PROJECT SUMMARY Dysfunction of the striated external urethral sphincter is the strongest predictor of stress urinary incontinence, defined as involuntary loss of urine due to increased intraabdominal pressure in the absence of bladder contraction. Stress urinary incontinence affects approximately 1 in 2 women at some point in their lives. The primary inciting event behind the development of chronic stress urinary incontinence has been unequivocally identified as injury to the urinary continence mechanism sustained by women during vaginal deliveries. This astoundingly prevalent condition dramatically decreases quality of life, causes significant morbidity, and is associated with large economic burden to the individuals and society. Despite this, preventative strategies are almost non-existent, and the available treatments are delayed and compensatory as they do not directly target the underlying pathophysiology. The above is largely due to the fact that our understanding of the pathways that lead to failure of the intrinsic muscular components of the external urethral sphincter following birth injury remains limited. Furthermore, the prevailing preclinical studies do not utilize biologically relevant pregnant animal model of birth injury. To address the existing unmet clinical need and knowledge gaps, we assembled a cross- disciplinary team with diverse but complimentary expertise to execute the current project at the interface between basic science, biomaterial development, and translational medicine. We will use a validated and biologically relevant pregnant pre-clinical model to investigate structural, molecular, and cellular events at multiple time points across a recovery continuum of the striated external urethral sphincter following birth injury. These basic processes will inform the development of and the critical time to deliver new, minimally invasive tissue- engineered therapy for the prevention and treatment of urethral muscle dysfunction. Specifically, we will test a novel pro-regenerative skeletal muscle-specific injectable extracellular matrix hydrogel, derived from decellularized porcine skeletal muscles, in preventing and reversing maladaptive recovery of the external urethral sphincter following birth injury. Collectively, this innovative study will provide fundamental knowledge of the biological processes involved in the regulation of external urethral sphincter muscle regeneration, and comprehensive functionally relevant assessments of the role of low-cost acellular minimally invasive regenerative therapy to counteract the existing epidemic of urinary incontinence.