Activity and expression of nitric oxide synthase in the hypertrophied rat bladder and the effect of nitric oxide on bladder smooth muscle growth.
OBJECTIVE
We investigated the expression and activity of nitric oxide synthase (NOS) and the localization of cyclic guanosine monophosphate (cGMP) in hypertrophied rat bladder. We also examined whether nitric oxide (NO) has a growth inhibitory effect in bladder smooth muscle cells.


METHODS
The urethra was partly ligated and the bladder was removed 3 days, 3 or 6 weeks after obstruction. NOS activity was determined as the conversion of L-[14C]citrulline from L-[14C]arginine (Amersham Life Science, Solna, Sweden). Neuronal NOS (nNOS) expression was studied with Western blot analysis and immunohistochemistry. The expression of inducible NOS (iNOS) and cGMP was evaluated by immunohistochemistry. The effect of NO on isolated bladder smooth muscle cell growth was assessed as protein and DNA synthesis by [3H]-leucine and [3H]-thymidine (NEN Life Science Products, Zaventem, Belgium) incorporation, respectively.


RESULTS
Ca independent iNOS activity increased after short-term obstruction. Immunohistochemical studies in obstructed bladders demonstrated iNOS expression primarily in urothelial and inflammatory cells. Ca dependent nNOS activity decreased after obstruction, as confirmed by Western blot analysis. The cGMP immunoreactive cells were mainly found within the serosal layer of obstructed bladders. The NO donor DETA-NONOate (Alexis Biochemicals, Lausen, Switzerland) (300 microM.) reduced [3H]-leucine and [ H]-thymidine incorporation by a mean of 29% +/- 2% and 95% +/- 2%, respectively, in cultured bladder smooth muscle cells.


CONCLUSIONS
Bladder obstruction caused a small increase in iNOS activity and a decrease in nNOS activity. NO was found to have a growth inhibitory effect in bladder smooth muscle cells, suggesting that changes in NOS activity may influence the progress of bladder hypertrophy.
Johansson
Rebecka
R
Department of Clinical Pharmacology, Lund University Hospital, Sweden.
Pandita
Raj Kumar
RK
Poljakovic
Mirjana
M
Garcia-Pascual
Angeles
A
De Vente
Jan
J
Persson
Katarina
K
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism; D005260:Female; D006834:Hydrazines / Q000494:pharmacology; D006984:Hypertrophy; D007150:Immunohistochemistry; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects; D009569:Nitric Oxide / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D009603:Nitroso Compounds; D051381:Rats; D017207:Rats, Sprague-Dawley; D001743:Urinary Bladder / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D001748:Urinary Bladder Neck Obstruction / Q000378:metabolism / Q000473:pathology
D006834:Hydrazines; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; C496322:Nos1 protein, rat; C496318:Nos2 protein, rat; D006152:Cyclic GMP; D002118:Calcium