THE EFFECTS OF LPS AND PROBUCOL ON INTERLEUKIN-1 (IL-1) AND PLATELET-DERIVED GROWTH-FACTOR (PDGF) GENE-EXPRESSION IN THE HUMAN MONOCYTIC CELL-LINE U-937
Sr. Li et al., THE EFFECTS OF LPS AND PROBUCOL ON INTERLEUKIN-1 (IL-1) AND PLATELET-DERIVED GROWTH-FACTOR (PDGF) GENE-EXPRESSION IN THE HUMAN MONOCYTIC CELL-LINE U-937, Biochimica et biophysica acta. Molecular basis of disease, 1225(3), 1994, pp. 271-274
We have investigated the effects of lipopolysaccharide (LPS) and probu
col (a lipid soluble antioxidant) on the gene expression of interleuki
n 1 alpha and beta (IL-1 alpha and IL-1 beta), and platelet-derived gr
owth factor A chain and B chain (PDGF-A and PDGF-B) in the human monoc
ytic cell line U-937. Steady-state mRNA levels were measured quantitat
ively by the reverse transcription-polymerase chain reaction (RT-PCR)
using a non-radioactive label. Cells were incubated with LPS, in the p
resence or absence of probucol for 20 h. The cells were harvested and
RNA was then prepared, reverse-transcribed in the presence of an inter
nal standard and subsequently amplified and labelled with digoxigenin-
11-dUTP by the PCR reaction. The PCR products were subjected to agaros
e gel electrophoresis, blotted onto nylon membranes and visualised by
immunological detection of digoxigenin followed by a chemiluminescent
reaction. We found that: (1) LPS treatment of U-937 cells caused an up
-regulation of gene expression of IL-1 beta and PDGF-A chain by approx
imately 250% and 100% respectively, although it did not stimulate the
expression of IL-1 alpha nor PDGF-B chain mRNA. (2) Probucol treatment
in vitro had no effect on the basal or LPS-stimulated mRNA levels of
IL-1 alpha, IL-1 beta, PDGF-A and PDGF-B despite its reported activity
in vivo. (3) PDGF-A and PDGF-B were expressed at a similar level in u
nstimulated U-937 cells approximately 10-50 copies/ng total RNA, where
as the expression of IL-1 beta mRNA was approximately 2-4 times higher
than IL-1 alpha mRNA. (4) Finally, in U-937 monocytic cells the expre
ssion of IL-1 alpha and IL-1 beta, and PDGF-A and PDGF-B appear to be
independently regulated.