Functional analysis of a conditionally transformed pancreatic beta-cell line
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Published on: June 8, 2010
N Fleischer1, C Chen, M Surana
1Department of Medicine, Albert Einstein College of Medicine, Bronx, New York, USA. fleisch@aecom.yu.edu
Abstract:
Development of beta-cell lines for cell therapy of diabetes is hindered by functional deviations of the replicating cells from the normal beta-cell phenotype. In a recently developed cell line, denoted betaTC-tet, derived from transgenic mice expressing the SV40 T antigen (Tag) under control of the tetracycline (Tc) gene regulatory system, growth arrest can be induced by shutting off Tag expression in the presence of Tc. Here, we compared differentiated cell functions in dividing and growth-arrested betaTC-tet cells, both in culture and in vivo. Proliferating cells stably maintained normal glucose responsiveness for >60 passages in culture. Growth-arrested cells survived for months in culture and in vivo and maintained normal insulin production and secretion. After growth arrest, the cells gradually increased their insulin content three- to fourfold. This occurred without significant changes in insulin biosynthetic rates. At high passage numbers, proliferating betaTC-tet cells exhibited an abnormal increase in hexokinase expression. However, the upregulation of hexokinase was reversible upon growth arrest. Growth-arrested cells transplanted intraperitoneally into syngeneic recipients responded to hyperglycemia by a significant increase in insulin secretion. These findings demonstrate that transformed beta-cells maintain function during long periods of growth arrest, suggesting that conditional transformation of beta-cells may be a useful approach for developing cell therapy for diabetes.
Insights
Conditional transformation of beta cells using the betaTC-tet cell line allows for sustained function during growth arrest. This approach shows promise for developing effective cell therapies for diabetes by maintaining insulin production and glucose responsiveness.
Area of Science:
- Biotechnology
- Endocrinology
- Cell Biology
Background:
- Developing functional beta-cell lines for diabetes cell therapy is challenging due to phenotypic drift in replicating cells.
- The betaTC-tet cell line, derived from transgenic mice, allows for conditional control of SV40 T antigen (Tag) expression via tetracycline (Tc).
Purpose of the Study:
- To compare the differentiated cell functions of dividing versus growth-arrested betaTC-tet cells in vitro and in vivo.
- To assess the potential of conditional beta-cell transformation for diabetes cell therapy.
Main Methods:
- Culturing and in vivo transplantation of betaTC-tet cells under conditions of proliferation and growth arrest.
- Assessment of insulin production, secretion, glucose responsiveness, and hexokinase expression.
- Monitoring cell survival and function over extended periods and passages.
Main Results:
- Proliferating betaTC-tet cells maintained glucose responsiveness for over 60 passages.
- Growth-arrested cells exhibited stable insulin production and secretion in vitro and in vivo for months.
- Insulin content increased 3-4 fold after growth arrest without altered biosynthesis; abnormal hexokinase expression in proliferating cells was reversible upon growth arrest.
- Transplanted growth-arrested cells responded to hyperglycemia with increased insulin secretion.
Conclusions:
- Transformed beta-cells retain essential functions during prolonged growth arrest.
- Conditional transformation offers a viable strategy for generating functional beta-cell lines for diabetes cell therapy.
