Using pancreas tissue slices for in situ studies of islet of Langerhans and acinar cell biology

Marciniak, Anja; Cohrs, Christian M.; Tsata, Vasiliki; Chouinard, Julie A.; Selck, Claudia; Stertmann, Julia; Reichelt, Saskia; Rose, Tobias; Ehehalt, Florian; Weitz, Jürgen; Solimena, Michele; Rupnik, Marjan Slak; Speier, Stephan

doi:10.1038/nprot.2014.195

Cited by 99 publications

(126 citation statements)

References 44 publications

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“…Our approach employs a pancreatic slice method (Marciniak et al, 2014) that preserves the native islet organisation, unlike the more widely used islet cultures which can cause structural changes [e.g. an increase in tight junctions induced by the enzyme treatments (Intveld et al, 1984)] and loss of endothelial cells (Lukinius et al, 1995).…”

Section: Discussionmentioning

confidence: 99%

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Cell polarity defines three distinct domains in pancreatic beta cells

Gan

Zavortink

Ludick

et al. 2016

Journal of Cell Science

147

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The structural organisation of pancreatic β-cells in the islets of Langerhans is relatively unknown. Here, using three-dimensional (3D) two-photon, 3D confocal and 3D block-face serial electron microscopy, we demonstrate a consistent in situ polarisation of β-cells and define three distinct cell surface domains. An apical domain located at the vascular apogee of β-cells, defined by the location of PAR-3 (also known as PARD3) and ZO-1 (also known as TJP1), delineates an extracellular space into which adjacent β-cells project their primary cilia. A separate lateral domain, is enriched in scribble and Dlg, and colocalises with E-cadherin and GLUT2 (also known as SLC2A2). Finally, a distinct basal domain, where the β-cells contact the islet vasculature, is enriched in synaptic scaffold proteins such as liprin. This 3D analysis of β-cells within intact islets, and the definition of distinct domains, provides new insights into understanding β-cell structure and function.

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Section: Discussionmentioning

confidence: 99%

“…Here, we have used a pancreatic slice preparation that maintains the native structural organisation of the islets (Marciniak et al, 2014). We have imaged in three dimensions, using three distinct methods; live-cell two-photon microscopy, immunofluorescence confocal microscopy and, finally, serial block-face electron microscopy.…”

Section: Introductionmentioning

confidence: 99%

Cell polarity defines three distinct domains in pancreatic beta cells

Gan

Zavortink

Ludick

et al. 2016

Journal of Cell Science

147

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“…Subsequently, the gel-embedded islets were cultured for 24 h in indicated conditions. Calcium imaging was performed in a custommade perifusion system (30). Briefly, coverslips with gelembedded islets were moved into a temperature-controlled perifusion chamber where the temperature was set at 37°C.…”

Section: In Vitro Imaging Of B-cell Cytosolic Free Calcium Dynamicsmentioning

confidence: 99%

Alterations in β-Cell Calcium Dynamics and Efficacy Outweigh Islet Mass Adaptation in Compensation of Insulin Resistance and Prediabetes Onset

et al. 2016

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Emerging insulin resistance is normally compensated by increased insulin production of pancreatic β-cells, thereby maintaining normoglycemia. However, it is unclear whether this is achieved by adaptation of β-cell function, mass, or both. Most importantly, it is still unknown which of these adaptive mechanisms fail when type 2 diabetes develops. We performed longitudinal in vivo imaging of β-cell calcium dynamics and islet mass of transplanted islets of Langerhans throughout diet-induced progression from normal glucose homeostasis, through compensation of insulin resistance, to prediabetes. The results show that compensation of insulin resistance is predominated by alterations of β-cell function, while islet mass only gradually expands. Hereby, functional adaptation is mediated by increased calcium efficacy, which involves Epac signaling. Prior to prediabetes, β-cell function displays decreased stimulated calcium dynamics, whereas islet mass continues to increase through prediabetes onset. Thus, our data reveal a predominant role of islet function with distinct contributions of triggering and amplifying pathway in the in vivo processes preceding diabetes onset. These findings support protection and recovery of β-cell function as primary goals for prevention and treatment of diabetes and provide insight into potential therapeutic targets.

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“…This technique allowed the study of islet- and β-cell-specific genes [16, 28, 29] and proteins [30]. LCM enables the extraction of samples from an environment which is well conserved and close to the natural condition, to better investigate cell physiology [31], cell biology [32], cell transcriptome [13], and proteome [30]. Islets obtained with this method are expected to reflect closely the in vivo molecular composition and pathology of in situ islets.…”

Section: Introductionmentioning

confidence: 99%

Proteomic profiling of human islets collected from frozen pancreata using laser capture microdissection

Zhang

Lanzoni

Battarra

et al. 2017

Journal of Proteomics

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The etiology of Type 1 Diabetes (T1D) remains elusive. Enzymatically isolated and cultured (EIC) islets cannot fully reflect the natural protein composition and disease process of in vivo islets, because of the stress from isolation procedures. In order to study islet protein composition in conditions close to the natural environment, we performed proteomic analysis of EIC islets, and laser capture microdissected (LCM) human islets and acinar tissue from fresh-frozen pancreas sections of three cadaveric donors. 1104 and 706 proteins were identified from 6 islets equivalents (IEQ) of LCM islets and acinar tissue, respectively. The proteomic profiles of LCM islets were reproducible within and among cadaveric donors. The endocrine hormones were only detected in LCM islets, whereas catalytic enzymes were significantly enriched in acinar tissue. Furthermore, high overlap (984 proteins) and similar function distribution were found between LCM and EIC islets proteomes, except that EIC islets had more acinar contaminants and stress-related signal transducer activity proteins. The comparison among LCM islets, LCM acinar tissue and EIC islets proteomes indicates that LCM combined with proteomic methods enables accurate and unbiased profiling of islet proteome from frozen pancreata. This paves the way for proteomic studies on human islets during the progression of T1D.

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Using pancreas tissue slices for in situ studies of islet of Langerhans and acinar cell biology

Cited by 99 publications

References 44 publications

Cell polarity defines three distinct domains in pancreatic beta cells

Cell polarity defines three distinct domains in pancreatic beta cells

Alterations in β-Cell Calcium Dynamics and Efficacy Outweigh Islet Mass Adaptation in Compensation of Insulin Resistance and Prediabetes Onset

Proteomic profiling of human islets collected from frozen pancreata using laser capture microdissection

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