In situ RT-PCR detection of inducible nitric oxide synthetase gene expression in lung during endotoxemia in rabbits
- Author:
Mei WAN
1
;
LINGYihng
;
Bin CONG
;
Junxia WANG
;
Lei JIN
;
Fenglian HAN
Author Information
1. Hebei Medical University
- Keywords:
endotatixemia;
inducible nitric oxide sunthetase;
gene erpression;
in situ RT PCR
- From:
Chinese Medical Journal
2000;113(12):1133-1137
- CountryChina
- Language:Chinese
-
Abstract:
Oblectlv To detect the Iocation of inducible nitlic oxide synthetase (iNOS) protein and mRNA in lung during endotoxemia in rabbits. Methods blotting was perfomed before, 1 hour and 5 houm after the intravenous administration of lipolysaccharide (LPS) in rabbits. Immuno histonbernical analysls (IA), in situ hybridization and in situ ireverse transcfipfiorl-polymerase chain reaction (in sltoJ RT-PCR) were also pedormed in lung sections. Results iNOS mRNA expression was found using Nodhem blotting in lung 5 hours after LPS injection, while It was not found in contrei. Tbe positive stain was found only in macrophages in lung 5 hours after LPS inljecfion by standard hybddization and IA; while by in situ RT- PCR, tbe amplifation products were found in macrophaoes, aJlway epitheiial oells, vascular endotbeliei cells, smooth muscle cells and leukocytes, in addition to macrophages distributed abundantly thriygout the lung. The signal was absent in control or samples. Cencluslons Using an in situ RT-PCR technique, iNOS expressson was riot only observed in macriogaqss also in many other kinds of cells in ybg during endotoxemla in rabbits. This suggests that n situ RT-PCR ls much more sensirtive than in situ hybridization, and can be usad to examine gones with Iow expreseion.
