Co-Culture of α TC-6 Cells and β TC-1 Cells: Morphology and Function

Abstract

Background: In vitro experiments using only β-cell lines instead of islets are limited because pancreatic islets are composed of four different types of endocrine cells. Several recent studies have focused on cellular interactions among these cell types, especially α- and β-cells. Because islet isolation needs time and experience, we tested a simple co-culture system with α- and β-cells. Their morphology and function were assessed by comparison to each single cell culture and pancreatic islets.

Methods: α TC-6 cells and β TC-1 cells were maintained in Dulbecco's Minimal Essential Medium containing 5 mM glucose and 10% fetal bovine serum. Cells were mixed at a 1:1 ratio (5×10⁵) in 6-well plates and cultured for 24, 48, and 72 hours. After culture, cells were used for insulin and glucagon immunoassays and tested for glucose-stimulated insulin secretion (GSIS).

Results: α TC-6 and β TC-1 cells became condensed by 24 hours and were more strongly compacted after 48 hours. β TC-1 cells showed both β-β and β-α cell contacts. GSIS increased with increasing glucose concentration in co-cultured cells, which showed lower secreted insulin levels than β TC-1 cells alone. The increase in the secreted insulin/insulin content ratio was significantly lower for co-cultured cells than for β-cells alone (P=0.04). Compared to islets, the α-/β-cell co-culture showed a higher ratio of GSIS to insulin content, but the difference was not statistically significant (P=0.09).

Conclusion: α TC-6 and β TC-1 cells in the co-culture system showed cell-to-cell contacts and a similar stimulated insulin secretion pattern to islets. The co-culture system may be used to better mimic pancreatic islets in in vitro assessments.

Keywords: Insulin; α-Cell; β-Cell.

Fig. 1. Gyratory shaker for cell mixing in co-culture.

Fig. 2. Morphology in the co-culture at (A) 2 hours and (B) 48 hours, as observed using an optical microscope (×200).

Fig. 3. Cellular composition of the aggregates, as studied by immunohistochemical staining for insulin and glucagon at (A) 24 hours, (B) 48 hours, and (C) 72 hours. Red, glucagon; green, insulin; blue, DAPI.

Fig. 4. Cellular composition of the aggregates in co-culture at (A) 24, (B) 48, and (C) 72 hours.

Fig. 5. Glucose-stimulated insulin secretion from β TC-1 cells and co-cultured cells.

Fig. 6. Increase in secreted insulin/insulin content in β TC-1 cells, co-cultured cells, and islets.