Neurogenic lower urinary tract dysfunction (NLUTD) is common in patients with different neurological diseases. Neurological conditions that affect the suprasacral segments of the spinal cord are often accompanied by detrusor overactivity and detrusor sphincter dyssynergia (DSD). The resulting high detrusor pressures that are seen in these patients are a well-known risk factor for bladder deterioration and upper urinary tract damage. In the long term, patients with such lesions may progress with bladder fibrosis and decreased bladder compliance. Evidence from studies with animal models of bladder outlet obstruction (BOO) and also from bladder samples from men with BOO due to benign prostatic hyperplasia indicate that hypoxia plays a significant role in BOO-induced bladder dysfunction. As the bladder intramural tension increases, there is decreased blood flow and consequent detrusor hypoxia. Chronic bladder ischemia may result in impaired detrusor contractility and decreased bladder compliance. The cellular adaptation that occurs as a response to hypoxic events is primarily mediated by the hypoxia-inducible factors (HIF’s). The HIF-related pathway is responsible for the activation of other key genes in the remodeling and adaptation process to hypoxia, such as vascular endothelial growth factor (VEGF), nitric oxide synthase (NOS) and connective tissue growth factor (CTGF). In the present study we evaluated the expression of genes associated with hypoxia and fibrosis in the bladder of patients with NLUTD requiring bladder augmentation.

We analyzed bladder specimens from 17 patients with NLUTD caused by diseases affecting the suprasacral spinal cord who underwent bladder augmentation due to low bladder compliance or detrusor overactivity refractory to conservative treatment. All subjects provided written informed consent and the study was approved by the Ethics Committee. Preoperative urodynamics was performed in all patients. A full thickness bladder sample was collected during the bladder augmentation surgery. The bladder biopsies were prepared for relative gene expression analysis with quantitative real-time polymerase chain reaction (RT-PCR) of MMP-1, TIMP-1, HIF1α, HIF2α, HIF3α, iNOS, eNOS, VEGF and CTGF. The control group included bladder specimens obtained from nine cadaveric organ donors. Expression levels of the RNA were analyzed by qRT-PCR using an ABI 7500 Fast RT-PCR System (Applied Biosystems). Data were expressed as medians and interquartile ranges or absolute values and fractions. Confidence interval were calculated with natural logarithm transformation.

The group was composed of 13 men and 4 women and the mean age was 37 years ± 18.33 (range 11-60). The cause of NLUTD was spinal cord injury in 10 patients (58.82%), myelomeningocele in 4 patients (23.52%) and inflammatory diseases in 2 patients (11.76%).  The median duration of neurological disease was 12 years (range 3-24). Urodynamic findings demonstrated mean maximum cystometric capacity of 260 ± 63.50 mL, mean bladder compliance of 17.95 ± 16.80 mL/cmH20 and 7 (41.17%) patients had detrusor overactivity. Patients with NLUTD had statistically significant overexpression of TIMP-1, HIF1α and HIF3α and HIF2α showed a trend to overexpression. MMP-1 was underexpressed in patients with NLUTD and the expressions of VEGF, CTGF, eNOS, and iNOS was heterogeneous. Data are shown in Table 1.

We investigated the expression of genes related to hypoxia and fibrosis in the bladder of patients with NLUTD who underwent bladder augmentation due to low bladder compliance or refractory detrusor overactivity. This is the first study to evaluate the HIF-related pathway in the bladder of patients with NLUTD. We showed overexpression of HIF 1 and 3 and a trend to overxpression of HIF 2. These findings are consistent with animal studies showing a role for hypoxia in the mechanisms leading to bladder fibrosis and seem to indicate that it is also important in patients with NLUTD. In our series, however, we did not find overexpression of VEGF, CTGF and nitric oxide synthase, which are important genes in the remodeling and adaptation process to hypoxia. These findings may be due to the heterogeneity of our population and to the long time of disease duration compared to the other models studied until this date.

Our study is the first to investigate the HIF-related pathway in the pathophysiology of the bladder decompensation in patients with NLUTD. We demonstrated the overexpression of HIF1α, HIF 3α and TIMP-1 and underexpression of MMP-1 in the detrusor layer of patients with NLUTD undergoing bladder augmentation.

Drzewiecki BA, Anumanthan G, Penn HA, Tanaka ST, Thomas JC, Adams MC, Brock JW 3rd, Pope JC 4th, Matusik RJ, Hayward S, Clayton DB. Modulation of the hypoxic response following partial bladder outlet obstruction. J Urol. 2012 Oct;188(4 Suppl):1549-54. doi: 10.1016/j.juro.2012.02.037. Epub 2012 Aug 19. PubMed PMID: 22910264.
Stephany HA, Strand DW, Ching CB, Tanaka ST, Milne GL, Cajaiba MM, Thomas JC,  Pope JC 4th, Adams MC, Brock JW 3rd, Hayward SW, Matusik RJ, Clayton DB. Chronic  cyclic bladder over distention up-regulates hypoxia dependent pathways. J Urol. 2013 Oct;190(4 Suppl):1603-9. doi: 10.1016/j.juro.2013.02.026. Epub 2013 Feb 19.  PubMed PMID: 23429070; PubMed Central PMCID: PMC4085185.
Wiafe B, Adesida A, Churchill T, Adewuyi EE, Li Z, Metcalfe P. Hypoxia-increased expression of genes involved in inflammation, dedifferentiation, pro-fibrosis, and extracellular matrix remodeling of human bladder smooth muscle cells. In Vitro Cell Dev Biol Anim. 2017 Jan;53(1):58-66. doi: 10.1007/s11626-016-0085-2. Epub 2016 Sep 8. PubMed PMID: 27632054.