Related Experiment Videos
Endotoxin-induced production of interleukin 6 by equine peritoneal macrophages in vitro
D D Morris1, N Crowe, J N Moore
1Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens 30602-7385.
Abstract:
A study was performed to determine whether equine peritoneal macrophages produce interleukin 6 (IL-6) in vitro in response to endotoxin. Peritoneal fluid was collected from 14 clinically normal adult horses and was used as the source of peritoneal macrophages. Macrophages from each horse were isolated and cultured separately in vitro in the absence or presence of various concentrations (0.5, 5, 500 ng/ml) of endotoxin (lipopolysaccharide from Escherichia coli 055:B5). Culture medium supernatants were collected after 3, 6, 12, and 24 hours' incubation and were frozen at -70 C until assayed for IL-6 activity. Supernatant IL-6 activity was determined by use of a modified colorimetric assay and the murine hybridoma cell line B 13.29 clone B.9, which is dependent on IL-6 for survival. Results indicated that equine peritoneal macrophages produce IL-6 in vitro and that supernatant medium IL-6 activity was significantly (P less than 0.05) increased by exposure to endotoxin. Significant (P less than 0.05) time and treatment effects on macrophage IL-6 production were apparent. The IL-6 activity peaked at 6 or 12 hours' incubation, then remained high through 24 hours' incubation, regardless of endotoxin exposure. Medium IL-6 activity during 3 and 6 hours' incubation was significantly (P less than 0.05) greater in macrophages exposed to 5 or 500 ng of endotoxin/ml than in those exposed to 0.5 ng of endotoxin/ml; however peak IL-6 activity was similar among all endotoxin concentrations. Endotoxin concentration did not have an effect on medium IL-6 activity from macrophages exposed to endotoxin for 12 or 24 hours.
Insights
Equine peritoneal macrophages produce interleukin-6 (IL-6) in vitro. Endotoxin exposure significantly increased IL-6 production, with peak activity observed at 6-12 hours post-stimulation.
Area of Science:
- Veterinary Immunology
- Equine Immunology
- Cellular Immunology
Background:
- Equine peritoneal macrophages play a role in local immune responses.
- Interleukin-6 (IL-6) is a key cytokine involved in inflammation and immunity.
Purpose of the Study:
- To investigate the in vitro production of IL-6 by equine peritoneal macrophages.
- To determine the effect of endotoxin on IL-6 production in these cells.
Main Methods:
- Peritoneal macrophages were isolated from healthy horses.
- Macrophages were cultured in vitro with varying concentrations of endotoxin (lipopolysaccharide).
- IL-6 activity in culture supernatants was measured over 24 hours using a bioassay.
Main Results:
- Equine peritoneal macrophages produced IL-6 in vitro.
- Endotoxin significantly increased IL-6 production compared to controls (P < 0.05).
- Peak IL-6 activity occurred at 6-12 hours and remained elevated for 24 hours.
Conclusions:
- Equine peritoneal macrophages are capable of producing IL-6.
- Endotoxin is a potent stimulus for IL-6 release from these cells.
- The findings contribute to understanding equine inflammatory responses.