Proteomic analysis of conditioned media from glucose responsive and glucose non‐responsive phenotypes reveals a panel of secreted proteins associated with beta cell dysfunction

Abstract

Media conditioned by dysfunctioning pancreatic beta cells offer an excellent source of potential protein markers associated with this phenotype. Proteins identified from cell culture model systems are often found to be of importance clinically. Previous work by us and others have shown that low-passage MIN-6 cells (MIN-6(L)) respond to changes in glucose concentrations, producing an approximately 5.5-fold glucose-stimulated insulin secretion (GSIS) in response to 26.7 mmol/L, compared with 3.3 mmol/L, glucose. After continuous culture or high-passage (MIN-(H)), this GSIS was no longer present and thus represents an excellent model system for investigating beta cell dysfunction. Employing 2-D difference gel electrophoresis and mass spectrometry a panel of protein markers were identified in conditioned media (CM) from MIN-6(L) and MIN-6(H) beta cells. These proteins, including secretogranin II, secretogranin III and transthyretin, are associated with secretory granule biogenesis and were found to have substantially increased levels in the CM from the non-responsive high-passage MIN-6 beta cells. A panel of protein markers found to have increased abundance levels in CM from MIN-6(H) compared with MIN-6(L) beta cells may have the potential to be used clinically for assessing beta cell function and to monitor the effects of specific therapeutics.

Systematic Proteomic Analysis Identifies β-Site Amyloid Precursor Protein Cleaving Enzyme 2 and 1 (BACE2 and BACE1) Substrates in Pancreatic β-Cells

“…6C). Thus, all major islet hormones (insulin, glucagon, somatostatin and pancreatic polypeptide) were detected among other known secretory proteins such as granins (chromogranins and secretogranins), transthyretin (TTR) (35), islet amyloid polypeptide (iAPP) (36), and neuropeptide Y (NPY) (37). In addition several secreted factors were identified of which the roles have yet to be studied in the pancreatic ␤-cell, including vitamin D-binding protein (DBP), a multifunctional plasma protein of which altered circulating levels are associated with type 1 diabetes (38).…”

Transthyretin: the servant of many masters

“…In type 2 diabetes where there is a reduced beta cell number and function, there are contradictory reports on the plasma TTR levels compared to controls. Some studies (with very small patient numbers) indicate that they are increased, while others suggest that they are decreased [65–67] (Buxbaum, unpublished). More studies with larger cohorts are necessary first to establish the facts and then to interpret them.…”

Microarray Analysis of Novel Candidate Genes Responsible for Glucose-Stimulated Insulin Secretion in Mouse Pancreatic β Cell Line MIN6

“…In addition, comparative proteomic analyses of the cell lysates [7] and the conditioned media [8] were used to investigate the relative changes in the proteome that accompany the loss of GSIS over time in culture. Recently, impairment of glucose and lipid oxidation was suggested to be involved in the loss of GSIS in high passage MIN6 cells [9].…”