Longitudinal intravital imaging of transplanted mesenchymal stem cells elucidates their functional integration and therapeutic potency in an animal model of interstitial cystitis/bladder pain syndrome

Abstract

Rationale: Mesenchymal stem cell (MSC) therapy may be a novel approach to improve interstitial cystitis/bladder pain syndrome (IC/BPS), an intractable disease characterized by severe pelvic pain and urinary frequency. Unfortunately, the properties of transplanted stem cells have not been directly analyzed in vivo, which hampers elucidation of the therapeutic mechanisms of these cells and optimization of transplantation protocols. Here, we monitored the behaviors of multipotent stem cells (M-MSCs) derived from human embryonic stem cells (hESCs) in real time using a novel combination of in vivo confocal endoscopic and microscopic imaging and demonstrated their improved therapeutic potency in a chronic IC/BPS animal model.Methods: Ten-week-old female Sprague-Dawley rats were instilled with 10 mg of protamine sulfate followed by 750 μg of lipopolysaccharide weekly for 5 weeks. The sham group was instilled with phosphate-buffered saline (PBS). Thereafter, the indicated dose (0.1, 0.25, 0.5, and 1×106 cells) of M-MSCs or PBS was injected once into the outer layer of the bladder. The distribution, perivascular integration, and therapeutic effects of M-MSCs were monitored by in vivo endoscopic and confocal microscopic imaging, awake cystometry, and histological and gene expression analyses.Results: A novel combination of longitudinal intravital confocal fluorescence imaging and microcystoscopy in living animals, together with immunofluorescence analysis of bladder tissues, demonstrated that transplanted M-MSCs engrafted following differentiation into multiple cell types and gradually integrated into a perivascular-like structure until 30 days after transplantation. The beneficial effects of transplanted M-MSCs on bladder voiding function and the pathological characteristics of the bladder were efficient and long-lasting due to the stable engraftment of these cells.Conclusion: This longitudinal bioimaging study of transplanted hESC-derived M-MSCs in living animals reveals their long-term functional integration, which underlies the improved therapeutic effects of these cells on IC/BPS.

Synergistic Effects of N-Acetylcysteine and Mesenchymal Stem Cell in a Lipopolysaccharide-Induced Interstitial Cystitis Rat Model

“…Bladder function was evaluated two weeks after the final instillation of LPS/PS under unrestrained and awake state in metabolic cages, as previously described [5][6][7]15].…”

“…Immediately after the awake cystometry, the bladder tissue was harvested for histological analysis, which evaluated the epithelial denudation, mast cell infiltration, tissue fibrosis, and apoptosis with cytokeratin immunostaining (Keratin, Pan Ab-1; Thermo Scientific, Foster City, CA, USA), toluidine blue staining (Toluidine blue-O; Daejung Chemicals and Metals, Seoul, Korea), Masson's trichrome staining (Junsei Chemical, Tokyo, Japan), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining (Roche, Mannheim, Germany), respectively [6,7,16]. Engraftment of the administered M-MSCs was determined by immunofluorescence analysis (EVOS, FL Color Imaging System, Life Technologies, Carlsbad, CA, USA) of human β2-microglobulin (hB2M) (mouse monoclonal, SC80668; Santa Cruz Biotechnology Inc., Paso Robles, CA, USA) and visualized by Alexa 488-conjugated (A11029) anti-mouse antibody (Molecular Probes, Grand Island, NY, USA), as previously described [2,9].…”

“…Stem cell therapy is considered a potential alternative treatment for chronic, degenerative disease which has limited therapeutic options with suboptimal efficacy [1][2][3]. Interstitial cystitis/bladder pain syndrome (IC/BPS), a symptomatic diagnosis characterized by chronic pelvic pain that lasts for more than six weeks and frequently accompanies lower urinary tract symptoms such as frequency and urgency in the absence of infection and other pathologies, is an ideal candidate for stem cell therapy [4][5][6][7]. Various treatment options, including behavioral therapy, oral pharmacotherapy, intravesical instillation, and surgical therapy, have been suggested for the treatment of IC/BPS, although none of them provide a definite cure for the disease.…”

“…In this regard, a treatment with mesenchymal stem cells (MSCs) is currently considered a new potential direction for treatment of IC/BPS in several preclinical settings [4][5][6][7][8]. The potential effects of MSCs in IC/BPS are related to their ability to produce a subset of biologically active substances with anti-inflammatory, anti-fibrosis, and tissue regenerating properties.…”

“…To overcome these issues, embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) have recently been employed as a promising alternative supply in order to provide sufficient cell numbers with enhanced in vivo survival, engraftment, and functionality for sustained therapeutic potency [11,12]. We recently proved the superior therapeutic effects and engraftment as well as long-term survival of human ESC-derived multipotent MSCs (M-MSCs) compared to their adult tissue counterpart, having employed several IC/BPS animal models that recapitulate the different pathogenesis by intravesical instillation of hydrochloride acid (HCl) or lipopolysaccharide/protamine sulfate (LPS/PS) as well as intravenous injection of ketamine [5][6][7].…”