scholarly journals Improved in vivo imaging method for individual islets across the mouse pancreas reveals a heterogeneous insulin secretion response to glucose

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Henriette Frikke-Schmidt ◽  
Peter Arvan ◽  
Randy J. Seeley ◽  
Corentin Cras-Méneur
Keyword(s):  
Insulin Secretion ◽  
In Vivo Imaging ◽  
Motion Blur ◽  
Isolated Islets ◽  
Imaging Method ◽  
Glucose Stimulation ◽  
Powerful Approach ◽  
Glucose Challenge ◽  
The Impact

AbstractWhile numerous techniques can be used to measure and analyze insulin secretion in isolated islets in culture, assessments of insulin secretion in vivo are typically indirect and only semiquantitative. The CpepSfGFP reporter mouse line allows the in vivo imaging of insulin secretion from individual islets after a glucose stimulation, in live, anesthetized mice. Imaging the whole pancreas at high resolution in live mice to track the response of each individual islet over time includes numerous technical challenges and previous reports were only limited in scope and non-quantitative. Elaborating on this previous model—through the development of an improved methodology addressing anesthesia, temperature control and motion blur—we were able to track and quantify longitudinally insulin content throughout a glucose challenge in up to two hundred individual islets simultaneously. Through this approach we demonstrate quantitatively for the first time that while isolated islets respond homogeneously to glucose in culture, their profiles differ significantly in vivo. Independent of size or location, some islets respond sharply to a glucose stimulation while others barely secrete at all. This platform therefore provides a powerful approach to study the impact of disease, diet, surgery or pharmacological treatments on insulin secretion in the intact pancreas in vivo.

scholarly journals In vivo imaging of individual islets across the mouse pancreas reveals a heterogeneous insulin secretion response to glucose

2020 ◽  
Author(s):  
Henriette Frikke-Schmidt ◽  
Peter Arvan ◽  
Randy J Seeley ◽  
Corentin Cras-Méneur
Keyword(s):  
Insulin Secretion ◽  
Dynamic Range ◽  
High Dynamic Range ◽  
Motion Blur ◽  
Isolated Islets ◽  
Computer Assisted ◽  
Powerful Approach ◽  
Glucose Challenge ◽  
The Impact

AbstractWhile numerous techniques can be used to measure and analyze insulin secretion in isolated islets in culture, assessments of insulin secretion in vivo are typically indirect and only semiquantitative. The CpepSfGFP reporter mouse line allows the in vivo imaging of insulin secretion from individual islets after a glucose challenge, in live, anesthetized mice, addressing secretion from the pancreas as a whole. Imaging the whole pancreas at high resolution in live mice includes numerous technical challenges. Using rapid-fire imaging in high dynamic range and tiling combined with computer-assisted masked morphometry, we have developed a method to overcome motion blur, and the effects of anesthesia, to be able to monitor and quantify insulin (CpepSfGFP) content simultaneously and longitudinally for the first time in hundreds of individual islets, throughout a glucose challenge.Through this approach we demonstrate that while isolated islets respond homogeneously to glucose in culture, their response profile differs significantly in vivo. Independent of size or location, some islets respond sharply to a glucose stimulation while others barely secrete at all. This platform therefore provides a powerful approach to study the impact of disease, diet, surgery or pharmacological treatments on insulin secretion in the intact pancreas in vivo.

scholarly journals Isoform-specific roles of prolyl hydroxylases in the regulation of pancreatic β-cell function

Endocrinology ◽  
2021 ◽  
Author(s):  
Monica Hoang ◽  
Emelien Jentz ◽  
Sarah M Janssen ◽  
Daniela Nasteska ◽  
Federica Cuozzo ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Katp Channel ◽  
Cell Mass ◽  
Isolated Islets ◽  
Second Phase ◽  
Β Cells ◽  
Β Cell ◽  
Prolyl Hydroxylases ◽  
Glucose Challenge

Abstract Pancreatic β-cells can secrete insulin via two pathways characterized as KATP channel-dependent and independent. The KATP channel-independent pathway is characterized by a rise in several potential metabolic signaling molecules, including the NADPH/NADP + ratio and α-ketoglutarate (αKG). Prolyl hydroxylases (PHDs), which belong to the αKG-dependent dioxygenase superfamily, are known to regulate the stability of hypoxia-inducible factor α (HIFα). In the current study, we assess the role of PHDs in vivo using the pharmacological inhibitor dimethyloxalylglycine (DMOG) and generated β-cell specific knockout (KO) mice for all three isoforms of PHD (β-PHD1 KO, β-PHD2 KO, and β-PHD3 KO mice). DMOG inhibited in vivo insulin secretion in response to glucose challenge and inhibited the 1 st phase of insulin secretion but enhanced the second-phase of insulin secretion in isolated islets. None of the β-PHD KO mice showed any significant in vivo defects associated with glucose tolerance and insulin resistance except for β-PHD2 KO mice which had significantly increased plasma insulin during a glucose challenge. Islets from both β-PHD1 KO and β-PHD3 KO had elevated β-cell apoptosis and reduced β-cell mass. Isolated islets from β-PHD1 KO and β-PHD3 KO had impaired glucose-stimulated insulin secretion and glucose-stimulated increases in the ATP/ADP and NADPH/NADP + ratio. All three PHD isoforms are expressed in β-cells, with PHD3 showing the most unique expression pattern. The lack of each PHD protein did not significantly impair in vivo glucose homeostasis. However, β-PHD1 KO and β-PHD3 KO mice had defective β-cell mass and islet insulin secretion, suggesting that these mice may be predisposed to developing diabetes.

Loss of growth hormone signaling in the mouse germline or in adulthood reduces islet mass and alters islet function with notable sex differences

2021 ◽  
Author(s):  
Silvana Duran-Ortiz ◽  
Kathryn L. Corbin ◽  
Ishrat Jahan ◽  
Nicholas B. Whitticar ◽  
Sarah E Morris ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Insulin Secretion ◽  
Insulin Sensitivity ◽  
Glucose Tolerance ◽  
Body Fat ◽  
Isolated Islets ◽  
Wild Type ◽  
Cross Sectional ◽  
Islet Mass ◽  
The Impact

In the endocrine pancreas, growth hormone (GH) is known to promote pancreatic islet growth and insulin secretion. In this study, we show that GH receptor (GHR) loss in the germline and in adulthood impacts islet mass in general but more profoundly in male mice. GHR knockout (GHRKO) mice have enhanced insulin sensitivity and low circulating insulin. We show that the total cross-sectional area of isolated islets (estimated islet mass) was reduced by 72% in male but by only 29% in female GHRKO mice compared to wild type controls. Also, islets from GHRKO mice secreted ~50% less glucose-stimulated insulin compared to size-matched islets from wild type mice. We next used mice with a floxed Ghr gene to knock down the GHR in adult mice at six-months of age (6mGHRKO) and examined the impact on glucose and islet metabolism. By 12-months of age, female 6mGHRKO mice had increased body fat and reduced islet mass but had no change in glucose tolerance or insulin sensitivity. However, male 6mGHRKO mice had nearly twice as much body fat, substantially reduced islet mass, and enhanced insulin sensitivity, but no change in glucose tolerance. Despite large losses in islet mass, glucose-stimulated insulin secretion from isolated islets was not significantly different between male 6mGHRKO and controls while isolated islets from female 6mGHRKO mice showed increased glucose-stimulated insulin release. Our findings demonstrate the importance of GH to islet mass throughout life and that unique sex-specific adaptations to the loss of GH signaling allow mice to maintain normal glucose metabolism.

scholarly journals Metabolic Effects of Selective Deletion of Group VIA Phospholipase A2 from Macrophages or Pancreatic Islet Beta-Cells

Biomolecules ◽  
2020 ◽  
Vol 10 (10) ◽  
pp. 1455
Author(s):  
John Turk ◽  
Haowei Song ◽  
Mary Wohltmann ◽  
Cheryl Frankfater ◽  
Xiaoyong Lei ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Glucose Tolerance ◽  
Phospholipase A2 ◽  
Ex Vivo ◽  
Isolated Islets ◽  
Metabolic Effects ◽  
Β Cell ◽  
Cell Functions ◽  
Normal Glucose

To examine the role of group VIA phospholipase A2 (iPLA2β) in specific cell lineages in insulin secretion and insulin action, we prepared mice with a selective iPLA2β deficiency in cells of myelomonocytic lineage, including macrophages (MØ-iPLA2β-KO), or in insulin-secreting β-cells (β-Cell-iPLA2β-KO), respectively. MØ-iPLA2β-KO mice exhibited normal glucose tolerance when fed standard chow and better glucose tolerance than floxed-iPLA2β control mice after consuming a high-fat diet (HFD). MØ-iPLA2β-KO mice exhibited normal glucose-stimulated insulin secretion (GSIS) in vivo and from isolated islets ex vivo compared to controls. Male MØ-iPLA2β-KO mice exhibited enhanced insulin responsivity vs. controls after a prolonged HFD. In contrast, β-cell-iPLA2β-KO mice exhibited impaired glucose tolerance when fed standard chow, and glucose tolerance deteriorated further when introduced to a HFD. β-Cell-iPLA2β-KO mice exhibited impaired GSIS in vivo and from isolated islets ex vivo vs. controls. β-Cell-iPLA2β-KO mice also exhibited an enhanced insulin responsivity compared to controls. These findings suggest that MØ iPLA2β participates in HFD-induced deterioration in glucose tolerance and that this mainly reflects an effect on insulin responsivity rather than on insulin secretion. In contrast, β-cell iPLA2β plays a role in GSIS and also appears to confer some protection against deterioration in β-cell functions induced by a HFD.

scholarly journals Islet macrophages are associated with islet vascular remodeling and compensatory hyperinsulinemia during diabetes

2019 ◽  
Vol 317 (6) ◽  
pp. E1108-E1120 ◽  
Author(s):  
Manesh Chittezhath ◽  
Divya Gunaseelan ◽  
Xiaofeng Zheng ◽  
Riasat Hasan ◽  
Vanessa S. Y. Tay ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Vascular Remodeling ◽  
Ex Vivo ◽  
Vascular Density ◽  
Rapid Progression ◽  
Peripheral Insulin Resistance ◽  
Diphtheria Toxin Receptor ◽  
Toxin Receptor ◽  
The Impact

β-Cells respond to peripheral insulin resistance by first increasing circulating insulin during diabetes. Islet remodeling supports this compensation, but its drivers remain poorly understood. Infiltrating macrophages have been implicated in late-stage type 2 diabetes, but relatively little is known on islet resident macrophages, especially during compensatory hyperinsulinemia. We hypothesized that islet resident macrophages would contribute to islet vascular remodeling and hyperinsulinemia during diabetes, the failure of which results in a rapid progression to frank diabetes. We used chemical (clodronate), genetics (CD169-diphtheria toxin receptor mice), or antibody-mediated (colony-stimulating factor 1 receptor α) macrophage ablation methods in diabetic (db/db) and diet-induced models of compensatory hyperinsulinemia to investigate the role of macrophages in islet remodeling. We transplanted islets devoid of macrophages into naïve diabetic mice and assessed the impact on islet vascularization. With the use of the above methods, we showed that macrophage depletion significantly and consistently compromised islet remodeling in terms of size, vascular density, and insulin secretion capacity. Depletion of islet macrophages reduced VEGF-A secretion in both human and mouse islets ex vivo, and this functionally translated to delayed revascularization upon transplantation in vivo. We revealed that islet resident macrophages were associated with islet remodeling and increased insulin secretion during diabetes. This suggests utility in harnessing islet macrophages during this phase to promote islet vascularization, remodeling, and insulin secretion.

scholarly journals Glucose Intolerance and Impaired Insulin Secretion in Pancreas-Specific Signal Transducer and Activator of Transcription-3 Knockout Mice Are Associated with Microvascular Alterations in the Pancreas

Endocrinology ◽  
2010 ◽  
Vol 151 (5) ◽  
pp. 2050-2059 ◽  
Author(s):  
Elena Kostromina ◽  
Natalia Gustavsson ◽  
Xiaorui Wang ◽  
Chun-Yan Lim ◽  
George K. Radda ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Glucose Intolerance ◽  
Endocrine Pancreas ◽  
Isolated Islets ◽  
Signal Transducer ◽  
Microvascular Network ◽  
Impaired Insulin Secretion ◽  
Stat3 Signaling ◽  
Impaired Insulin

Maintenance of glucose homeostasis depends on adequate amount and precise pattern of insulin secretion, which is determined by both β-cell secretory processes and well-developed microvascular network within endocrine pancreas. The development of highly organized microvasculature and high degrees of capillary fenestrations in endocrine pancreas is greatly dependent on vascular endothelial growth factor-A (VEGF-A) from islet cells. However, it is unclear how VEGF-A production is regulated in endocrine pancreas. To understand whether signal transducer and activator of transcription (STAT)-3 is involved in VEGF-A regulation and subsequent islet and microvascular network development, we generated a mouse line carrying pancreas-specific deletion of STAT3 (p-KO) and performed physiological analyses both in vivo and using isolated islets, including glucose and insulin tolerance tests, and insulin secretion measurements. We also studied microvascular network and islet development by using immunohistochemical methods. The p-KO mice exhibited glucose intolerance and impaired insulin secretion in vivo but normal insulin secretion in isolated islets. Microvascular density in the pancreas was reduced in p-KO mice, along with decreased expression of VEGF-A, but not other vasotropic factors in islets in the absence of pancreatic STAT3 signaling. Together, our study suggests that pancreatic STAT3 signaling is required for the normal development and maintenance of endocrine pancreas and islet microvascular network, possibly through its regulation of VEGF-A.

scholarly journals Optimized murine lung preparation for detailed structural evaluation via micro-computed tomography

2012 ◽  
Vol 112 (1) ◽  
pp. 159-166 ◽  
Author(s):  
Dragoş M. Vasilescu ◽  
Lars Knudsen ◽  
Matthias Ochs ◽  
Ewald R. Weibel ◽  
Eric A. Hoffman
Keyword(s):  
In Vivo Imaging ◽  
Airway Pressure ◽  
Ex Vivo ◽  
Three Dimensional ◽  
Imaging Method ◽  
Small Airways ◽  
Micro Computed Tomography ◽  
Vascular Perfusion ◽  
Murine Lung

Utilizing micro-X-ray CT (μCT) imaging, we sought to generate an atlas of in vivo and intact/ex vivo lungs from normal murine strains. In vivo imaging allows visualization of parenchymal density and small airways (15–28 μm/voxel). Ex vivo imaging of the intact lung via μCT allows for improved understanding of the three-dimensional lung architecture at the alveolar level with voxel dimensions of 1–2 μm. μCT requires that air spaces remain air-filled to detect alveolar architecture while in vivo structural geometry of the lungs is maintained. To achieve these requirements, a fixation and imaging methodology that permits nondestructive whole lung ex vivo μCT imaging has been implemented and tested. After in vivo imaging, lungs from supine anesthetized C57Bl/6 mice, at 15, 20, and 25 cmH2O airway pressure, were fixed in situ via vascular perfusion using a two-stage flushing system while held at 20 cmH2O airway pressure. Extracted fixed lungs were air-dried. Whole lung volume was acquired at 1, 7, 21, and >70 days after the lungs were dried and served as validation for fixation stability. No significant shrinkage was observed: +8.95% change from in vivo to fixed lung ( P = 0.12), −1.47% change from day 1 to day 7 ( P = 0.07), −2.51% change from day 1 to day 21 ( P = 0.05), and −4.90% change from day 1 to day 70 and thereafter ( P = 0.04). μCT evaluation showed well-fixed alveoli and capillary beds correlating with histological analysis. A fixation and imaging method has been established for μCT imaging of the murine lung that allows for ex vivo morphometric analysis, representative of the in vivo lung.

scholarly journals Islet macrophages drive islet vascular remodeling and compensatory hyperinsulinemia in the early stages of diabetes

2019 ◽  
Author(s):  
Manesh Chittezhath ◽  
Divya Gunaseelan ◽  
Xiaofeng Zheng ◽  
Riasat Hasan ◽  
Vanessa SY Tay ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Insulin Secretion ◽  
Vascular Remodeling ◽  
Ex Vivo ◽  
Vascular Density ◽  
Rapid Progression ◽  
List Type ◽  
The Impact

Abstractβ-cells respond to peripheral insulin resistance by increasing circulating insulin in early type-2 diabetes (T2D). Islet remodeling supports this compensation but the drivers of this process remain poorly understood. Infiltrating macrophages have been implicated in late stage T2D but relatively little is known on islet resident macrophages, especially in early T2D. We hypothesize that islet resident macrophages contribute to islet vascular remodeling and hyperinsulinemia, the failure of which results in a rapid progression to T2D. Using genetic and diet-induced models of compensatory hyperinsulinemia we show that its depletion significantly compromises islet remodeling in terms of size, vascular density and insulin secretion capacity. Depletion of islet macrophages reduces VEGF-A secretion from both human and mouse islets ex vivo and the impact of reduced VEGF-A functionally translates to delayed re-vascularization upon transplantation in vivo. Hence, we show a new role of islet resident macrophages in the context of early T2D and suggest that there is considerable utility in harnessing islet macrophages to promote islet remodeling and islet insulin secretion capacity.HighlightsThe compensatory hyperinsulinemic phase of type-2 diabetes is accompanied with significant pancreatic islet remodeling.Bona fide islet resident macrophages are increased during the diabetic compensation phase by largely in situ proliferation.Ablating macrophages severely compromises the islet remodeling process and exacerbates glycemic control in vivo.Mouse and human islet macrophages contribute VEGF-A to the islet environment.Specific removal of islet macrophages delays islet vascularization in compensatory hyperinsulinemic mice.

Abstract PO039: Novel in vivo imaging method to evaluate "Don't eat me" signal of tumor against microglia

2021 ◽  
Author(s):  
Takahiro Tsuji ◽  
Hiroaki Wake ◽  
Mariko Shindo ◽  
Daisuke Kato ◽  
Hiroaki Ozasa ◽  
...  
Keyword(s):  
In Vivo Imaging ◽  
Imaging Method
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