Mouse pancreatic islet macrophages use locally released ATP to monitor beta cell activity.

AIMS/HYPOTHESIS: Tissue-resident macrophages sense the microenvironment and respond by producing signals that act locally to maintain a stable tissue state. It is now known that pancreatic islets contain their own unique resident macrophages, which have been shown to promote proliferation of the insulin-secreting beta cell. However, it is unclear how beta cells communicate with islet-resident macrophages. Here we hypothesised that islet macrophages sense changes in islet activity by detecting signals derived from beta cells. METHODS: To investigate how islet-resident macrophages respond to cues from the microenvironment, we generated mice expressing a genetically encoded Ca2+ indicator in myeloid cells. We produced living pancreatic slices from these mice and used them to monitor macrophage responses to stimulation of acinar, neural and endocrine cells. RESULTS: Islet-resident macrophages expressed functional purinergic receptors, making them exquisite sensors of interstitial ATP levels. Indeed, islet-resident macrophages responded selectively to ATP released locally from beta cells that were physiologically activated with high levels of glucose. Because ATP is co-released with insulin and is exclusively secreted by beta cells, the activation of purinergic receptors on resident macrophages facilitates their awareness of beta cell secretory activity. CONCLUSIONS/INTERPRETATION: Our results indicate that islet macrophages detect ATP as a proxy signal for the activation state of beta cells. Sensing beta cell activity may allow macrophages to adjust the secretion of factors to promote a stable islet composition and size.

Secretory Functions of Macrophages in the Human Pancreatic Islet Are Regulated by Endogenous Purinergic Signaling.

Endocrine cells of the pancreatic islet interact with their microenvironment to maintain tissue homeostasis. Communication with local macrophages is particularly important in this context, but the homeostatic functions of human islet macrophages are not known. In this study, we show that the human islet contains macrophages in perivascular regions that are the main local source of the anti-inflammatory cytokine interleukin-10 (IL-10) and the metalloproteinase MMP9. Macrophage production and secretion of these homeostatic factors are controlled by endogenous purinergic signals. In obese and diabetic states, macrophage expression of purinergic receptors MMP9 and IL-10 is reduced. We propose that in those states, exacerbated ß-cell activity due to increased insulin demand and increased cell death produce high levels of ATP that downregulate purinergic receptor expression. Loss of ATP sensing in macrophages may reduce their secretory capacity.

Gene Expression in Patient-Derived Neural Progenitors Implicates WNT5A Signaling in the Etiology of Schizophrenia.

BACKGROUND: Genome-wide association studies of schizophrenia have demonstrated that variations in noncoding regions are responsible for most of the common variation heritability of the disease. It is hypothesized that these risk variants alter gene expression. Therefore, studying alterations in gene expression in schizophrenia may provide a direct approach to understanding the etiology of the disease. In this study we use cultured neural progenitor cells derived from olfactory neuroepithelium (CNON cells) as a genetically unaltered cellular model to elucidate the neurodevelopmental aspects of schizophrenia. METHODS: We performed a gene expression study using RNA sequencing of CNON cells from 111 control subjects and 144 individuals with schizophrenia. Differentially expressed genes were identified with DESeq2 software, using covariates to correct for sex, age, library batches, and 1 surrogate variable component. RESULTS: A total of 80 genes were differentially expressed (false discovery rate < 10%), showing enrichment in cell migration, cell adhesion, developmental process, synapse assembly, cell proliferation, and related Gene Ontology categories. Cadherin and Wnt signaling pathways were positive in overrepresentation test, and, in addition, many genes were specifically involved in WNT5A signaling. The differentially expressed genes were modestly, but significantly, enriched in the genes overlapping single nucleotide polymorphisms with genome-wide significant association from the Psychiatric Genomics Consortium genome-wide association study of schizophrenia. We also found substantial overlap with genes associated with other psychiatric disorders or brain development, enrichment in the same Gene Ontology categories as genes with mutations de novo in schizophrenia, and studies of induced pluripotent stem cell-derived neural progenitor cells. CONCLUSIONS: CNON cells are a good model of the neurodevelopmental aspects of schizophrenia and can be used to elucidate the etiology of the disorder.

Paracrine Interactions within the Pancreatic Islet Determine the Glycemic Set Point.

Every animal species has a signature blood glucose level or glycemic set point. These set points are different, and the normal glycemic levels (normoglycemia) of one species would be life threatening for other species. Mouse normoglycemia can be considered diabetic for humans. The biological determinants of the glycemic set point remain unclear. Here we show that the pancreatic islet imposes its glycemic set point on the organism, making it the bona fide glucostat in the body. Moreover, and in contrast to rodent islets, glucagon input from the alpha cell to the insulin-secreting beta cell is necessary to fine-tune the distinctive human set point. These findings affect transplantation and regenerative approaches to treat diabetes because restoring normoglycemia may require more than replacing only the beta cells. Furthermore, therapeutic strategies using glucagon receptor antagonists as hypoglycemic agents need to be reassessed, as they may reset the overall glucostat in the organism.

Resealable, optically accessible, PDMS-free fluidic platform for ex vivo interrogation of pancreatic islets.

We report the design and fabrication of a robust fluidic platform built out of inert plastic materials and micromachined features that promote optimized convective fluid transport. The platform is tested for perfusion interrogation of rodent and human pancreatic islets, dynamic secretion of hormones, concomitant live-cell imaging, and optogenetic stimulation of genetically engineered islets. A coupled quantitative fluid dynamics computational model of glucose stimulated insulin secretion and fluid dynamics was first utilized to design device geometries that are optimal for complete perfusion of three-dimensional islets, effective collection of secreted insulin, and minimization of system volumes and associated delays. Fluidic devices were then fabricated through rapid prototyping techniques, such as micromilling and laser engraving, as two interlocking parts from materials that are non-absorbent and inert. Finally, the assembly was tested for performance using both rodent and human islets with multiple assays conducted in parallel, such as dynamic perfusion, staining and optogenetics on standard microscopes, as well as for integration with commercial perfusion machines. The optimized design of convective fluid flows, use of bio-inert and non-absorbent materials, reversible assembly, manual access for loading and unloading of islets, and straightforward integration with commercial imaging and fluid handling systems proved to be critical for perfusion assay, and particularly suited for time-resolved optogenetics studies.

Targeting AR Variant-Coactivator Interactions to Exploit Prostate Cancer Vulnerabilities.

Castration-resistant prostate cancer (CRPC) progresses rapidly and is incurable. Constitutively active androgen receptor splice variants (AR-Vs) represent a well-established mechanism of therapeutic resistance and disease progression. These variants lack the AR ligand-binding domain and, as such, are not inhibited by androgen deprivation therapy (ADT), which is the standard systemic approach for advanced prostate cancer. Signaling by AR-Vs, including the clinically relevant AR-V7, is augmented by Vav3, an established AR coactivator in CRPC. Using mutational and biochemical studies, we demonstrated that the Vav3 Diffuse B-cell lymphoma homology (DH) domain interacted with the N-terminal region of AR-V7 (and full length AR). Expression of the Vav3 DH domain disrupted Vav3 interaction with and enhancement of AR-V7 activity. The Vav3 DH domain also disrupted AR-V7 interaction with other AR coactivators: Src1 and Vav2, which are overexpressed in PC. This Vav3 domain was used in proof-of-concept studies to evaluate the effects of disrupting the interaction between AR-V7 and its coactivators on CRPC cells. This disruption decreased CRPC cell proliferation and anchorage-independent growth, caused increased apoptosis, decreased migration, and resulted in the acquisition of morphological changes associated with a less aggressive phenotype. While disrupting the interaction between FL-AR and its coactivators decreased N-C terminal interaction, disrupting the interaction of AR-V7 with its coactivators decreased AR-V7 nuclear levels.Implications: This study demonstrates the potential therapeutic utility of inhibiting constitutively active AR-V signaling by disrupting coactivator binding. Such an approach is significant, as AR-Vs are emerging as important drivers of CRPC that are particularly recalcitrant to current therapies. Mol Cancer Res; 15(11); 1469-80. ©2017 AACR.