Beneficial islet inflammation in health depends on pericytic TLR/MyD88 signaling.

While inflammation is beneficial for insulin secretion during homeostasis, its transformation adversely affects ß-cells and contributes to diabetes. However, the regulation of islet inflammation for maintaining glucose homeostasis remains largely unknown. Here, we identified pericytes as pivotal regulators of islet immune and ß-cell function in health. Islets and pancreatic pericytes express various cytokines in healthy humans and mice. To interfere with the pericytic inflammatory response, we selectively inhibited the TLR/MyD88 pathway in these cells in transgenic mice. The loss of MyD88 impaired pericytic cytokine production. Furthermore, MyD88-deficient mice exhibited skewed islet inflammation with fewer cells, an impaired macrophage phenotype, and reduced IL-1ß production. This aberrant pericyte-orchestrated islet inflammation was associated with ß-cell dedifferentiation and impaired glucose response. Additionally, we found that Cxcl1, a pericytic MyD88-dependent cytokine, promoted immune IL-1ß production. Treatments with either Cxcl1 or IL-1ß restored the mature ß-cell phenotype and glucose response in transgenic mice, suggesting a potential mechanism through which pericytes and immune cells regulate glucose homeostasis. Our study revealed pericyte-orchestrated islet inflammation as a crucial element in glucose regulation, implicating this process as a potential therapeutic target for diabetes.

Pericytes modulate islet immune cells and insulin secretion through Interleukin-33 production in mice.

Introduction: Immune cells were recently shown to support ß-cells and insulin secretion. However, little is known about how islet immune cells are regulated to maintain glucose homeostasis. Administration of various cytokines, including Interleukin-33 (IL-33), was shown to influence ß-cell function. However, the role of endogenous, locally produced IL-33 in pancreatic function remains unknown. Here, we show that IL-33, produced by pancreatic pericytes, is required for glucose homeostasis. Methods: To characterize pancreatic IL-33 production, we employed gene expression, flow cytometry, and immunofluorescence analyses. To define the role of this cytokine, we employed transgenic mouse systems to delete the Il33 gene selectively in pancreatic pericytes, in combination with the administration of recombinant IL-33. Glucose response was measured in vivo and in vitro, and morphometric and molecular analyses were used to measure ß-cell mass and gene expression. Immune cells were analyzed by flow cytometry. Resuts: Our results show that pericytes are the primary source of IL-33 in the pancreas. Mice lacking pericytic IL-33 were glucose intolerant due to impaired insulin secretion. Selective loss of pericytic IL-33 was further associated with reduced T and dendritic cell numbers in the islets and lower retinoic acid production by islet macrophages. Discussion: Our study demonstrates the importance of local, pericytic IL-33 production for glucose regulation. Additionally, it proposes that pericytes regulate islet immune cells to support ß-cell function in an IL-33-dependent manner. Our study reveals an intricate cellular network within the islet niche.

The postnatal pancreatic microenvironment guides ß cell maturation through BMP4 production.

Glucose homeostasis depends on regulated insulin secretion from pancreatic ß cells, which acquire their mature phenotype postnatally. The functional maturation of ß cells is regulated by a combination of cell-autonomous and exogenous factors; the identity of the latter is mostly unknown. Here, we identify BMP4 as a critical component through which the pancreatic microenvironment regulates ß cell function. By combining transgenic mouse models and human iPSCs, we show that BMP4 promotes the expression of core ß cell genes and is required for proper insulin production and secretion. We identified pericytes as the primary pancreatic source of BMP4, which start producing this ligand midway through the postnatal period, at the age ß cells mature. Overall, our findings show that the islet niche directly promotes ß cell functional maturation through the timely production of BMP4. Our study highlights the need to recapitulate the physiological postnatal islet niche for generating fully functional stem-cell-derived ß cells for cell replacement therapy for diabetes.

Pericytes contribute to the islet basement membranes to promote beta-cell gene expression.

ß-Cells depend on the islet basement membrane (BM). While some islet BM components are produced by endothelial cells (ECs), the source of others remains unknown. Pancreatic pericytes directly support ß-cells through mostly unidentified secreted factors. Thus, we hypothesized that pericytes regulate ß-cells through the production of BM components. Here, we show that pericytes produce multiple components of the mouse pancreatic and islet interstitial and BM matrices. Several of the pericyte-produced ECM components were previously implicated in ß-cell physiology, including collagen IV, laminins, proteoglycans, fibronectin, nidogen, and hyaluronan. Compared to ECs, pancreatic pericytes produce significantly higher levels of α2 and α4 laminin chains, which constitute the peri-islet and vascular BM. We further found that the pericytic laminin isoforms differentially regulate mouse ß-cells. Whereas α2 laminins promoted islet cell clustering, they did not affect gene expression. In contrast, culturing on Laminin-421 induced the expression of ß-cell genes, including Ins1, MafA, and Glut2, and significantly improved glucose-stimulated insulin secretion. Thus, alongside ECs, pericytes are a significant source of the islet BM, which is essential for proper ß-cell function.

GIP regulates inflammation and body weight by restraining myeloid-cell-derived S100A8/A9.

Enteroendocrine cells relay energy-derived signals to immune cells to signal states of nutrient abundance and control immunometabolism. Emerging data suggest that the gut-derived nutrient-induced incretin glucose-dependent insulinotropic polypeptide (GIP) operates at the interface of metabolism and inflammation. Here we show that high-fat diet (HFD)-fed mice with immune cell-targeted GIP receptor (GIPR) deficiency exhibit greater weight gain, insulin resistance, hepatic steatosis and significant myelopoiesis concomitantly with impaired energy expenditure and inguinal white adipose tissue (WAT) beiging. Expression of the S100 calcium-binding protein S100A8 was increased in the WAT of mice with immune cell-targeted GIPR deficiency and co-deletion of GIPR and the heterodimer S100A8/A9 in immune cells ameliorated the aggravated metabolic and inflammatory phenotype following a HFD. Specific GIPR deletion in myeloid cells identified this lineage as the target of GIP effects. Furthermore, GIP directly downregulated S100A8 expression in adipose tissue macrophages. Collectively, our results identify a myeloid-GIPR-S100A8/A9 signalling axis coupling nutrient signals to the control of inflammation and adaptive thermogenesis.

Pancreatic Pericytes Support ß-Cell Function in a Tcf7l2-Dependent Manner.

Polymorphism in TCF7L2, a component of the canonical Wnt signaling pathway, has a strong association with ß-cell dysfunction and type 2 diabetes through a mechanism that has yet to be defined. ß-Cells rely on cells in their microenvironment, including pericytes, for their proper function. Here, we show that Tcf7l2 activity in pancreatic pericytes is required for ß-cell function. Transgenic mice in which Tcf7l2 was selectively inactivated in their pancreatic pericytes exhibited impaired glucose tolerance due to compromised ß-cell function and glucose-stimulated insulin secretion. Inactivation of pericytic Tcf7l2 was associated with impaired expression of genes required for ß-cell function and maturity in isolated islets. In addition, we identified Tcf7l2-dependent pericytic expression of secreted factors shown to promote ß-cell function, including bone morphogenetic protein 4 (BMP4). Finally, we show that exogenous BMP4 is sufficient to rescue the impaired glucose-stimulated insulin secretion of transgenic mice, pointing to a potential mechanism through which pericytic Tcf7l2 activity affects ß-cells. To conclude, we suggest that pancreatic pericytes produce secreted factors, including BMP4, in a Tcf7l2-dependent manner to support ß-cell function. Our findings thus propose a potential cellular mechanism through which abnormal TCF7L2 activity predisposes individuals to diabetes and implicates abnormalities in the islet microenvironment in this disease.

Neonatal pancreatic pericytes support ß-cell proliferation.

OBJECTIVE: The maintenance and expansion of ß-cell mass rely on their proliferation, which reaches its peak in the neonatal stage. ß-cell proliferation was found to rely on cells of the islet microenvironment. We hypothesized that pericytes, which are components of the islet vasculature, support neonatal ß-cell proliferation. METHODS: To test our hypothesis, we combined in vivo and in vitro approaches. Briefly, we used a Diphtheria toxin-based transgenic mouse system to specifically deplete neonatal pancreatic pericytes in vivo. We further cultured neonatal pericytes isolated from the neonatal pancreas and combined the use of a ß-cell line and primary cultured mouse ß-cells. RESULTS: Our findings indicate that neonatal pancreatic pericytes are required and sufficient for ß-cell proliferation. We observed impaired proliferation of neonatal ß-cells upon in vivo depletion of pancreatic pericytes. Furthermore, exposure to pericyte-conditioned medium stimulated proliferation in cultured ß-cells. CONCLUSIONS: This study introduces pancreatic pericytes as regulators of neonatal ß-cell proliferation. In addition to advancing current understanding of the physiological ß-cell replication process, these findings could facilitate the development of protocols aimed at expending these cells as a potential cure for diabetes.

Isolating and Analyzing Cells of the Pancreas Mesenchyme by Flow Cytometry.

The pancreas is comprised of epithelial cells that are required for food digestion and blood glucose regulation. Cells of the pancreas microenvironment, including endothelial, neuronal, and mesenchymal cells were shown to regulate cell differentiation and proliferation in the embryonic pancreas. In the adult, the function and mass of insulin-producing cells were shown to depend on cells in their microenvironment, including pericyte, immune, endothelial, and neuronal cells. Lastly, changes in the pancreas microenvironment were shown to regulate pancreas tumorigenesis. However, the cues underlying these processes are not fully defined. Therefore, characterizing the different cell types that comprise the pancreas microenvironment and profiling their gene expression are crucial to delineate the tissue development and function under normal and diseased states. Here, we describe a method that allows for the isolation of mesenchymal cells from the pancreas of embryonic, neonatal, and adult mice. This method utilizes the enzymatic digestion of mouse pancreatic tissue and the subsequent fluorescence-activated cell sorting (FACS) or flow-cytometric analysis of labeled cells. Cells can be labeled by either immunostaining for surface markers or by the expression of fluorescent proteins. Cell isolation can facilitate the characterization of genes and proteins expressed in cells of the pancreas mesenchyme. This protocol was successful in isolating and culturing highly enriched mesenchymal cell populations from the embryonic, neonatal, and adult mouse pancreas.

Pancreatic Mesenchyme Regulates Islet Cellular Composition in a Patched/Hedgehog-Dependent Manner.

Pancreas development requires restrained Hedgehog (Hh) signaling activation. While deregulated Hh signaling in the pancreatic mesenchyme has been long suggested to be detrimental for proper organogenesis, this association was not directly shown. Here, we analyzed the contribution of mesenchymal Hh signaling to pancreas development. To increase Hh signaling in the pancreatic mesenchyme of mouse embryos, we deleted Patched1 (Ptch1) in these cells. Our findings indicate that deregulated Hh signaling in mesenchymal cells was sufficient to impair pancreas development, affecting both endocrine and exocrine cells. Notably, transgenic embryos displayed disrupted islet cellular composition and morphology, with a reduced ß-cell portion. Our results indicate that the cell-specific growth rates of α- and ß-cell populations, found during normal development, require regulated mesenchymal Hh signaling. In addition, we detected hyperplasia of mesenchymal cells upon elevated Hh signaling, accompanied by them acquiring smooth-muscle like phenotype. By specifically manipulating mesenchymal cells, our findings provide direct evidence for the non-autonomous roles of the Hh pathway in pancreatic epithelium development. To conclude, we directly show that regulated mesenchymal Hh signaling is required for pancreas organogenesis and establishment of its proper cellular composition.

Islet Pericytes Are Required for ß-Cell Maturity.

ß-Cells rely on the islet microenvironment for their functionality and mass. Pericytes, along with endothelial cells, make up the dense islet capillary network. However, although the role of endothelial cells in supporting ß-cell homeostasis has been vastly investigated, the role of pericytes remains largely unknown. Here, we focus on contribution of pericytes to ß-cell function. To this end, we used a transgenic mouse system that allows diphtheria toxin-based depletion of pericytes. Our results indicate that islets depleted of their pericytes have reduced insulin content and expression. Additionally, isolated islets displayed impaired glucose-stimulated insulin secretion, accompanied by a reduced expression of genes associated with ß-cell function. Importantly, reduced levels of the transcription factors MafA and Pdx1 point to ß-cell dedifferentiation in the absence of pericytes. Ex vivo depletion of pericytes in isolated islets resulted in a similar impairment of gene expression, implicating their direct, blood flow-independent role in maintaining ß-cell maturity. To conclude, our findings suggest that pericytes are pivotal components of the islet niche, which are required for ß-cell maturity and functionality. Abnormalities of islet pericytes, as implicated in type 2 diabetes, may therefore contribute to ß-cell dysfunction and disease progression.