PD27-01 DEFINING SYNAPTOTAGMIN-1 AS THE RECEPTOR FOR BOTULINUM NEUROTOXIN B IN THE BLADDER

Abstract

You have accessJournal of UrologyUrodynamics/Lower Urinary Tract Dysfunction/Female Pelvic Medicine: Basic Research & Pathophysiology (PD27)1 Sep 2021PD27-01 DEFINING SYNAPTOTAGMIN-1 AS THE RECEPTOR FOR BOTULINUM NEUROTOXIN B IN THE BLADDER Hatim Thaker, Jie Zhang, Shin-Ichiro Miyashita, Vivian Cristofaro, Maryrose Sullivan, Rosalyn Adam, and Min Dong Hatim ThakerHatim Thaker More articles by this author , Jie ZhangJie Zhang More articles by this author , Shin-Ichiro MiyashitaShin-Ichiro Miyashita More articles by this author , Vivian CristofaroVivian Cristofaro More articles by this author , Maryrose SullivanMaryrose Sullivan More articles by this author , Rosalyn AdamRosalyn Adam More articles by this author , and Min DongMin Dong More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002020.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Neurogenic and idiopathic detrusor overactivity (DO) is currently treated with onabotulinumtoxin A (BoNT/A) as third line therapy. In cases of treatment failure, alternatives to BoNT/A are limited. BoNT/B is another serotype that could serve as an alternative to BoNT/A. BoNT/B shares the same mode of action with BoNT/A, but uses different receptors. Two homologous proteins, synaptotagmin-1 and -2 (Syt-1, -2), have been identified as BoNT/B receptors. However, the dominant receptor form in bladder tissues remains unknown. Here, we aim to determine the specific Syt responsible for BoNT/B action in skeletal and smooth muscle, allowing us to engineer and improve BoNT/B efficacy in recognizing its specific receptor in the bladder. METHODS: Transgenic knock-in (KI) mice were generated by CRISPR/Cas9 approach, where a triple-residue mutation on Syt-1 and Syt-2 receptor abolishes BoNT/B binding. Wild type (WT) and KI mouse bladders were harvested, incubated with BoNT/B, and subjected to immunostaining with confocal microscopy or ex vivo nerve-evoked contraction studies via electrical field stimulation. To evaluate smooth muscle response in vivo, intramural bladder injection of BoNT/B (3.25 pg in 5 uL) via laparotomy was done. Post-op, mice underwent voiding spot assay (VSA) to assess toxin action on bladder function. RESULTS: WT bladders demonstrated high immunoreactivity to Syt-1 (Panel A), and Syt-1 KI bladders failed to bind BoNT/B in vitro (Panel B). Diaphragm skeletal muscle tissues showed a Syt-2 predominance. Ex vivo contraction studies with 0.3 nM BoNT/B showed a 90% reduction in nerve-evoked contractions in WT and Syt-2 KI, but not Syt-1 KI, bladders (Panel D). VSA after bladder injection of BoNT/B reduced voided volumes, akin to retention, in WT and Syt-2 KI mice only (Panel E). In contrast, BoNT/B produced full paralysis in hindlimb skeletal muscles in WT and Syt-1 KI mice; Syt-2 KI mice tolerated BoNT/B at 30x higher doses without paralysis (Panel C). CONCLUSIONS: We demonstrate that Syt-1 is the physiologically relevant receptor for BoNT/B in bladder tissues. The differential expression of Syt-1 vs Syt-2 provides an opportunity to engineer BoNT/B to improve its efficacy on bladder tissues and minimize its effect on skeletal muscles. Source of Funding: AUA/Urology Care Foundation Research Scholar Award and SUFU Chemodenervation Grant to H.T.R01NS080833, R01AI132387, R01AI139087, and R21NS106159 to M.D © 2021 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 206Issue Supplement 3September 2021Page: e443-e443 Advertisement Copyright & Permissions© 2021 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hatim Thaker More articles by this author Jie Zhang More articles by this author Shin-Ichiro Miyashita More articles by this author Vivian Cristofaro More articles by this author Maryrose Sullivan More articles by this author Rosalyn Adam More articles by this author Min Dong More articles by this author Expand All Advertisement Loading ...