About me
Optimizing NIT-1 Cell Culture Conditions
Location: 1
Mentor: Dr. Alice Tomei
NIT-1 insulinoma cell clusters, derived from non-obese diabetic mice, are promising candidates for type 1 diabetes (T1D) research due to their low production cost, availability (immortalized line), and consistent insulin secretion in response to glucose stimulation. These pseudoislets can be used in vitro to study the interactions between islets and immune cells at the islet transplant site in beta cell replacement therapies for T1D treatment. To ensure accurate experimental results, I aimed to optimize the culture conditions to maximize viability and functionality of these NIT-1 clusters during 48-hour in vitro culture. Obtaining consistently and reproducibly high cell functionality is critical for future studies aimed at testing the effects of immune cells and immunomodulatory drugs on NIT-1 cluster viability and functionality in an in vitro model of the islet transplant site. Various 96-well plates, including gas permeable and U-bottom plates, were tested under static and dynamic (shaking incubator) culture conditions. NIT-1 functionality was assessed using the gold standard Glucose Stimulated Insulin Secretion (GSIS) assay. Insulin secretion in cell culture supernatants was quantified by ELISA. Stimulation indexes (S.I.) were computed by dividing the insulin secretion under high glucose (stimulated) to the insulin secretion under low glucose (basal) conditions. While fresh cells showed the higher functionality (S.I.: 5.148), our results indicated that gas permeable plates maintained higher cell functionality (S.I. of 2.121) than U-bottom plates in static (S.I.: 1.386) or dynamic (S.I.: 1.298) conditions. Therefore, gas permeable 96-well plates are recommended for in vitro experiments involving NIT-1 pseudoislets to ensure optimal viability and functionality, likely due to their higher oxygen permeability.
