Nitric oxide, enhanced by macrophage-colony stimulating factor, mediates renal damage in reflux nephropathy.
BACKGROUND
Reflux nephropathy (RN) is a major cause of end-stage renal failure in children and young adults. Nitric oxide (NO) is an important mediator of tissue injury and inflammation. NO production is enhanced by hematopoietic growth factor including macrophage colony stimulating factor (M-CSF). M-CSF plays a pivotal role in the development of nephritis via macrophage activation. The aim of this study was to investigate the expression of inducible NO synthase (iNOS) and M-CSF in the refluxing kidney, in order to further understand the pathogenesis of RN.


METHODS
The kidney specimens from 6 patients with severe RN and 6 controls were examined by NADPH-diaphorase histochemistry and immunohistochemistry using ABC method with anti-M-CSF antibody. Double staining using NADPH-diaphorase histochemistry/M-CSF immunohistochemistry and M-CSF/iNOS fluorescence immunohistochemistry also was performed. In situ hybridization was performed using digoxigenin labeled M-CSF specific probe. RT-PCR was performed to evaluate the relative amount of iNOS mRNA expression. Apoptosis was determined using the in situ end-labeling technique.


RESULTS
The most striking difference between tissues from RN patients and controls was the marked increase in NADPH-d activity, iNOS immunoreactivity and mRNA and M-CSF immunoreactivity and mRNA expression in the kidneys of RN patients, particularly in the distal tubules, collecting system. Apoptotic cells were markedly increased in RN compared to controls.


CONCLUSIONS
Our findings suggest that the increase in M-CSF-stimulated local production of nitric oxide may be a major mediator in the development of reflux nephropathy.
Children's Research Centre, Our Lady's Hospital for Sick Children, Dublin, Ireland.
Udo
U
Rolle
Hideki
H
Shima
Prem
P
Puri
D000293:Adolescent; D017209:Apoptosis; D002648:Child; D002675:Child, Preschool; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007672:Kidney Cortex / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D016173:Macrophage Colony-Stimulating Factor / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D009252:NADPH Dehydrogenase / Q000032:analysis; D009569:Nitric Oxide / Q000032:analysis / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000032:analysis / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014718:Vesico-Ureteral Reflux / Q000378:metabolism* / Q000473:pathology*
D012333:RNA, Messenger; D009569:Nitric Oxide; D016173:Macrophage Colony-Stimulating Factor; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D009252:NADPH Dehydrogenase