Effects Of Prostaglandin A₁ On Cytokines And Nuclear Factor Kappa B

Objective To evaluate anti-inflammatory property of PGA1, effects ofPGA1 pretreatment on LPS-inducible expression of inflammatorycytokines and LPS-mediated nuclear translocation of NF- Κ B wereinvestigated. Methods Peripheral venous blood was collected fromhealthy volunteers into heparin anticoagulant solution. PBMC were thenpartially purified from red blood cells by sedimentation withHetastarch, and further purified according to the method ofFicoll-Hypaque. Then divided into two parts. Part 1: PBMC wereincubated with PGA1 of different concentrations ( 4, 8, 12, 16 and 20μmol/L) for 4h before LPS (10 μmol/L) stimulated. Supernatants werecollected at 8 and 24 h after start of stimulation with LPS fordetermination of TNF-α, IL-1β and IL-6 by ELISA. Part 2: PBMC addedto six-well plates were randomly divided into three groups: ①control,② LPS alone, and ③ PGA1 pretreatment. Cells in the LPS alone groupwere treated with LPS for 2 h. Cells in the PGA1 pretreatment group wereincubated with PGA1 for 4h before LPS stimulated. After treatment withLPS for 2h, protein levels of NF-ΚB in the nucleus and cytoplasm wereevaluated by western bolt analysis . Results In the LPS alone group,the concentrations of all the cytokines studied were significant 5广西医科大学 2001 级硕士研究生学位论文increased , compared with those in the control group. At 8 and 24 hafter LPS stimulated, PGA1 caused a reduction of the production ofTNF-α, IL-1βand IL-6 induced by LPS. These inhibitory effects of PGA1were dose-dependent. At a final concentration of 12 μmol/L, PGA1significantly inhibited LPS-induced release of all the inflammatorycytokines tested. Compared with control group, NF-ΚB level wassignificantly enhanced in the nucleus and decreased in the cytoplasmin the LPS alone group, indicating that LPS mediated nucleartranslocation of NF-ΚB. Prior treatment with PGA1 inhibited theincrease in nucleus protein level for NF-ΚB which was significant at12 and 20μmol/L. This inhibition occurred in dose-dependent manner.Conclusions These results suggest that pretreatment with PGA1dose-dependently inhibits release of the proinflammatory cytokines andinhibited LPS-mediated nuclear translocation of NF- Κ B. Thisinhibition by PGA1 may attenuate LPS-elicited NF- Κ B activity,resulting in suppressing LPS-induced cytokine release from PBMC.