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Hypoxia and endotoxin induce macrophage-mediated suppression of fibroblast proliferation
Abstract:
Cellular mechanisms and environmental factors contributing to wound failure following shock and wound contamination are unclear. The activation of macrophages by exposure to hypoxia (pO2 less than 20) and/or lipopolysaccharide (10 micrograms/ml) in vitro was investigated for its effect on macrophage regulation of fibroblast proliferation. The effect on fibroblast proliferation of conditioned medium from activated murine macrophages or co-culture with activated macrophages was tested by measuring 3T3 fibroblast incorporation of 3H-thymidine and culture DNA content. Unstimulated macrophages produced growth factors that increase fibroblast proliferation (proliferation index (PI) = 1.4 +/- 0.15, p less than 0.05 vs. control). Activation by hypoxia alone had little effect on macrophage regulation of fibroplasia (PI = 1.55 +/- 0.28, N.S. vs. unstimulated macrophages). LPS activated macrophages suppressed fibroplasia and the combination of hypoxia with LPS augmented the suppression (PI = 0.5 +/- 0.11, LPS alone, p less than 0.05 and 0.25 +/- 0.05, LPS + hypoxia, p less than 0.01). In addition, hypoxia + LPS treated co-cultures had reduced DNA contents, suggesting reduced cell numbers (12.5 +/- 2.6 micrograms vs. 8.2 +/- 2.0 micrograms). We screened several macrophage cytokines for their direct effect on 3T3 proliferation and found that mr-Tumor Necrosis Factor-alpha (150 units) also suppressed proliferation. Conditioned supernatants from LPS activated macrophages contained 12 +/- 2 units of mrTNF as measured by L929 cytolysis; however, this was significantly less than required to induce suppression of proliferation by direct addition. The regulatory role of the macrophage appears to be dependent on its level of activation. Activation by hypoxia and LPS altered macrophage regulation of fibroblast proliferation from stimulation to suppression.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Macrophage activation by lipopolysaccharide (LPS) and hypoxia alters their regulation of fibroblast proliferation, shifting from stimulation to suppression. This finding is crucial for understanding wound healing and failure in contaminated wounds.
Area of Science:
- Wound healing research
- Cellular immunology
- Biomedical engineering
Background:
- Wound failure after shock and contamination involves complex cellular and environmental factors.
- The role of macrophages in regulating fibroblast proliferation during wound healing is not fully understood.
Purpose of the Study:
- To investigate how macrophage activation by hypoxia and lipopolysaccharide (LPS) affects fibroblast proliferation.
- To elucidate the cellular mechanisms underlying wound healing regulation.
Main Methods:
- Murine macrophages were activated in vitro using hypoxia (low oxygen) and/or LPS.
- Fibroblast proliferation was assessed by measuring 3H-thymidine incorporation and DNA content in 3T3 fibroblasts.
- Conditioned media and co-culture systems were used to evaluate macrophage-fibroblast interactions.
Main Results:
- Unstimulated macrophages promoted fibroblast proliferation.
- LPS activation, particularly combined with hypoxia, significantly suppressed fibroblast proliferation and reduced cell numbers.
- Tumor Necrosis Factor-alpha (TNF-α) was identified as a factor involved in this suppression, though its direct effect was less potent than that of activated macrophages.
Conclusions:
- Macrophage regulatory function on fibroblast proliferation is dependent on activation state.
- Hypoxia and LPS co-activation induce a shift in macrophage behavior, suppressing rather than stimulating fibroblast proliferation.
- These findings offer insights into impaired wound healing mechanisms in complex clinical scenarios.