TY - JOUR
T1 - Nitric oxide synthase induction and intestinal epithelial cell viability in rats
AU - Tepperman, B. L.
AU - Brown, J. F.
AU - Whittle, Brendan J. R.
PY - 1993/8/1
Y1 - 1993/8/1
N2 - The present study determined the presence of constitutive and inducible nitric oxide (NO) synthase activities in intestinal isolated epithelial cells and the effects of NO induction on intestinal epithelial cell viability. Epithelial cells were isolated from rat proximal small intestine by dispersion using citrate and EDTA. Constitutive NO synthase activity, determined by [14C]arginine conversion to citrulline that was inhibited by in vitro incubation with the arginine analogue N(G)-monomethyl-L-arginine (L- NMMA; 300 μM) or ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'- tetraacetic acid (EGTA; 1 mM), was observed in these cells. Administration of Escherichia coli lipopolysaccharide (LPS; 3 mg/kg iv) significantly augmented NO synthase activity (determined 4 h later), which was inhibited in vitro by incubation with L-NMMA but not by EGTA. The highest level of constitutive and inducible NO synthase activity occurred in intestinal villus cells, and the lowest was in crypt cells. Induction of NO synthase activity was associated with a decrease in cellular viability as assessed by a decrease in trypan blue exclusion. Dexamethasone pretreatment (1 mg/kg iv 2 h before LPS administration) significantly reduced both induction of NO synthase activity and the reduction in cellular viability. Likewise concurrent administration of the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (10 mg/kg sc) ameliorated the reduction in cell viability induced by LPS administration, an effect abolished by pretreatment with the NO substrate L-arginine (350 mg/kg sc). Whereas constitutively formed NO may have a physiological role in these cells, the results in this study suggest that induction of NO synthase in epithelial cells may represent a mechanism of local intestinal damage.
AB - The present study determined the presence of constitutive and inducible nitric oxide (NO) synthase activities in intestinal isolated epithelial cells and the effects of NO induction on intestinal epithelial cell viability. Epithelial cells were isolated from rat proximal small intestine by dispersion using citrate and EDTA. Constitutive NO synthase activity, determined by [14C]arginine conversion to citrulline that was inhibited by in vitro incubation with the arginine analogue N(G)-monomethyl-L-arginine (L- NMMA; 300 μM) or ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'- tetraacetic acid (EGTA; 1 mM), was observed in these cells. Administration of Escherichia coli lipopolysaccharide (LPS; 3 mg/kg iv) significantly augmented NO synthase activity (determined 4 h later), which was inhibited in vitro by incubation with L-NMMA but not by EGTA. The highest level of constitutive and inducible NO synthase activity occurred in intestinal villus cells, and the lowest was in crypt cells. Induction of NO synthase activity was associated with a decrease in cellular viability as assessed by a decrease in trypan blue exclusion. Dexamethasone pretreatment (1 mg/kg iv 2 h before LPS administration) significantly reduced both induction of NO synthase activity and the reduction in cellular viability. Likewise concurrent administration of the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester (10 mg/kg sc) ameliorated the reduction in cell viability induced by LPS administration, an effect abolished by pretreatment with the NO substrate L-arginine (350 mg/kg sc). Whereas constitutively formed NO may have a physiological role in these cells, the results in this study suggest that induction of NO synthase in epithelial cells may represent a mechanism of local intestinal damage.
KW - cell viability
KW - dexamethasone
KW - intestinal cell
KW - lipopolysaccharide
UR - http://www.scopus.com/inward/record.url?scp=0027161939&partnerID=8YFLogxK
U2 - 10.1152/ajpgi.1993.265.2.G214
DO - 10.1152/ajpgi.1993.265.2.G214
M3 - Article
C2 - 7690186
AN - SCOPUS:0027161939
SN - 0002-9513
VL - 265
SP - G214-G218
JO - American Journal of Physiology - Gastrointestinal and Liver Physiology
JF - American Journal of Physiology - Gastrointestinal and Liver Physiology
IS - 2
ER -