Gene expression variance based on random sequencing in rat remnant kidney.
BACKGROUND
Several examinations have been performed to identify the genes involved in chronic renal failure using 5/6 nephrectomized rats. Recently, many systematic techniques for examining molecular expression have been developed. They might also be effective in elucidating the molecular mechanism of progressive renal failure. In this study, digital expression profiling was carried out to construct a subtractive mRNA expression database for the 5/6 nephrectomized kidney.


METHODS
One thousand clones were randomly sequenced from 5/6 nephrectomized and sham-operated rat kidney cDNA libraries, respectively, and defined by BLAST search. In silico subtractive analysis was performed to search for genes up- or down-regulated in the 5/6 nephrectomized kidney.


RESULTS
The growth factor-related mRNAs and the mRNAs encoding cytoskeletal or membrane proteins were up-regulated, but the transporter-related mRNAs were down-regulated in the 5/6 nephrectomized kidney database. In silico subtraction revealed that 63 mRNAs were increased and 59 were decreased in the 5/6 nephrectomized kidney. To confirm whether the in silico subtractive database reflected the actual expression of mRNA or protein, 12 known genes were examined by Northern blotting or immunoblotting, respectively. The actual expression of the 12 genes was comparable with the results of in silico subtraction. In addition, we successfully isolated five unknown genes, two up-regulated and three down-regulated in the 5/6 nephrectomized kidney.


CONCLUSIONS
We constructed a subtractive mRNA expression database for 5/6 nephrectomized kidney, which reflects the actual alterations in mRNA expression after subtotal nephrectomy. This database may be useful for elucidation of the molecular mechanism of progressive renal failure.
Horiba
Naoshi
N
Department of Pharmacy, Kyoto University Hospital, Faculty of Medicine, Kyoto University, Sakyo-ku, Kyoto, Japan.
Masuda
Satohiro
S
Takeuchi
Ayako
A
Saito
Hideyuki
H
Okuda
Masahiro
M
Inui
Ken-Ichi
K
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics; D003598:Cytoskeletal Proteins / Q000235:genetics; D030541:Databases, Genetic; D015536:Down-Regulation; D015870:Gene Expression; D020869:Gene Expression Profiling; D014644:Genetic Variation; D006133:Growth Substances / Q000235:genetics; D007668:Kidney / Q000503:physiopathology*; D008297:Male; D008565:Membrane Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D009392:Nephrectomy / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis; D011897:Random Allocation; D051381:Rats; D017208:Rats, Wistar; D015854:Up-Regulation
D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D006133:Growth Substances; D008565:Membrane Proteins