Kinetic study of porcine GM-CSF expression in porcine alveolar macrophages and spleen cells.
Granulocyte-macrophage colony-stimulating factor (GM-CSF) is an important regulator in inducing differentiation and proliferation of immune cells. The functional roles of porcine GM-CSF (pGM-CSF) have not yet been revealed. Therefore, expression patterns of pGM-CSF were investigated in immune cells after cloning and sequencing of whole pGM-CSF cDNA. Whole cDNA of pGM-CSF was amplified from porcine alveolar macrophages stimulated by lipopolysaccharide (LPS), using 5'- and 3'-rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR) methods. The products of 5'- and 3'-RACE-PCR were cloned, and the nucleotide sequence of whole pGM-CSF cDNA was determined (GenBank accession number AY116504). The kinetics of pGM-CSF mRNA expression were studied in porcine immune cells such as alveolar macrophages and spleen cells, using a real-time quantitative PCR. The expression of pGM-CSF in LPS-, phytohemagglutinin (PHA)-, or concanavalin A (ConA)-stimulated cells was always higher as compared to the control cells. The expression levels of pGM-CSF in alveolar macrophages were highest at 5 h after LPS stimulation and then continuously decreased in the late phase. In spleen cells, the LPS-stimulated group showed the highest levels after 5 h, but the PHA- and the ConA-stimulated groups showed slightly increased expression levels at the early phase and peaked at 24 h. To our knowledge, this is the first published report describing the nucleotide sequence of whole cDNA and the expression pattern of pGM-CSF using real-time quantitative PCR. These results indicate that pGM-CSF has its own characteristic expression profile in different immune cells.
Cho
Young Wook
YW
Department of Infectious Diseases, College of Veterinary Medicine and School of Agricultural Biotechnology, Seoul National University, San 56-1, Shilim-dong, Kwanak-ku, Seoul 151-742, South Korea.
Lee
Deog Yong
DY
Shin
Sung Jae
SJ
Shin
Na Ri
NR
Kang
Sang Gyun
SG
Choi
In Soo
IS
Yoo
Han Sang
HS
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003208:Concanavalin A / Q000494:pharmacology; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics*; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D010835:Phytohemagglutinins / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D013154:Spleen / Q000187:drug effects / Q000378:metabolism*; D034421:Sus scrofa
D018076:DNA, Complementary; D008070:Lipopolysaccharides; D010835:Phytohemagglutinins; D003208:Concanavalin A; D016178:Granulocyte-Macrophage Colony-Stimulating Factor