Macrophages respond to both environmental (temperature, ethanol, metals, lipopolysaccharides) and physiological (cytokines, phagocytosis, oxygen radicals) cues in the production of heat shock protein 70 (hsp70). The macrophage is terminally differentiated, and its responses to challenges cause changes in its level of activation and function. In this study, Fischer rat alveolar macrophages (AMs), were incubated in suspension and adherent cultures. Freshly isolated AMs showed low levels of hsp70 mRNA, even when AMs were maintained in suspension cultures. AMs, that are allowed to adhere, undergo an increased level of activation and the function of these cells appears to be directed towards inflammatory and tumorcidal activity. Adherence (ADH) also induced the expression of hsp70. Induction can be detected as early as 30 minutes and peak at 6 hours after ADH. To determine the factors that regulate ADH-induction of hsp70, various agents were added to the cultures and total RNA was analyzed by northern blot analysis. Addition of either cycloheximide (1-10 μg/ml), which inhibits de novo protein synthesis, or indomethacin (0.3-30μM) which inhibits prostaglandin synthesis, had no effect on ADH-induced expression of hsp70. The addition of cytochalasin b (5μg/ml), which disrupts the cytoskeletal structure, did not alter the ADH-induction of hsp 70. The thiol antioxidants, β-2-mercaptoethanol (0.05-5.0mM β-2-ME) and dithiothreitol (DTD, drastically reduced 80-85% of the ADH-induced expression of hsp 70. By altering the time of exposure to β-2-ME, we found that the first 15 minutes of ADH were critical for ADH-induction. Other antioxidants were tested (superoxide dismutase, vitamin C, vitamin E, nitric oxide synthetase inhibitors) and found to have no effect on ADH-induced expression of hsp70. These results suggest that the inhibitory effects of β-2-ME and DTT maybe due to effects on sulfhydryl groups and not due to radical scavenging activities; and may have profound effects on inflammatory and immune processes of AMs.
|Original language||English (US)|
|Journal||Journal of Investigative Medicine|
|State||Published - Jan 1 1996|
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)