Hypothesis / aims of study
Intrinsic sphincter deficiency is common in patients with spina bifida have a different pathophysiology and impact than in other neurogenic and non neurogenic populations. The objective of this study was to evaluate the association between five urinary biomarkers (NGF, BDNF, TIMP-2, TGF-B1 and PGE2) and intrinsic sphincter deficiency (ISD) in adults with spina bifida.
Study design, materials and methods
A single center prospective study was conducted between March 2015 and March 2017 including all consecutive adult spina bifida patients who were seen for urodynamic testing. Urinary tract imaging was also performed in all patients. At the end of the inclusion period, urines were defrosted and urinary NGF, BDNF, TIMP-2, TGF-B1 were assessed using validated ELISA kits.
Urine samples were taken from the initial void/catheterization immediately preceding the urodynamic test and frozen at -80°c. Patients were asked to come with a full bladder so that the urinary markers level would be more interpretable.
At the end of the inclusion period, they were defrosted and the urinary marker levels were measured using dedicated ELISA kits: PGE2 (Cayman Chemical) TIMP-2, TGFβ-1, NGF, BDNF (RayBioTech). Briefly, urine samples were pipetted into the wells of a 96-well plate coated with an antibody specific for each marker. If the marker was present in a sample, it was bound to the wells by the immobilized antibody. The wells were washed and a biotinylated anti-marker antibody was added. After washing away unbound biotinylated antibody, HRP-conjugated streptavidin was pipetted to the wells. The wells were washed again, and a TMB substrate solution was added to the wells and color develops in proportion to the amount of marker bound. The Stop Solution changed the color from blue to yellow, and the intensity of the color was measured at 450 nm. Results were adjusted on the urinary creatinine level (Cr). The urinary level of MMP-2 were assessed by zymography, using the Biotrak Activity Assay System
Associations between urinary markers levels and clinical (stress urinary incontinence (SUI)) and/or urodynamic (lowered maximum urethral closure pressure (MUCP)) of intrinsic sphincter deficiency were sought.
Results
Forty patients were included. The mean patients’ age was 37.7 years and there was 52.5% and 48.5% of male and female patients respectively. The majority of patients were self-catheterizing (70%) and the predominant neurological level was lumbar (79%). At baseline, the mean Qualiveen-SF was 1.23 (/4) and the mean maximum voided volume on the voiding diary was 419.7 ml. Most patients were self-catheterizing (70%), and had been on antimuscarinics (62.5%) with nine having an history of previous intradetrusor botulinum toxin injections (22.5%).
17 patients had SUI (42.5%). The mean MUCP was 56.7 cmH2O. Urinary TIMP-2/Cr and urinary BDNF/Cr were the only two markers significantly associated with MUCP (r=-0.37; p=0.02 in both cases, Figure 1). TIMP-2/Cr was significantly higher in the SUI group (p=0.003; ), as was BDNF/Cr (p=0.02). Urinary NGF/Cr, TGF-B1/Cr and MMP2 did not differ significantly between SUI and non-SUI groups.
Interpretation of results
Urinary TIMP-2 and BDNF may be associated with intrinsic sphincter deficiency in adults with spina bifida. These results suggest a possible pathophysiological role of extracellular matrix remodeling and both a fibrosis and a neurogenic component in the intrinsic sphincter deficiency of adults with spina bifida.