Endocrine Abstracts

Background: It is widely believed that increased circulatory concentrations of long-chain saturated fatty acids (LC-SFA) significantly contribute to the death and dysfunction of pancreatic β-cells in the development of type 2 diabetes (T2D). The mechanism by which LC-SFA cross the β-cell plasma membrane has not been fully established. This work aims to characterise the mechanism underpinning this uptake. Ultimately, regulation of LC-SFA entry may maintain β-cellviability, thereby slowing, or potentially halting, the progression of T2D.

Methodology: To determine whether the fluorescent LC-SFA analogue, BODIPY FL C₁₆, crosses plasma membranes using simple diffusion, cell-sized giant unilamellar vesicles (GUVs) were formed using microfluidics. RNA-seq analysis, with differential gene expression analysis of six transcriptomics datasets of human-derived EndoC-βH1 β-cells, human islets, adipocytes, and hepatocytes, determined quantitative differences in the expression of candidate LC-SFA transport proteins (CTPs). Changes in CTP gene expression was assessed using qRT-PCR in EndoC-βH1 cells following a 6h exposure to LC-FFA C16:0, C18:0 and C18:1 in low (5.5 mM) and high (20 mM) glucose. The rate of uptake of LC-SFA in EndoC-βH1 cells was assessed using a pHrodo^TM Green AM intracellular pH indicator and dynamin inhibited with Dyngo^TM-4a.

Results: BODIPY FL C₁₆ rapidly permeated GUV membranes. The profile expressions of LC-FFA transport proteins in hepatocytes and adipocytes significantly differed to those of human islets and EndoC-βH1 cells indicating that the mechanism of uptake differs in these cell types. Human islets and EndoC-βH1 cells had a similar pattern of expression with CD36, ACSL1-4 and FATP4 being upregulated in both. Exposure to LC-FFA in high and low glucose did not alter CTP expression. There was a rapid (<20s) uptake of LC-SFA, which was inhibited when the cells were pre-incubated with the dynamin inhibitor.

Conclusion: LC-FFA can cross artificial membranes using simple diffusion, and LC-FFA uptake seemingly occurs via a dynamin-mediated process in β-cells.