Rs, and NAADPRs [187], which could potentially be prime targets of cytokine action. Considering the complex interconnectivity and the correlation of sizeable changes in nuclear calcium with nutrient availability, it stands to reason that cytokine-induced disruptions in nuclear calcium handling could lead to significant dysfunction in beta-cells and possibly induce cell death.

8. Final thoughts

As we have reviewed, glucose stimulates dynamic changes in cytosolic calcium that occur through extracellular influx and uptake/release from the ER, mitochondria, and nucleus (see Figure 3). The potential for calcium-mediated interplay among these organelles suggests that tremendously intricate communication may be necessary to carryout the dynamic events involved with the triphasic response to glucose stimulation and subsequent oscillations in [Ca²⁺]_i and insulin release [188,189]. The flipside of this intricate give-and-take is that disruptions in any of these calcium-regulating organelles can have consequences on all aspects of the cellular response. Proinflammatory cytokines utilize calcium in their signaling pathways and can also impact calcium handling in each of these organelles. The ER is widely appreciated for its dynamic calcium regulation through action of the SERCA pump, a direct cytokine target. Likewise, the mitochondrial permeability transition pore triggers apoptosis in response to mitochondrial calcium overload, which also may occur in response to cytokine action. Given the impact of cytokines on gene expression and nuclear signaling, elucidating the effect of cytokines on nuclear calcium is a particularly intriguing area for future research.

Figure 3.

Calcium interactions. Cytokines are known to affect calcium handling within the mitochondria and ER (✔) and may disrupt nuclear calcium (?). Cytokines may also influence calcium signaling between these organelles (arrows).

With regard to diabetes therapy, several anti-inflammatory compounds are being considered in clinical trials. For example, a recent pilot study showed that the TNF inhibitor, etanercept, improved A1C and islet function in new-onset T1D patients [190]. The IL-1 receptor antagonist, Anakinra, has shown similar success in improving glycemia and beta-cell secretory function in patients with T2D [191] and in mouse models of high-fat-diet-induced hyperglycemia [192]. Since calcium is important to both IL-1B and TNF-a signaling, and their respective cytokine receptor antagonists improve islet function, it is reasonable to suspect that calcium also plays a role in these improvements. To our knowledge, however, no studies have examined calcium changes directly. Anti-hypertensive drugs, including calcium-channel blockers, are also being examined for their anti-diabetic and anti-inflammatory effects. A recent meta-analysis of clinical studies showed that calcium-channel blockers were neutral in patients with hypertension with regard to new-onset diabetes [193], although it should be noted that beta-cell function was not a specific endpoint. This study also showed that the risk of developing diabetes was increased with diuretics and decreased with ARBs or ACE inhibitors [193]. ARBs are particularly intriguing because recent studies suggest they have anti-inflammatory properties and can improve beta-cell function [194,195].

Although it is not clear whether targeting calcium channels would be therapeutically beneficial to the beta-cell in diabetes patients, it is clear that the role of calcium, though not fully understood, is nevertheless central to many mechanisms of cytokine-induced beta-cell dysfunction and death. Measuring these dynamic changes in [Ca²⁺]_i can thus provide valuable insight into beta-cell function under normal and pathological circumstances and may even serve as a biomarker of islet function in vivo. A recently developed technique involving manganese-enhanced magnetic resonance imaging can be used to detect pancreatic responses to glucose stimulation by measuring changes in the activity of voltage-gated calcium channels [196] as a surrogate measurement of insulin secretion. This technique may one day be used clinically to identify functional beta-cell mass in diabetic patients.