TY - JOUR
T1 - Cytokine-inducible nitric oxide synthase (iNOS) expression in cardiac myocytes
T2 - Characterization and regulation of iNOS expression and detection of iNOS activity in single cardiac myocytes in vitro
AU - Balligand, Jean Luc
AU - Ungureanu-Longroisi, Dan
AU - Simmons, William W.
AU - Pimental, David
AU - Malinski, Tadeusz A.
AU - Kapturczak, Matthias
AU - Taha, Ziad
AU - Lowenstein, Charles J.
AU - Davidoff, Amy J.
AU - Kelly, Ralph A.
AU - Smith, Thomas W.
AU - Michel, Thomas
PY - 1994/11/4
Y1 - 1994/11/4
N2 - Cellular constituents of heart muscle contain both constitutive and inducible nitric oxide (NO) signaling pathways that modulate the contractile properties of cardiac myocytes. The identities of the inducible NO synthase (iNOS) isoform(s) expressed in cardiac muscle, and of the specific cell types expressing iNOS activity, remain poorly characterized. We amplified a 217-base pair cDNA by reverse transcriptase-polymerase chain reaction from primary cultures of inflammatory cytokine-pretreated adult rat ventricular myocytes (ARVM) that was nearly identical to other iNOS cDNA sequences. Using this 217-base pair cDNA as a probe in Northern blots, we found no evidence of iNOS mRNA in control myocytes, but both interleukin-1β and interferon-γ individually increased iNOS mRNA abundance in primary cultures of ARVM, with maximal expression at 12 h. The half-life of iNOS mRNA in actinomycin C1-treated cells was 4 h. Both dexamethasone and transforming growth factor-β attenuated the induction of iNOS mRNA abundance and enzyme activity by IL-1β and INFγ. Pretreatment with dexamethasone also abolished the induction of iNOS mRNA, but not the increase in GTP cyclohydrolase mRNA in purified cardiac myocytes from lipopolysaccharide-injected rats. In order to further characterize the specific cell type producing NO, we used a NO-specific porphyrinic/Nafion-coated microsensor to record NO release from a single, isolated ARVM pretreated with IL-1β and IFNγ in L-arginine-depleted medium. NO release could be detected following microinjection of L-arginine in the vicinity of the cell juxtaposed to the NO microsensor, but not following microinjection of D-arginine, and not from ARVM pretreated with L-N-monomethylarginine. Cytokine-pretreated ARVM that had been maintained in L-arginine-depleted medium also exhibited a depressed contractile response to isoproterenol after addition of L-arginine, but not o-arginine. These results indicate that altered contractile function of cardiac myocytes following exposure to specific inflammatory cytokines is due to induction of myocyte iNOS.
AB - Cellular constituents of heart muscle contain both constitutive and inducible nitric oxide (NO) signaling pathways that modulate the contractile properties of cardiac myocytes. The identities of the inducible NO synthase (iNOS) isoform(s) expressed in cardiac muscle, and of the specific cell types expressing iNOS activity, remain poorly characterized. We amplified a 217-base pair cDNA by reverse transcriptase-polymerase chain reaction from primary cultures of inflammatory cytokine-pretreated adult rat ventricular myocytes (ARVM) that was nearly identical to other iNOS cDNA sequences. Using this 217-base pair cDNA as a probe in Northern blots, we found no evidence of iNOS mRNA in control myocytes, but both interleukin-1β and interferon-γ individually increased iNOS mRNA abundance in primary cultures of ARVM, with maximal expression at 12 h. The half-life of iNOS mRNA in actinomycin C1-treated cells was 4 h. Both dexamethasone and transforming growth factor-β attenuated the induction of iNOS mRNA abundance and enzyme activity by IL-1β and INFγ. Pretreatment with dexamethasone also abolished the induction of iNOS mRNA, but not the increase in GTP cyclohydrolase mRNA in purified cardiac myocytes from lipopolysaccharide-injected rats. In order to further characterize the specific cell type producing NO, we used a NO-specific porphyrinic/Nafion-coated microsensor to record NO release from a single, isolated ARVM pretreated with IL-1β and IFNγ in L-arginine-depleted medium. NO release could be detected following microinjection of L-arginine in the vicinity of the cell juxtaposed to the NO microsensor, but not following microinjection of D-arginine, and not from ARVM pretreated with L-N-monomethylarginine. Cytokine-pretreated ARVM that had been maintained in L-arginine-depleted medium also exhibited a depressed contractile response to isoproterenol after addition of L-arginine, but not o-arginine. These results indicate that altered contractile function of cardiac myocytes following exposure to specific inflammatory cytokines is due to induction of myocyte iNOS.
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M3 - Article
C2 - 7525557
AN - SCOPUS:0028135094
SN - 0021-9258
VL - 269
SP - 27580
EP - 27588
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 44
ER -