Islet function and insulin release: a matter of β-cell maturity

In a recent study conducted in the laboratory of Dr Hodson at the Institute of Metabolism and Systems Research (IMSR, University of Birmingham, UK) and published in Nature Communications (Nasteska et al, Nat Commun, 2021), subtle differences in β-cell maturity were found to contribute to islet function and insulin release. Recombinant genetic and chemogenetic disruption strategies were used to alter the ratio between PDX1LOW/MAFALOW immature β-cells and PDX1HIGH/MAFAHIGH mature β-cells in the islet. Of note, Pancreas/duodenum homeobox protein 1 (PDX1) and V-maf musculoaponeurotic fibrosarcoma oncogene homolog A (MAFA) are transcription factors whose expression levels are critical for β-cell identity, development and maturation. Both PDX1 and MAFA act by binding regulatory elements of the insulin gene driving, which is important for glucose-stimulated insulin secretion.

In the adult pancreatic islet, there is a heterogeneous population of mature and immature b-cells. Nasteska et al demonstrate that it is crucial to preserve the heterogeneity in β-cell maturity for the maintenance of normal islet function. In particular, a key islet function is glucose-regulated insulin secretion, which, at the cellular level, is triggered by Ca2+ channel opening in association with glucose metabolism and ADP/ATP generation. In brief, the authors provide evidence that both PDX1LOW/MAFALOW and PDX1HIGH/MAFAHIGH β-cells contribute to proper islet function and insulin release. Noteworthy, the study also has an impact on understanding the mechanisms behind type II diabetes (i.e. a disease characterized by a high blood sugar level over a prolonged period of time) since this metabolic disorder occurs if b-cells in the pancreas are not able to release enough insulin to drive glucose into the cells.

In Nasteska et al., CrestOptics X-Light spinning disk was used to acquire images from rodent and human islets. Specifically, fast live imaging was used to perform Ca2+ flux analysis, ATP/ADP ratio and cAMP level measurements over time in different experimental conditions (see section “Microscopy Methods” for further details). For the in-depth functional analysis carried out in the work, please click here to read the whole publication.

Here we show examples of spinning disk Ca2+ live imaging experiments (Figure A) and analysis (Figure B) similar to experiments more extensively reported in Nasteska et al. paper.

Figure A: Representative movie from fast multicellular spinning disk Ca2+ imaging experiments on islets (acquisition every 125 msec; scale bar, 30 µm).

Maximum Intensity Projection in time

(Rainbow dark LUTs)

Movie

(denoising algorithm applied)

Figure B: Representative analysis of Ca2+ flux analysis on specific cells of interest (A-E) using the genetically-encoded Ca2+ indicator, GCaMP6. GCaMP6 fluorescence intensity from spinning disk acquisitions was measured at 3 and 17 minutes (min) in response to glucose (G) and after KCl stimulus (20 min).

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Microscopy methods

For Ca2+ imaging, islets were loaded with Fluo8 (AAT Bioquest Cat# 21082-AAT) or Fura2 (HelloBio HB0780-1mg), or adenovirally-transduced with GCaMP6m, before imaging using a Crest X-Light spinning disk system coupled to a Nikon Ti-E base and 10 x / 0.4 / air or 25 x / 0.8 / air objective.  

ATP/ADP imaging was performed as for Fluo8, except islets were infected with adenovirus harboring the ATP/ADP sensor, Perceval (a kind gift from Prof. Gary Yellen, Harvard), for 48 h.

For cAMP imaging, islets were infected with adenovirus harboring Epac2-camps (a kind gift from Prof. Dermot Cooper, Cambridge) and analyzed using a Crest X-Light spinning disk system.

All experiments were carried out in an ethical manner and approved by corresponding Research Ethics Committees.

Reference

PDX1LOW MAFALOW β-cells contribute to islet function and insulin release

Daniela Nasteska, Nicholas H. F. Fine, Fiona B. Ashford, Federica Cuozzo, Katrina Viloria, Gabrielle Smith, Aisha Dahir, Peter W. J. Dawson, Yu-Chiang Lai, Aimée Bastidas-Ponce, Mostafa Bakhti, Guy A. Rutter, Remi Fiancette, Rita Nano, Lorenzo Piemonti, Heiko Lickert, Qiao Zhou, Ildem Akerman & David J. Hodson

Nature Communications, 12(1):674 (2021 Jan 29) doi: 10.1038/s41467-020-20632-z

Any content re-use from the article is done under the terms and conditions of the Creative Commons Attribution (CC BY) license.

 

Images and data courtesy of David J. Hodson.

Acknowledgement to Lorenzo Piemonti and Rita Nano (Milan, Italy) for the islet preparation shown here.

 

The application note has been prepared in collaboration with David J. Hodson and Cairn Research

Institute of Metabolism and Systems Research (IMSR), University of Birmingham, Birmingham, UK.

Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK

Centre for Endocrinology, Diabetes and Metabolism, Birmingham Health Partners, Birmingham, UK

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