Chemokines Are Selectively Regulated during the Progression of Acute Inflammation
DL Horton, DG Remick. Boston University Medical School, Boston, MA
Background: Upon traumatic or infectious injury, secreted cytokines and chemokines display distinct kinetic profiles during the inflammatory response. However, it remains unclear whether this distinction confers differential regulation of inflammation. Using human whole blood as a model system, we examined the effects of delayed DEX on LPS-induced cytokines and chemokines. We hypothesized that chemokines are selectively regulated during the progression of acute inflammation in the whole blood model.
Design: Venous blood was collected from healthy volunteers into heparinzed (10U/ml) syringes. 50 ng/ml LPS or RPMI 1640 was added to the blood in 1.5 ml Eppendorf tubes, followed by continuous mixing with 5% CO2 and ambient air at 37oC. Blood was spun at 1000 x g for 5 minutes at various time points and plasma collected and stored at 20oC for later cytokine analysis. For dex experiments, 10-6M dex was added to blood simultaneously with LPS at time 0, or after 6 h.r LPS stimulation. Plasma was collected at various time points and stored until later analysis. These studies have been approved by the Institutional Review Board of Boston University.
Results: Blood gas analysis revealed unchanged partial pressure of O2 for up to 24 hours, while pCO2 increased from 50 3 to 74 4 mmHg. Glucose and pH decreased significantly (99 6 to 0.50.2 mg/dL, and 7.30.02 to 7.00.02, respectively) between 0 and 24 h. Cellular viability remained unchanged after 24 hours of incubation with DEX. LPS (50 ng/ml) stimulation induced TNF, IL-1, and IL-6, which peaked by 6 hours. In contrast, chemokines (GRO and IL-8) increased steadily up to 24 hours. Maximal TNF, IL-1, and IL-6 mRNA levels were observed at 2 hours and were undetectable by 6 hours. GRO and IL-8 mRNA displayed an initial peak at 2 h, and subsequently decreased by 12 hours. Between 12 and 24 hours, chemokine mRNA increased steadily. Simultaneous DEX (10-6M) and LPS resulted in significant suppression of cytokines and chemokines at 24 hours. However, delaying DEX until 6 hours after LPS abrogated its suppression of TNF, IL-1, and IL-6. In contrast, delayed DEX significantly suppressed both GRO and IL-8 levels.
Conclusions: In conclusion, these data suggest that the regulation of progressive inflammation occurs via modulation of chemokines, but not cytokines, in the whole blood model of acute inflammation. Thus, future experiments should be aimed at targeting therapies specifically to chemokines.
Monday, March 9, 2009 1:00 PM
Poster Session II # 199, Monday Afternoon