Experimental bladder regeneration using a poly-l-lactide/silk fibroin scaffold seeded with nanoparticle-labeled allogenic bone marrow stromal cells

Abstract

In the present study, a poly-l-lactide/silk fibroin (PL-SF) bilayer scaffold seeded with allogenic bone marrow stromal cells (BMSCs) was investigated as a potential approach for bladder tissue engineering in a model of partial bladder wall cystectomy in rabbits. The inner porous layer of the scaffold produced from silk fibroin was designed to promote cell proliferation and the outer layer produced from poly-l-lactic acid to serve as a waterproof barrier. To compare the feasibility and efficacy of BMSC application in the reconstruction of bladder defects, 12 adult male rabbits were divided into experimental and control groups (six animals each) that received a scaffold seeded with BMSCs or an acellular one, respectively. For BMSC tracking in the graft in in vivo studies using magnetic resonance imaging, cells were labeled with superparamagnetic iron oxide nanoparticles. In vitro studies demonstrated high intracellular incorporation of nanoparticles and the absence of a toxic influence on BMSC viability and proliferation. Following implantation of the graft with BMSCs into the bladder, we observed integration of the scaffold with surrounding bladder tissues (as detected by magnetic resonance imaging). During the follow-up period of 12 weeks, labeled BMSCs resided in the implanted scaffold. The functional activity of the reconstructed bladder was confirmed by electromyography. Subsequent histological assay demonstrated enhanced biointegrative properties of the PL-SF scaffold with cells in comparison to the control graft, as related to complete regeneration of the smooth muscle and urothelium tissues in the implant. Confocal microscopy studies confirmed the presence of the superparamagnetic iron oxide nanoparticle-labeled BMSCs in newly formed bladder layers, thus indicating the role of stem cells in bladder regeneration. The results of this study demonstrate that application of a PL-SF scaffold seeded with allogenic BMSCs can enhance biointegration of the graft in vivo and support bladder tissue regeneration and function.

Keywords:

• bladder
• bone marrow stromal cells
• scaffold
• superparamagnetic iron oxide nanoparticles
• tissue engineering
• stem cells

Supplementary materials

Figure S1 Assessment of SPIONs’ cytotoxicity on BMSCs.

Notes: Following co-incubation of magnetic nanoparticles with BMSCs (at concentration 150 μg/mL) for 1, 3, 12, and 24 h the toxicity was assessed using MTT assay. Values are expressed as mean ± SD.

Abbreviations: BMSCs, bone marrow stromal cells; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; SD, standard deviation; SPIONs, superparamagnetic iron oxide nanoparticles; h, hours.

Figure S2 Lymphocytic infiltration at the implantation site of the PL-SF acellular scaffold. Notes: Infiltration is pointed by black solid arrows. Scale bar, 500 μm.

Abbreviation: PL-SF, poly-l-lactide/silk fibroin.

Acknowledgments

This study was supported by a grant from the Russian Science Foundation (No 14-50-00068) and with financial support from the Federal Agency of Scientific Organizations (Russia). The authors are grateful to Marina Parr, Liudmila Yakovleva, Yaroslav Marchenko, and Boris Nikolaev for synthesis of the magnetic nanoparticles, Olga Genbach for assistance in animal experiments and Yuliya Idrisova for analysis of scaffold properties.

Disclosure

The authors report no conflicts of interest in this work.