MOLECULAR ANALYSIS OF COLLAGENS IN BLADDER FIBROSIS

Purpose: Fibrosis of bladder tissue is characterized by an abnormal de position of connective tissue within different layers of the bladder w all, resulting in a low volume, high pressure vesical which may ultima tely contribute to renal scarring and failure. These bladders are func tionally referred to as ''non-compliant'' and may result from differen t etiologies: neurogenic, which encompasses myelodysplasia and spinal cord injury, or non-neurogenic, owing to obstruction or radiation ther apy. To examine the molecular mechanisms responsible for this fibrosis , we have analyzed a well-characterized pediatric patient population f or alteration(s) in collagen types I and III regulation at the protein and nucleic acid levels. Materials and Methods: Immunohistochemical l ocalization of collagen subtypes (I, III, and IV) was carried out usin g type specific monoclonal antibodies. Total collagen was determined b y hydroxyproline analysis, and subtype specific expression of collagen ous proteins, following cyanogen bromide extraction procedures, was qu antified by competitive ELISA. Total RNA was extracted by guanidinium/ phenol/chloroform, and slot blot hybridization analyses with radiolabe led human cDNA probes were quantified by densitometry of resulting aut oradiograms.Results: Connective tissue infiltration of detrusor smooth muscle bundles was specific for type III collagen. Protein analyses d emonstrated: 1) an increase in total collagen, 2) a statistically sign ificant increase in the type III: type I collagen ratio, and 3), an ab solute increase in type III collagen protein in non-compliant bladder tissue. At the mRNA level, there was a coordinate increase in both col lagen I and III steady-state mRNAs in non-neurogenic bladder tissue, w hereas neurogenic bladder tissue was characterized by an increase in t he type III: type I mRNA transcript ratio. Conclusions: These data sug gest that regulation of collagen synthesis in bladder fibrosis is comp lex and is characterized by both transcriptional and post-transcriptio nal mechanisms, depending upon the etiology of the fibrosis.