Altered expression of thin filament-associated proteins in hypertrophied urinary bladder smooth muscle.
OBJECTIVE
Obstruction of the urinary bladder outlet induces detrusor smooth muscle (DSM) hypertrophy. The goal of this study was to determine whether the composition of thin filament-associated proteins, known to play important roles in cytoskeletal structure and/or the regulation of contraction, is altered in DSM during hypertrophy.


METHODS
DSM hypertrophy was induced in male rabbits by partial ligation of the urethra. Sham-operated rabbits served as a control. Reverse transcriptase-polymerase chain reaction (RT-PCR) and real-time PCR revealed a significant increase in the expression of mRNAs for basic (h1) calponin (CaP), and alpha-isoform of tropomyosin (Tm) in hypertrophied DSM compared to controls. Western blotting and two-dimensional (2-D) gel electrophoresis showed enhanced expression of these proteins and also a significant increase in the expression of beta-non muscle and gamma-smooth muscle actin in the DSM from obstructed bladders, while alpha-actin remained constant.


RESULTS
Enhanced expression of these proteins in the DSM from obstructed bladders was confirmed by immunofluorescence microscopy. Double immunostaining with Cap/Tm and alpha/beta-actin-specific antibodies showed co-localization of these proteins in myocytes. Colocalization of smooth muscle specific myosin and CaP to cytoplasmic filaments in cells dissociated from the hypertrophied DSM indicated that these cells are differentiated smooth muscle cells.


CONCLUSIONS
The change in the isoforms of actin, Cap, and Tm may be part of the molecular mechanism for bladder compensation in increased urethral resistance.
Mannikarottu
Anita S
AS
Division of Urology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Disanto
Michael E
ME
Zderic
Stephen A
SA
Wein
Alan J
AJ
Chacko
Samuel
S
D000199:Actins / Q000096:biosynthesis; D000818:Animals; D002135:Calcium-Binding Proteins / Q000096:biosynthesis; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D006984:Hypertrophy; D066298:In Vitro Techniques; D007381:Intermediate Filament Proteins / Q000096:biosynthesis*; D008297:Male; D008840:Microfilament Proteins; D008856:Microscopy, Fluorescence; D009130:Muscle, Smooth / Q000473:pathology / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014335:Tropomyosin / Q000096:biosynthesis; D001743:Urinary Bladder / Q000473:pathology / Q000502:physiology*; D001748:Urinary Bladder Neck Obstruction / Q000473:pathology; D000096985:Calponins
D000199:Actins; D002135:Calcium-Binding Proteins; D007381:Intermediate Filament Proteins; D008840:Microfilament Proteins; D012333:RNA, Messenger; D014335:Tropomyosin; D000096985:Calponins