Type

Journal Article

Authors

Martin Clynes
Sweta Rani
Per Bendix Jeppesen
Eoin Ryan
Eadaoin McKiernan
Patrick Gammell
Lorraine O'Driscoll

Subjects

Biochemistry

Topics
cell proliferation cell line reverse transcriptase polymerase chain reaction cytology biological markers secretion humans cells insulin secreting cells gene expression regulation gene expression passage chemistry metabolism insulin alkaline phosphatase phenotype oligonucleotide array sequence analysis cell differentiation dose response relationship drug profile pharmacology stem cells glucose global proinsulin gene expression profiling

Phenotypic and global gene expression profile changes between low passage and high passage MIN-6 cells. (2006)

Abstract The long-term potential to routinely use replacement beta cells/islets as cell therapy for type 1 diabetes relies on our ability to culture such cells/islets, in vitro, while maintaining their functional status. Previous beta cell studies, by ourselves and other researchers, have indicated that the glucose-stimulated insulin secretion (GSIS) phenotype is relatively unstable, in long-term culture. This study aimed to investigate phenotypic and gene expression changes associated with this loss of GSIS, using the MIN-6 cell line as model. Phenotypic differences between MIN-6(L, low passage) and MIN-6(H, high passage) were determined by ELISA (assessing GSIS and cellular (pro)insulin content), proliferation assays, phase contrast light microscopy and analysis of alkaline phosphatase expression. Differential mRNA expression was investigated using microarray, bioinformatics and real-time PCR technologies. Long-term culture was found to be associated with many phenotypic changes, including changes in growth rate and cellular morphology, as well as loss of GSIS. Microarray analyses indicate expression of many mRNAs, including many involved in regulated secretion, adhesion and proliferation, to be significantly affected by passaging/ long-term culture. Loss/reduced levels, in high passage cells, of certain transcripts associated with the mature beta cell, together with increased levels of neuron/glia-associated mRNAs, suggest that, with time in culture, MIN-6 cells may revert to an early (possibly multi-potential), poorly differentiated, 'precursor-like' cell type. This observation is supported by increased expression of the stem cell marker, alkaline phosphatase.
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Full list of authors on original publication

Martin Clynes, Sweta Rani, Per Bendix Jeppesen, Eoin Ryan, Eadaoin McKiernan, Patrick Gammell, Lorraine O'Driscoll

Experts in our system

1
Martin Clynes
Dublin City University
Total Publications: 209
 
2
Sweta Rani
Trinity College Dublin
Total Publications: 22
 
3
Eoin Ryan
University College Dublin
 
4
Lorraine O'Driscoll
Trinity College Dublin
Total Publications: 152