Regulation of adherence-stimulated HSP-70 mRNA expression in alveolar macrophages

A. Lopez, S. A. Moore, A. Richardson

Research output: Contribution to journalArticle

Abstract

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 languageEnglish (US)
JournalJournal of Investigative Medicine
Volume44
Issue number1
StatePublished - 1996

Fingerprint

HSP70 Heat-Shock Proteins
Alveolar Macrophages
Mercaptoethanol
Messenger RNA
Macrophages
Suspensions
Antioxidants
Chemical activation
Cytochalasins
Dithiothreitol
Scavenging
Inbred F344 Rats
Cycloheximide
Vitamin E
Phagocytosis
Sulfhydryl Compounds
Nitric Oxide Synthase
Indomethacin
Northern Blotting
Ascorbic Acid

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Regulation of adherence-stimulated HSP-70 mRNA expression in alveolar macrophages. / Lopez, A.; Moore, S. A.; Richardson, A.

In: Journal of Investigative Medicine, Vol. 44, No. 1, 1996.

Research output: Contribution to journalArticle

@article{533fde4e74224c3ca052b0af47059c0e,
title = "Regulation of adherence-stimulated HSP-70 mRNA expression in alveolar macrophages",
abstract = "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.",
author = "A. Lopez and Moore, {S. A.} and A. Richardson",
year = "1996",
language = "English (US)",
volume = "44",
journal = "Journal of Investigative Medicine",
issn = "1081-5589",
publisher = "Lippincott Williams and Wilkins",
number = "1",

}

TY - JOUR

T1 - Regulation of adherence-stimulated HSP-70 mRNA expression in alveolar macrophages

AU - Lopez, A.

AU - Moore, S. A.

AU - Richardson, A.

PY - 1996

Y1 - 1996

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=33749576162&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33749576162&partnerID=8YFLogxK

M3 - Article

AN - SCOPUS:33749576162

VL - 44

JO - Journal of Investigative Medicine

JF - Journal of Investigative Medicine

SN - 1081-5589

IS - 1

ER -