Characterization of Human Pancreatic Islet Cells using a Single-Cell Western Blot Platform.

Abstract

OBJECTIVE Islet transplantation is an effective treatment for type 1 diabetes. However, transplant success depends on quick islet assessment because islets deteriorate 2-3 days after isolation. A new tool, single-cell Western blot (scWestern), offers results within one day. In this study, we aimed to test the suitability of scWestern to detect protein markers for beta (insulin), alpha (glucagon), and delta (somatostatin) cells, the three major endocrine cell types in islets. METHODS We characterized the antibody specificity, signal intensity, and cell identification on the scWestern platform, and then compared the islet cell composition analysis between scWestern and immunohistochemistry performed by the Integrated Islet Distribution Program (IIDP). RESULTS Islet cell composition is comparable for alpha and beta cells, but not delta cells. Protein expression levels of insulin, glucagon, and somatostatin in individual islet cells varied greatly, highlighting cell type heterogeneity. Surprisingly, scWestern revealed double-hormonal cells (~1%), co-expressing insulin and somatostatin or insulin and glucagon, in non-diabetic and non-obese adult human islets, which was confirmed by confocal immunofluorescence microscopy. CONCLUSIONS These results demonstrate that each alpha, beta, and delta cells express varying levels of peptide hormones, and a small subpopulation co-expresses double hormones in normal human islets. The scWestern platform will enable timely assessment of beta cell mass in isolated islets before clinical transplantation.