Investigating the suspension culture on aggregation and function of mouse pancreatic β‐cells

Abstract

The integrity and hierarchical structure of islet influence β-cells physiology dramatically. A culture substrate which can maintain or improve β-cells aggregation shall benefit cell therapy for diabetics. In this study, nontreated, type IV collagen, Lipidure, and ultralow attachment dishes were used to culture a murine β-cell line, MIN-6. The formation and biological performances of pseudoislets were investigated. Results showed that β-cells formed loose and irregular aggregates on nontreated dishes. Oppositely, pseudoislets formed on other three substrates. Most pseudoislets on Lipidure and type IV collagen dishes had a diameter between 100-150 μm with high survival rate, while large pseudoislets (>250 μm) with seriously central necrosis were found on ultralow attachment dishes. Western blot analysis revealed that pseudoislets had relatively higher connexin 36 protein productions relative to single cells. The glucose-stimulated insulin secretion test showed pseudoislets on type IV collagen have high stimulation index. Monolayers from TCPS dishes and pseudoislets from type IV collagen or Lipidure dishes were further transplanted into diabetic mice. Animals received both single cells and pseudoislets had decreasing blood glucose level and regained body weight. Histologic examination revealed that all implants successfully engrafted with positive insulin staining. Interestingly, the area under curve for the intraperitoneal glucose tolerance test showed pseudoislets had superior glucose disappearance rate. This study reveals that isolated islets or insulin-producing cells can be cultured on type IV collagen or Lipidure dishes to improve/maintain integrity prior to transplantation.

A multiple-funnels cell culture insert for the scale-up production of uniform cell spheroids

“…Similarly, Yang et al. reported serious central apoptosis in cell spheres cultured on ultra-low attachment and bacterial (non-treated polystyrene) dishes [38] . Using an oxygen-permeable material to fabricate a cell spheroid formation device, Anada et al.…”

Engineering Functional Pseudo-Islets of Defined Sizes from Primary Murine Cells Using PEG Microwell Devices

“…These hydrogel microwell devices possess many benefits over alternative methods of β-cell aggregation, such as the hanging drop method [33,49] and spontaneous aggregations [14,30,50], including simplicity of fabrication, high fidelity of pattern transfer across a range of sizes and shapes, and the ease of aggregate culture and removal due to use of PEG, a polymer commonly used in biomaterials for it's high water content and bio-inert nature. We have translated the use of these microwells from mouse insulinoma 6 (MIN6) cells, a β-cell line, to primary islets.…”

A biomimetic honeycomb‐like scaffold prepared by flow‐focusing technology for cartilage regeneration

“…After being cultured for 7 days, chondrocytes in microfluidic scaffolds were underwent live staining in media containing calcein‐AM (C3099, Invitrogen Corp., Carlsbad, CA) for 30 min to assess cell survival (Yang et al, ). The chondrocytes/microfluidic scaffold constructs were subsequently stained with 2 µmol/L of DAPI (sc‐3598, Santa Cruz Biotechnology, Dallas, TX) for 7 min.…”