Decreases in Gap Junction Coupling Recovers Ca2+ and Insulin Secretion in Neonatal Diabetes Mellitus, Dependent on Beta Cell Heterogeneity and Noise.

Diabetes is caused by dysfunction to ß-cells in the islets of Langerhans, disrupting insulin secretion and glucose homeostasis. Gap junction-mediated electrical coupling between ß-cells in the islet plays a major role in coordinating a pulsatile secretory response at elevated glucose and suppressing insulin secretion at basal glucose. Previously, we demonstrated that a critical number of inexcitable cells can rapidly suppress the overall islet response, as a result of gap junction coupling. This was demonstrated in a murine model of Neonatal Diabetes Mellitus (NDM) involving expression of ATP-insensitive KATP channels, and by a multi-cellular computational model of islet electrical activity. Here we examined the mechanisms by which gap junction coupling contributes to islet dysfunction in NDM. We first verified the computational model against [Ca2+] and insulin secretion measurements in islets expressing ATP-insensitive KATP channels under different levels of gap junction coupling. We then applied this model to predict how different KATP channel mutations found in NDM suppress [Ca2+], and the role of gap junction coupling in this suppression. We further extended the model to account for stochastic noise and insulin secretion dynamics. We found experimentally and in the islet model that reductions in gap junction coupling allow progressively greater glucose-stimulated [Ca2+] and insulin secretion following expression of ATP-insensitive KATP channels. The model demonstrated good correspondence between suppression of [Ca2+] and clinical presentation of different NDM mutations. Significant recoveries in [Ca2+] and insulin secretion were predicted for many mutations upon reductions in gap junction coupling, where stochastic noise played a significant role in the recoveries. These findings provide new understanding how the islet functions as a multicellular system and for the role of gap junction channels in exacerbating the effects of decreased cellular excitability. They further suggest novel therapeutic options for NDM and other monogenic forms of diabetes.

Phase transitions in the multi-cellular regulatory behavior of pancreatic islet excitability.

The pancreatic islets of Langerhans are multicellular micro-organs integral to maintaining glucose homeostasis through secretion of the hormone insulin. ß-cells within the islet exist as a highly coupled electrical network which coordinates electrical activity and insulin release at high glucose, but leads to global suppression at basal glucose. Despite its importance, how network dynamics generate this emergent binary on/off behavior remains to be elucidated. Previous work has suggested that a small threshold of quiescent cells is able to suppress the entire network. By modeling the islet as a Boolean network, we predicted a phase-transition between globally active and inactive states would emerge near this threshold number of cells, indicative of critical behavior. This was tested using islets with an inducible-expression mutation which renders defined numbers of cells electrically inactive, together with pharmacological modulation of electrical activity. This was combined with real-time imaging of intracellular free-calcium activity [Ca2+]i and measurement of physiological parameters in mice. As the number of inexcitable cells was increased beyond ∼15%, a phase-transition in islet activity occurred, switching from globally active wild-type behavior to global quiescence. This phase-transition was also seen in insulin secretion and blood glucose, indicating physiological impact. This behavior was reproduced in a multicellular dynamical model suggesting critical behavior in the islet may obey general properties of coupled heterogeneous networks. This study represents the first detailed explanation for how the islet facilitates inhibitory activity in spite of a heterogeneous cell population, as well as the role this plays in diabetes and its reversal. We further explain how islets utilize this critical behavior to leverage cellular heterogeneity and coordinate a robust insulin response with high dynamic range. These findings also give new insight into emergent multicellular dynamics in general which are applicable to many coupled physiological systems, specifically where inhibitory dynamics result from coupled networks.

Dimensionality and size scaling of coordinated Ca(2+) dynamics in MIN6 ß-cell clusters.

Pancreatic islets of Langerhans regulate blood glucose homeostasis by the secretion of the hormone insulin. Like many neuroendocrine cells, the coupling between insulin-secreting ß-cells in the islet is critical for the dynamics of hormone secretion. We have examined how this coupling architecture regulates the electrical dynamics that underlie insulin secretion by utilizing a microwell-based aggregation method to generate clusters of a ß-cell line with defined sizes and dimensions. We measured the dynamics of free-calcium activity ([Ca(2+)]i) and insulin secretion and compared these measurements with a percolating network model. We observed that the coupling dimension was critical for regulating [Ca(2+)]i dynamics and insulin secretion. Three-dimensional coupling led to size-invariant suppression of [Ca(2+)]i at low glucose and robust synchronized [Ca(2+)]i oscillations at elevated glucose, whereas two-dimensional coupling showed poor suppression and less robust synchronization, with significant size-dependence. The dimension- and size-scaling of [Ca(2+)]i at high and low glucose could be accurately described with the percolating network model, using similar network connectivity. As such this could explain the fundamentally different behavior and size-scaling observed under each coupling dimension. This study highlights the dependence of proper ß-cell function on the coupling architecture that will be important for developing therapeutic treatments for diabetes such as islet transplantation techniques. Furthermore, this will be vital to gain a better understanding of the general features by which cellular interactions regulate coupled multicellular systems.

Trisomy 21 causes changes in the circulating proteome indicative of chronic autoinflammation.

Trisomy 21 (T21) causes Down syndrome (DS), but the mechanisms by which T21 produces the different disease spectrum observed in people with DS are unknown. We recently identified an activated interferon response associated with T21 in human cells of different origins, consistent with overexpression of the four interferon receptors encoded on chromosome 21, and proposed that DS could be understood partially as an interferonopathy. However, the impact of T21 on systemic signaling cascades in living individuals with DS is undefined. To address this knowledge gap, we employed proteomics approaches to analyze blood samples from 263 individuals, 165 of them with DS, leading to the identification of dozens of proteins that are consistently deregulated by T21. Most prominent among these proteins are numerous factors involved in immune control, the complement cascade, and growth factor signaling. Importantly, people with DS display higher levels of many pro-inflammatory cytokines (e.g. IL-6, MCP-1, IL-22, TNF-α) and pronounced complement consumption, resembling changes seen in type I interferonopathies and other autoinflammatory conditions. Therefore, these results are consistent with the hypothesis that increased interferon signaling caused by T21 leads to chronic immune dysregulation, and justify investigations to define the therapeutic value of immune-modulatory strategies in DS.

Decreasing cx36 gap junction coupling compensates for overactive KATP channels to restore insulin secretion and prevent hyperglycemia in a mouse model of neonatal diabetes.

Mutations to the ATP-sensitive K(+) channel (KATP channel) that reduce the sensitivity of ATP inhibition cause neonatal diabetes mellitus via suppression of ß-cell glucose-stimulated free calcium activity ([Ca(2+)]i) and insulin secretion. Connexin-36 (Cx36) gap junctions also regulate islet electrical activity; upon knockout of Cx36, ß-cells show [Ca(2+)]i elevations at basal glucose. We hypothesized that in the presence of overactive ATP-insensitive KATP channels, a reduction in Cx36 would allow elevations in glucose-stimulated [Ca(2+)]i and insulin secretion to improve glucose homeostasis. To test this, we introduced a genetic knockout of Cx36 into mice that express ATP-insensitive KATP channels and measured glucose homeostasis and islet metabolic, electrical, and insulin secretion responses. In the normal presence of Cx36, after expression of ATP-insensitive KATP channels, blood glucose levels rapidly rose to >500 mg/dL. Islets from these mice showed reduced glucose-stimulated [Ca(2+)]i and no insulin secretion. In mice lacking Cx36 after expression of ATP-insensitive KATP channels, normal glucose levels were maintained. Islets from these mice had near-normal glucose-stimulated [Ca(2+)]i and insulin secretion. We therefore demonstrate a novel mechanism by which islet function can be recovered in a monogenic model of diabetes. A reduction of gap junction coupling allows sufficient glucose-stimulated [Ca(2+)]i and insulin secretion to prevent the emergence of diabetes.