Although clinical trials have demonstrated an association between intradetrusor BoNT/A treatments and urinary tract infections (UTIs) in idiopathic overactive bladder (OAB) patients, observational studies in neurogenic incontinence patients suggested a decrease in UTIs, while randomized controlled trials support a lack of effect of BoNT/A on UTIs. Chronic inflammation is a common histological finding in both neurogenic and non-neurogenic overactive human bladders, but BoNT/A was not found to affect human bladder histological inflammation or edema. By contrast, in animal models intravesical BoNT/A reduced inflammatory markers such as cyclooxygenase-2 (COX-2), the EP4 receptor and Prostaglandin E2 (PGE2). Additionally, in OAB patients a decrease of blood levels of PGE2 was found following intradetrusor BoNT/A injections.  
As associations between UTIs, chronic inflammation and bladder overactivity are acknowledged, but not yet elucidated, we conducted a study to explore whether intradetrusor BoNT/A injections affect the background for increased incidence of UTIs. As Toll-like receptors (TLRs) can detect pathogens in the urinary tract, we studied the expression of TLR2, TLR4 and TLR5 in the urine of patients before and after intradetrusor BoNT/A injections. We also explored the expression of inflammatory cytokines IL-1β, IL-6, TNFα, as well as PGE2, which play a significant role as intermediates of immune response and as promoters of inflammatory reactions, in the urine of patients before and after an intradetrusor BoNT/A injection.

This pilot, prospective study was approved by the Hospital Scientific Board (ref. No: 89/22.01.2021) and all participants were recruited following written informed consent. Patients with neurogenic detrusor overactivity (NDO) associated incontinence refractory to oral pharmacotherapy who received bladder BonT/A injections were recruited. As per routine protocol approved by the Hospital Scientific Board all patients were submitted to urodynamic investigation at baseline, 4-6 weeks and 6 months post BoNT/A treatment. Urine specimens were obtained at baseline during the cystoscopy for the BoNT/A treatment, and upon routine urodynamic follow-up visits at 4-6 weeks and at 6 months post treatment. Specimens were preserved using the Urine Preservative Single Dose® by Norgen Biotek Corporation, Canada, and stored at room temperature. The expression of the genes of interest in the urine was studied by RNA isolation from urine samples, reverse transcription and Real-Time PCR (qRT-PCR).  GAPDH was used as house-keeping gene. 
The 2−∆∆CT algorithm was used to analyze the relative changes in gene expression.

Eighteen patients were recruited, who had at least one bladder BoNT/A treatment. They all had urine specimens adequate for analysis at one month post treatment, while specimens from ten patients were evaluable for processing at 6 months post BoNT/A. Four patients had a 2nd BoNT/A treatment and had specimens obtained upon the 2nd injection (clinical relapse time) and one-month post-treatment. All genes, but for TNFα, showed progressive downregulation at 1 and 6 months, which was more significant at 6 months post treatment (Figure 1A,B). TNFα appeared to significantly increase at 1 month after treatment, followed by a dramatic reduction at 6 months. A similar trend was found for TNFα after the 2nd BoNT/A injection.  Interestingly, the expression of all genes, apart from IL-1β, before the 2nd injection, remained significantly lower than before the 1st injection, showing no tendency for ‘relapse’ at the time of clinical relapse (Figure 1A,B). The latter was established by urodynamic investigation at all cases.

Our results suggest that intradetrusor BoNT/A injections may positively and significantly affect the background associated with the incidence of UTIs. Significant post-BoNT/A reductions in the expression of Toll-like receptors TLR2, 4 and 5, which can detect pathogens in the urinary tract, may indicate a reduced need for those receptors due to reduced incidence of UTIs. Also, the reduction in the urine expression of inflammatory cytokines IL-1β, IL-6 and of prostaglandin PGE2 which act as intermediates of immune response as well as promoters of inflammatory reactions, suggest a less inflammatory background in the patients’ bladders but may also indicate a reduced immune response, a.k.a. reduced incidence of UTIs. Results need to be evaluated in association with clinical data related to symptomatic and asymptomatic bacteriuria before and after the BoNT/A treatments, as well as the patients’ clinical response to BoNT/A.

In our cohort of patients with NDPO associated incontinence, intradetrusor BoNT/A injections achieved significant downregulation of the expression of Toll-like receptors associated with the detection of urinary tract pathogens, as well as of inflammatory cytokines and prostaglandin PGE2 associated with the mediation of immune response and inflammation, in the urine of patients. Our findings suggest that intradetrusor BoNT/A injections may affect the background associated with the incidence of UTIs in neurogenic bladders. Depending on the clinical significance of these results, the studied genes may be further explored as potential biomarkers for inflammation and UTIs in the neurogenic population.

Continence 12S (2024) 101432
DOI: 10.1016/j.cont.2024.101432