Generation of human pancreatic beta cell lines and studies of islet cell

Abstract

Effective drug discovery begins with effective in vitro experimentation. Whilst many disease states, such as cancer, have a plethora of biologically relevant cell lines, diabetes research does not have this luxury. Although, rodent beta cell lines are available for in vitro research, their use to effectively mimic human disease can be debated. This project aims to improve upon existing cell lines, 1.1B4 and 1.4E7, by the overexpression of INS and/or PDX1.

Overexpression of INS and/or PDX1 generated stably transfected 1.1B4 derived cell lines with enhanced insulin secretion and responsiveness to secretagogues. Unfortunately, questions surrounding 1.1B4 authenticity arose and therefore it was necessary to discontinue experimentation with these novel cell lines.

1.4E7 cells and the overexpression of INS and/or PDX1, resulted in the generation of novel1.4E7 derived cell lines. Initial investigations identified an optimal clone from each transfection. Subsequently, these identified clones were pitched against each other to determine the ultimate clone derived from 1.4E7 cells.

Dual transfection with INS and PDX1 successfully generated a novel cell line capable of enhanced insulin secretion in response to glucose and decreased insulin secretion in response to inhibitors. 1.4E7_DUAL Clone 15 was elucidated as the ultimate clone derived from transfection of 1.4E7 cells, achieving the primary aim of this project.

A secondary aim of this project, chicken islets were employed as an effective model of alpha cell function. Early research on alpha cell toxins was conducted prior to modern experimental methodologies and therefore potentially effective treatments maybe long forgotten. Recent unpublished investigations have revitalised local interest in synthalin. In vitro experimentation with synthalin on alpha-TC1.9 cells failed to demonstrate it as a selective alpha cell toxin, as historically believed. However, a combination of in vivo and ex vivo supports, synthalin is metabolised in vivo and a metabolite is a selective alpha cell toxin.

Date of Award	May 2023
Original language	English
Supervisor	Neville Mc Clenaghan (Supervisor) & Peter Flatt (Supervisor)