Identification of variations of gene expression of visceral adipose and renal tissue in type 2 diabetic rats using cDNA representational difference analysis.
OBJECTIVE
To identify differences in gene expression in renal and visceral adipose tissue in type 2 diabetic rats using cDNA representational difference analysis (RDA) and to explore the molecular pathogenesis of type 2 diabetes and its chronic vascular complications.


METHODS
A rat model of type 2 diabetes was generated by administration of a high fat and calorie diet combined with a low dose of streptozocin (STZ) injected into the tail vein. The difference bands were generated by cDNA representational difference analysis (cDNA RDA). The final difference products were ligated into the pUC-18 vector and sequenced. A bioformatics analysis was performed on the obtained expressed sequence tags (ESTs), and then the expression levels of known and novel genes were verified by semi-quantitative reverse transcription-PCR (RT-PCR). At the same time, full-length cDNA of a novel gene was cloned in silico.


RESULTS
The type 2 diabetic rats in this experiment experienced hyperglycemia, lipidemia, lower insulin sensitivity and normal body weight. We obtained 9 novel ESTs and 2 novel genes from renal tissue of rats and 6 novel ESTs and 1 known gene, the rat lipoprotein lipase (LPL) gene from their visceral adipose tissue. The 2 novel genes (RS91 and RS2) from the renal tissue were both very similar to serine (or cysteine) proteinase inhibitor, clade F and eukaryotic translation initiation factor 3 and subunit 5 (EIF-3 epsilon). The expression of both novel genes and the LPL gene were upregulated in renal and visceral adipose tissue of type 2 diabetic and fat-enriched rats. Full-length cDNA of the novel gene RS91 was cloned in silico.


CONCLUSIONS
(1) The rat model of type 2 diabetes generated in this study was ideal because the disease in the animals closely mimicked type 2 diabetic patients. (2) cDNA RDA is a flexible, inexpensive, more accurate, sensitive and highly effective technique for identifying differences in gene expression. (3) Six novel ESTs and 1 known gene were obtained from rat visceral adipose tissue. The LPL gene was upregulated in adipose tissue of type 2 diabetic and fat-enriched rats, a result possibly related to the diabetic animals' high fat and calorie diet, lipidemia, insulin resistance (RI) and molecular pathogenesis. (4) Nine novel ESTs and 2 novel genes were obtained from the renal tissue of rat. We believe the 2 novel genes to be the serine proteinase inhibitors clade F and EIF-3 epsilon in rats. The upregulation of the 2 novel genes in renal tissue of type 2 diabetic rats and may have been related to their renal impairment.
Yang
Jialin
J
Institute of Endocrinology, Shanghai Ruijin Hospital, Shanghai Second Medical University, Shanghai 200025, China.
Li
Guo
G
Zhang
Fanglin
F
Liu
Youping
Y
Zhang
Di
D
Zhou
Wenzhong
W
Xu
Guangwu
G
Yang
Yisheng
Y
Luo
Min
M
D000273:Adipose Tissue / Q000378:metabolism*; D000818:Animals; D003001:Cloning, Molecular; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism*; D020224:Expressed Sequence Tags; D020869:Gene Expression Profiling; D007668:Kidney / Q000378:metabolism*; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis; D010957:Plasmids; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014781:Viscera