Justin P. Annes

Adenosine kinase inhibition selectively promotes rodent and porcine islet β-cell replication

Diabetes is a pathological condition characterized by relative insulin deficiency, persistent hyperglycemia, and, consequently, diffuse micro- and macrovascular disease. One therapeutic strategy is to amplify insulin-secretion capacity by increasing the number of the insulin-producing β cells without triggering a generalized proliferative response. Here, we present the development of a small-molecule screening platform for the identification of molecules that increase β-cell replication. Using this platform, we identify a class of compounds [adenosine kinase inhibitors (ADK-Is)] that promote replication of primary β cells in three species (mouse, rat, and pig). Furthermore, the replication effect of ADK-Is is cell type-selective: treatment of islet cell cultures with ADK-Is increases replication of β cells but not that of α cells, PP cells, or fibroblasts. Short-term in vivo treatment with an ADK-I also increases β-cell replication but not exocrine cell or hepatocyte replication. Therefore, we propose ADK inhibition as a strategy for the treatment of diabetes.

A liver Hif-2α-Irs2 pathway sensitizes hepatic insulin signaling and is modulated by Vegf inhibition

Insulin initiates diverse hepatic metabolic responses, including gluconeogenic suppression and induction of glycogen synthesis and lipogenesis. The liver possesses a rich sinusoidal capillary network with a higher degree of hypoxia and lower gluconeogenesis in the perivenous zone as compared to the rest of the organ. Here, we show that diverse vascular endothelial growth factor (VEGF) inhibitors improved glucose tolerance in nondiabetic C57BL/6 and diabetic db/db mice, potentiating hepatic insulin signaling with lower gluconeogenic gene expression, higher glycogen storage and suppressed hepatic glucose production. VEGF inhibition induced hepatic hypoxia through sinusoidal vascular regression and sensitized liver insulin signaling through hypoxia-inducible factor-2α (Hif-2α, encoded by Epas1) stabilization. Notably, liver-specific constitutive activation of HIF-2α, but not HIF-1α, was sufficient to augment hepatic insulin signaling through direct and indirect induction of insulin receptor substrate-2 (Irs2), an essential insulin receptor adaptor protein. Further, liver Irs2 was both necessary and sufficient to mediate Hif-2α and Vegf inhibition effects on glucose tolerance and hepatic insulin signaling. These results demonstrate an unsuspected intersection between Hif-2α−mediated hypoxic signaling and hepatic insulin action through Irs2 induction, which can be co-opted by Vegf inhibitors to modulate glucose metabolism. These studies also indicate distinct roles in hepatic metabolism for Hif-1α, which promotes glycolysis, and Hif-2α, which suppresses gluconeogenesis, and suggest new treatment approaches for type 2 diabetes mellitus.

Erdheim-Chester disease presenting with cutaneous involvement: a case report and literature review.

Erdheim–Chester disease (ECD) is a rare, systemic, non‐familial histiocytic disorder, first described by Jakob Erdheim and William Chester in 1930. Most patients have multiple sites of involvement at presentation. The most common site of involvement is the long bones of the axial skeleton, which is seen almost universally, followed by the nervous system, heart, lungs, orbit and retroperitoneum, which are seen in up to 50% of cases. 1 Cutaneous involvement is rarely a presenting symptom of ECD, with two reported cases in the English literature. 2 The diagnosis of ECD is rarely made by skin biopsy because of the relative rarity of cutaneous involvement as a presenting feature, and also perhaps because of the difficulty in distinguishing the histopathological appearance from potential mimics. The importance of distinguishing ECD from other cutaneous disorders with similar pathology lies in the implications for both treatment and prognosis. ECD is an aggressive, often fatal disorder, with death from disease occurring in greater than 50% of patients.

Repurposing cAMP-Modulating Medications to Promote β-Cell Replication

Loss of β-cell mass is a cardinal feature of diabetes. Consequently, developing medications to promote β-cell regeneration is a priority. cAMP is an intracellular second messenger that modulates β-cell replication. We investigated whether medications that increase cAMP stability or synthesis selectively stimulate β-cell growth. To identify cAMP-stabilizing medications that promote β-cell replication, we performed high-content screening of a phosphodiesterase (PDE) inhibitor library. PDE3, -4, and -10 inhibitors, including dipyridamole, were found to promote β-cell replication in an adenosine receptor-dependent manner. Dipyridamoles action is specific for β-cells and not α-cells. Next we demonstrated that norepinephrine (NE), a physiologic suppressor of cAMP synthesis in β-cells, impairs β-cell replication via activation of α(2)-adrenergic receptors. Accordingly, mirtazapine, an α(2)-adrenergic receptor antagonist and antidepressant, prevents NE-dependent suppression of β-cell replication. Interestingly, NEs growth-suppressive effect is modulated by endogenously expressed catecholamine-inactivating enzymes (catechol-O-methyltransferase and l-monoamine oxidase) and is dominant over the growth-promoting effects of PDE inhibitors. Treatment with dipyridamole and/or mirtazapine promote β-cell replication in mice, and treatment with dipyridamole is associated with reduced glucose levels in humans. This work provides new mechanistic insights into cAMP-dependent growth regulation of β-cells and highlights the potential of commonly prescribed medications to influence β-cell growth.

The influence of sodium- and calcium-regulatory hormone interventions on adipocytokines in obesity and diabetes

OBJECTIVE The renin-angiotensin-aldosterone system (RAAS), vitamin D, and parathyroid hormone have all been implicated as regulators of adipocytokines and inflammation. We evaluated human interventional study protocols to investigate whether controlled modulations of these calcium- and sodium-regulatory hormones could influence adipocytokines and inflammation in obesity and diabetes. METHODS Post-hoc analyses of two separate human protocols (Protocol 1, n=14; Protocol 2, n=24) conducted in a clinical research setting after rigorous control of diet, posture, medications, and diurnal rhythm, were performed. Protocol 1 evaluated obese hypertensives with vitamin D deficiency who received an infusion of angiotensin II (AngII) before and after 1month of vitamin D3 therapy. Protocol 2 evaluated obese subjects with type 2 diabetes who also received AngII. Adipocytokines and inflammatory markers were measured before and after vitamin D3 therapy, and also before and after infusions of AngII. RESULTS Vitamin D3 therapy significantly raised 25(OH)D and 1,25(OH)2D concentrations, and lowered parathyroid hormone, but had no effect on concentrations of adiponectin, resistin, leptin, IL-6, PAI-1, urinary TGFβ1, or HOMA-IR. AngII infusions, despite significant elevations in blood pressure and serum aldosterone, did not influence adipocytokine concentrations in either protocol. CONCLUSION In contrast to prior studies conducted in healthy populations, or those that could not control major regulators of the RAAS or adipocytokines, we observed that robust modulations in calcium- and sodium-regulatory hormones did not influence adipocytokines or inflammation in obesity or diabetes. Adipose-tissue physiology in these conditions may alter the hormonal regulation of inflammatory parameters.

Adult tissue sources for new β cells

The diabetes pandemic incurs extraordinary public health and financial costs that are projected to expand for the foreseeable future. Consequently, the development of definitive therapies for diabetes is a priority. Currently, a wide spectrum of therapeutic strategies-from implantable insulin delivery devices to transplantation-based cell replacement therapy, to β-cell regeneration-focus on replacing the lost insulin-producing capacity of individuals with diabetes. Among these, β-cell regeneration remains promising but heretofore unproved. Indeed, recent experimental work has uncovered surprising biology that underscores the potential therapeutic benefit of β-cell regeneration. These studies have elucidated a variety of sources for the endogenous production of new β cells from existing cells. First, β cells, long thought to be postmitotic, have demonstrated the potential for regenerative capacity. Second, the presence of pancreatic facultative endocrine progenitor cells has been established. Third, the malleability of cellular identity has availed the possibility of generating β cells from other differentiated cell types. Here, we review the exciting developments surrounding endogenous sources of β-cell production and consider the potential of realizing a regenerative therapy for diabetes from adult tissues.

Genetics of adrenocortical disease: an update.

Purpose of reviewDisease states characterized by abnormal cellular function or proliferation frequently reflect aberrant genetic information. By revealing disease-specific DNA mutations, we gain insight into normal physiology, pathophysiology, potential therapeutic targets and are better equipped to evaluate an individuals disease risks. This review examines recent advances in our understanding of the genetic basis of adrenal cortical disease. Recent findingsImportant advances made in the past year have included identification of KCNJ5 potassium channel mutations in the pathogenesis of both aldosterone-producing adenomas and familial hyperaldosteronism type III; characterization of phosphodiesterase 11A as a modifier of phenotype in Carney complex caused by protein kinase, cAMP-dependent, regulatory subunit, type-I mutations; the finding of 11&bgr;-hydroxysteroid dehydrogenase type I mutations as a novel mechanism for cortisone reductase deficiency; and demonstration of potential mortality benefit in pursuing comprehensive presymptomatic screening for patients with Li–Fraumeni syndrome, including possible reduction in risks associated with adrenocortical carcinoma. SummaryThis research review provides a framework for the endocrinologist to maintain an up-to-date understanding of adrenal cortical disease genetics.

Hyaluronan content governs tissue stiffness in pancreatic islet inflammation

We have identified a novel role for hyaluronan (HA), an extracellular matrix polymer, in governing the mechanical properties of inflamed tissues. We recently reported that insulitis in type 1 diabetes of mice and humans is preceded by intraislet accumulation of HA, a highly hygroscopic polymer. Using the double transgenic DO11.10 × RIPmOVA (DORmO) mouse model of type 1 diabetes, we asked whether autoimmune insulitis was associated with changes in the stiffness of islets. To measure islet stiffness, we used atomic force microscopy (AFM) and developed a novel “bed of nails”-like approach that uses quartz glass nanopillars to anchor islets, solving a long-standing problem of keeping tissue-scale objects immobilized while performing AFM. We measured stiffness via AFM nanoindentation with a spherical indenter and found that insulitis made islets mechanically soft compared with controls. Conversely, treatment with 4-methylumbelliferone, a small-molecule inhibitor of HA synthesis, reduced HA accumulation, diminished swelling, and restored basal tissue stiffness. These results indicate that HA content governs the mechanical properties of islets. In hydrogels with variable HA content, we confirmed that increased HA leads to mechanically softer hydrogels, consistent with our model. In light of recent reports that the insulin production of islets is mechanosensitive, these findings open up an exciting new avenue of research into the fundamental mechanisms by which inflammation impacts local cellular responses.

CC-401 Promotes β-Cell Replication via Pleiotropic Consequences of DYRK1A/B Inhibition

Pharmacologic expansion of endogenous β cells is a promising therapeutic strategy for diabetes. To elucidate the molecular pathways that control β-cell growth we screened ∼2400 bioactive compounds for rat β-cell replication-modulating activity. Numerous hit compounds impaired or promoted rat β-cell replication, including CC-401, an advanced clinical candidate previously characterized as a c-Jun N-terminal kinase inhibitor. Surprisingly, CC-401 induced rodent (in vitro and in vivo) and human (in vitro) β-cell replication via dual-specificity tyrosine phosphorylation-regulated kinase (DYRK) 1A and 1B inhibition. In contrast to rat β cells, which were broadly growth responsive to compound treatment, human β-cell replication was only consistently induced by DYRK1A/B inhibitors. This effect was enhanced by simultaneous glycogen synthase kinase-3β (GSK-3β) or activin A receptor type II-like kinase/transforming growth factor-β (ALK5/TGF-β) inhibition. Prior work emphasized DYRK1A/B inhibition-dependent activation of nuclear factor of activated T cells (NFAT) as the primary mechanism of human β-cell-replication induction. However, inhibition of NFAT activity had limited effect on CC-401-induced β-cell replication. Consequently, we investigated additional effects of CC-401-dependent DYRK1A/B inhibition. Indeed, CC-401 inhibited DYRK1A-dependent phosphorylation/stabilization of the β-cell-replication inhibitor p27Kip1. Additionally, CC-401 increased expression of numerous replication-promoting genes normally suppressed by the dimerization partner, RB-like, E2F and multivulval class B (DREAM) complex, which depends upon DYRK1A/B activity for integrity, including MYBL2 and FOXM1. In summary, we present a compendium of compounds as a valuable resource for manipulating the signaling pathways that control β-cell replication and leverage a DYRK1A/B inhibitor (CC-401) to expand our understanding of the molecular pathways that control β-cell growth.

Genetic Disruption of Adenosine Kinase in Mouse Pancreatic β-Cells Protects Against High-Fat Diet–Induced Glucose Intolerance

Islet β-cells adapt to insulin resistance through increased insulin secretion and expansion. Type 2 diabetes typically occurs when prolonged insulin resistance exceeds the adaptive capacity of β-cells. Our prior screening efforts led to the discovery that adenosine kinase (ADK) inhibitors stimulate β-cell replication. Here, we evaluated whether ADK disruption in mouse β-cells affects β-cell mass and/or protects against high-fat diet (HFD)–induced glucose dysregulation. Mice targeted at the Adk locus were bred to Rip-Cre and Ins1-Cre/ERT1Lphi mice to enable constitutive (βADKO) and conditional (iβADKO) disruption of ADK expression in β-cells, respectively. Weight gain, glucose tolerance, insulin sensitivity, and glucose-stimulated insulin secretion (GSIS) were longitudinally monitored in normal chow (NC)–fed and HFD-fed mice. In addition, β-cell mass and replication were measured by immunofluorescence-based islet morphometry. NC-fed adult βADKO and iβADKO mice displayed glucose tolerance, insulin tolerance and β-cell mass comparable to control animals. By contrast, HFD-fed βADKO and iβADKO animals had improved glucose tolerance and increased in vivo GSIS. Improved glucose handling was associated with increased β-cell replication and mass. We conclude that ADK expression negatively regulates the adaptive β-cell response to HFD challenge. Therefore, modulation of ADK activity is a potential strategy for enhancing the adaptive β-cell response.