Delineating biological pathways unique to embryonic stem cell-derived insulin-producing cell lines from their noninsulin-producing progenitor cell lines.
To identify unique biochemical pathways in embryonic stem cell-derived insulin-producing cells as potential therapeutic targets to prevent or delay beta-cell dysfunction or death in diabetic patients, comparative genome-wide gene expression studies of recently derived mouse insulin-producing cell lines and their progenitor cell lines were performed using microarray technology. Differentially expressed genes were functionally clustered to identify important biochemical pathways in these insulin-producing cell lines. Biochemical or cellular assays were then performed to assess the relevance of these pathways to the biology of these cells. A total of 185 genes were highly expressed in the insulin-producing cell lines, and computational analysis predicted the pentose phosphate pathway (PPP), clathrin-mediated endocytosis, and the peroxisome proliferator-activated receptor (PPAR) signaling pathway as important pathways in these cell lines. Insulin-producing ERoSHK cells were more resistant to hydrogen peroxide (H(2)O(2))-induced oxidative stress. Inhibition of PPP by dehydroepiandrosterone and 6-aminonicotinamide abrogated this H(2)O(2) resistance with a concomitant decrease in PPP activity as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Clathrin-mediated endocytosis, which is essential in maintaining membrane homeostasis in secreting cells, was up-regulated by glucose in ERoSHK but not in their progenitor ERoSH cells. Its inhibition by chlorpromazine at high glucose concentration was toxic to the cells. Troglitazone, a PPARG agonist, up-regulated expression of Ins1 and Ins2 but not Glut2. Gene expression analysis has identified the PPP, clathrin-mediated endocytosis, and the PPAR signaling pathway as the major delineating pathways in these insulin-producing cell lines, and their biological relevance was confirmed by biochemical and cellular assays.
Chen
Tian Sheng
TS
Institute of Medical Biology, Agency for Science, Technology, and Research, Singapore.
Tan
Soon Sim
SS
Yeo
Ronne Wee Yeh
RW
Teh
Bao Ju
BJ
Luo
Ruihua
R
Li
GuoDong
G
Lim
Sai Kiang
SK
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics*; D002460:Cell Line; D016000:Cluster Analysis; D004622:Embryo, Mammalian; D053595:Embryonic Stem Cells / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D004705:Endocytosis / Q000187:drug effects / Q000235:genetics; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology*; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D010427:Pentose Phosphate Pathway / Q000187:drug effects / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics*
D006861:Hydrogen Peroxide; D005947:Glucose