Long‐term cryopreservation of pyramidalis muscle specimens as a source of striated muscle stem cells for treatment of post‐prostatectomy stress urinary incontinence

Abstract

Stem-cell injection into the degenerated external urethral sphincter is a new treatment modality for stress urinary incontinence (SUI). We examined the possibility of long-term cryopreserved pyramidalis muscle (PM) specimens as a source of striated muscle stem cells for the treatment of post-prostatectomy SUI. PM specimens were obtained from five male patients (mean age, 61-70 years) who underwent radical prostatectomy for prostate cancer. Specimens (volume, approximately 125 mm³ ) were obtained from the incisional edge, minced, and stored at -80°C in a freezing medium (Cell Banker 1®). After 24-60 months, the specimens were thawed and directly incubated at 37°C. Satellite cells were selectively cultured by magnetic affinity cell sorting using an anti-neural cell adhesion molecule (NCAM) antibody. Osteogenic and adipogenic differentiation were induced by bone morphogenic protein-7 (BMP-7) and γ-linolenic acid, respectively. NCAM-positive cells (>99% purity) were selectively cultured from all cryopreserved PM specimens and confirmed as being of striated muscle origin by the expression of desmin and MyoD. They fused and differentiated into multinucleated myotubes 7 days after incubation in a differentiation induction medium. Stimulation by BMP-7 and γ-linolenic acid induced expression of alkaline phosphatase (osteoblast marker) and lipid deposition within the cytoplasm (adipocyte characteristic), respectively. Long-term cryopreserved PM specimens can be used to culture muscle stem cells. Therefore, this method may be utilized for SUI treatment when necessary. Moreover, complete remove of the prostate gland without fear of injury to the urethral sphincter may be possible in patients with apical cancer or T3 prostate cancer.