Inducible Loss Of Integrin beta 1 From Bladder Smooth Muscle Causes Increased Voiding Frequency And Impaired Muscarinic Contractility

Integrins are integral membrane proteins which, in dynamic ways, physically link, the actin cytoskeleton to the extracellular matrix. They exhibit broad functional diversity and are involved in cellular differentiation, adhesion, proliferation and mechanotransduction. Diabetic bladder dysfunction (DBD) is a very common progressive complication of diabetes mellitus resulting in overactivity during the early compensatory phase and ultimately in an atonic bladder which is unable to empty properly. Both genomic and proteomic approaches focused on bladder smooth muscle (BSM) have demonstrated altered expression of cell adhesion proteins and extracellular matrix as well as compromised contractility, as molecular markers of DBD. To begin to understand the role of cell adhesion and mechanosensitive signaling in BSM we created a mouse with tamoxifen (TMX) inducible, smooth muscle‐conditional deletion of β1‐integrin using Cre‐lox (Txβ1‐cKO). Loss of β1‐integrin mRNA and protein following TMX induction by i.p. injection (5 × 2 mg on consecutive days) were confirmed by PCR and Western blotting. Void‐spotting assays on filter paper revealed a progressive increase in number of voids from 2.5/4 h to 8.3/4 h over a period of 60 days following Cre‐induction (n=5 mice/group, P <0.05). Littermate controls injected with vehicle (no TMX) showed no change. The mean volume/void decreased from 264 μl to 158 μl over 60 days in the integrin knockouts. An increase in the number of voids with reduced volume/void is often taken to indicate some form of overactivity. Urodynamic measurements using cystometry under anesthesia, revealed significantly reduced peak bladder pressures during voiding (20 cmH 2 O vs. 26 cmH 2 O, P <0.05), reduced intercontraction interval (44 s vs. 249 s, P <0.01), and reduced compliance (17 s/cmH 2 O vs. 68 s/cmH 2 O, N.S.) in TMX injected vs. control mice. Thus, in vivo voiding frequency was dramatically increased at the same time as the bladders showed increased resistance to filling and lower expulsive pressures. Urothelium dissected muscle‐strip force measurements by myography revealed a significant loss of contractile force in integrin‐deficient bladders in response to electrical field stimulation (45% and 62% mean reductions at 25 and 60 days post Cre‐induction respectively, for stimulation frequencies of 1 – 50 Hz). Selective inhibition of the muscarinic force component by incubation of strips with 0.5 μM atropine, indicated that muscarinic contractility was severely inhibited in integrin‐deficient BSM (17.2% of force generation was due to muscarinic receptors compared to 50% in controls) while purinergic contractility was largely intact. Masson trichrome staining of paraffin embedded bladder sections revealed a substantial expansion of collagenous connective tissue in BSM and disruption to close packing of smooth muscle bundles. We conclude that β1‐integrin is necessary for BSM structural integrity and for muscarinic signaling during force generation. These findings begin to offer insights into the physiological role of integrin mechanical signaling in BSM and may be relevant to certain bladder pathologies in which integrin expression levels have been shown to be dysregulated. Support or Funding Information Support for this work was provided by NIH grants DK‐083299 and DK‐095922. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .