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Characterization of rabbit urine-derived stem cells for potential application in lower urinary tract tissue regeneration

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Abstract

Tissue-engineered urethra with autologous cells seeded on biodegradable scaffolds offers an alternative for lower urinary tract reconstruction. Rabbit is most commonly used as an animal model in urethra and bladder tissue repair. The goal of this study is to characterize rabbit urine-derived stem cells (rUSC) and induce these cells to differentiate into urothelial and smooth muscle cells as an autologous cell source for potential use in lower urinary tract tissue regeneration in a rabbit model. We successfully cultured rUSC from 12 urine samples and 13 bladder wash samples of six rabbits. rUSC colonies appeared more in the bladder wash solution (2–4/15 ml) than those in the urine samples (1–2 clones/15 ml urine). The cells displayed rice grain-like in morphology. Mean population doubling of rUSC was 48.5 ± 6.2 and average doubling time was 25.7 ± 8.4 h, indicating that a single of rUSC clone generated about 4 × 1014 cells in 50 days. The rUSC were positive for CD29, CD90 and CD105 but negative for CD31, CD34 and CD45 in flow cytometry. When exposed to PDGF-BB and TGF-β1, these cells could differentiate into spindle-like cells, expressing smooth muscle-specific proteins, including α-smooth muscle action, desmin and myosin. Urothelially differentiated rUSC expressed urothelial-specific proteins, i.e., AE1/AE3 and E-cadherin when exposed to epidermal growth factor (EGF). Osteogenic-differentiated rUSC expressed osteogenic marker, i.e., alkaline phosphatase when exposed to serum containing DMEM low-glucose medium with osteogenic supplements. In conclusion, rUSC can be isolated from bladder wash or urine samples and cultured in vitro. There stem cells possess strong proliferative ability and are capable of differentiating in urothelial, myogenic and osteogenic lineages. Thus, rUSC are a potential alternative autologous cell source for lower urinary tract repair with tissue engineering technology in a rabbit model.

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Acknowledgements

The authors acknowledge funding support from the National Natural Science Foundation of China (No. 81570650 and No. 81371704).

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Author notes

  1. Huai Yang and Bote Chen contributed equally to this work.

Authors and Affiliations

  1. Department of Urology, General Hospital of Guangzhou Military Command of PLA, Guangzhou, Guangdong, China

    Huai Yang, Bote Chen, Guiwu Zhuang & Shijian Wu

  2. Department of Urology, Guangdong Second Provincial General Hospital, Guangzhou, Guangdong, China

    Bote Chen, Guosheng Yang & Xiaofu Qiu

  3. Department of Andrology, Guangzhou First People’s Hospital, Guangzhou, Guangdong, China

    Junhong Deng

  4. Reproductive Medicine Research Center, the Sixth Affiliated Hospital of Sun Yat-sen University, Guangzhou, China

    Guihua Liu

  5. Department of Urology, First Affiliated Hospital of Sun Yat-Sen University, Guangzhou, Guangdong, China

    Chunhua Deng

  6. School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, China

    Pinqing Wei

  7. Department of Urology, Shenzhen Longhua District Central hospital, Shenzhen, Guangdong, China

    Xisheng Wang

  8. Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA

    Yuanyuan Zhang

Authors
  1. Huai Yang
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  2. Bote Chen
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  3. Junhong Deng
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  4. Guiwu Zhuang
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  6. Guihua Liu
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  7. Chunhua Deng
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  11. Xisheng Wang
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  12. Yuanyuan Zhang
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Contributions

Administrative support: HY.

Editorial help: DC, DJ and YYZ.

Conceived and designed the experiments: YYZ.

Performed the experiments: BC, GZ and LG.

Analyzed the data: CD, DJ, SW, PW, XW and YYZ.

Contributed reagents/materials/analysis tools: HY.

Writing the paper: BC, GL and YYZ.

Corresponding authors

Correspondence to Huai Yang or Yuanyuan Zhang.

Ethics declarations

The protocol for collection of human urine samples from healthy donors was approved by the Wake General Hospital of Guangzhou Military Area (GHGMA) Institutional Review Board. The study protocol conforms to the ethical guidelines of the Declaration of Helsinki. Written informed consent was obtained from the urine donors. Experiments in rabbit were approved by the Animal Care and Use Committee at GHGMA. All the animal experiments were conducted in accordance with guidelines of the international committee of use of laboratory animals.

Competing interest

The authors declare that they have no competing interests.

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Yang, H., Chen, B., Deng, J. et al. Characterization of rabbit urine-derived stem cells for potential application in lower urinary tract tissue regeneration. Cell Tissue Res 374, 303–315 (2018). https://doi.org/10.1007/s00441-018-2885-z

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  • DOI: https://doi.org/10.1007/s00441-018-2885-z

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