June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
A simple method for the isolation, purification and cultivation of retinal microvascular pericytes from rat
Author Affiliations & Notes
  • Xiaoling Liu
    School of Optometry and Ophthalmology and Eye Hospital, Wenzhou Medical College, Wenzhou, China
    State Key Laboratory Cultivation Base and Key Laboratory of Vision Science,Ministry of Health P.R. China, Wenzhou, China
  • Guanghui Liu
    School of Optometry and Ophthalmology and Eye Hospital, Wenzhou Medical College, Wenzhou, China
    Department of Ophthalmology, Affiliated People's Hospital, Fujian University of Traditional Chinese Medicine, Fuzhou, China
  • Chun Meng
    Department of Bioengineering, College of Biological Science and Biotechnology, Fuzhou University, Fuzhou, China
  • Mingdong Pan
    Department of Ophthalmology, Affiliated People's Hospital, Fujian University of Traditional Chinese Medicine, Fuzhou, China
  • Yongzheng Zheng
    Department of Ophthalmology, Affiliated People's Hospital, Fujian University of Traditional Chinese Medicine, Fuzhou, China
  • Ling Lin
    Department of Bioengineering, College of Biological Science and Biotechnology, Fuzhou University, Fuzhou, China
  • Li Zhao
    Department of Cardiology, Affiliated People's Hospital, Fujian University of Traditional Chinese Medicine, Fuzhou, China
  • Footnotes
    Commercial Relationships Xiaoling Liu, None; Guanghui Liu, None; Chun Meng, None; Mingdong Pan, None; Yongzheng Zheng, None; Ling Lin, None; Li Zhao, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 5146. doi:https://doi.org/
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Xiaoling Liu, Guanghui Liu, Chun Meng, Mingdong Pan, Yongzheng Zheng, Ling Lin, Li Zhao; A simple method for the isolation, purification and cultivation of retinal microvascular pericytes from rat. Invest. Ophthalmol. Vis. Sci. 2013;54(15):5146. doi: https://doi.org/.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose: to establish a simple method for isolation, purification and cultivation of primary retinal microvascular pericytes(RMPs)from a readily available tissue source, rat, and to facilitate the study of their properties in vitro.

Methods: Retinas were isolated from weanling rats by ophthalmic microscopic surgical instruments followed by mechanical morcel and collagenase digestion. After homogeneous retinal microvascular fragments of 50-100μm were obtained from the digested retinas by filtered through nylon sieve, they were incubated in Dulbecco's modified Eagle's medium with 20% fetal b

Results: Migrating cells emerged from microvascular fragments after 24-48 hours of plating and formed loose colonies by 1 week. And they formed a pro-confluent multilayer on day 14-16. The subcultures grew faster and reached confluence on day 12-14. The cultured cells and their subpassages showed typical pericyte morphology with irregular triangular cell bodies and multiple long processes. They uniformly expressed the cellular markersα-smooth muscle actin (α-SMA), platelet-derived growth factor receptor-β(PDGFR-β), nerve/glial antigen 2 (NG2) and desmin, but lack of markers stained positive for von Willebrand’s factor(vWF), glutamine synthetase (GS) and glial fibrillary acidic protein(GFAP).The purity of cultured cells was up to 99% demonstrated by positive double staining for α-SMA and PDGFR-β. The cells can be cryopreserved, recultured and passaged repeatedly for 7 passages without a significant loss in expression of pericyte markers and for 9 passages without loss of typical features.

Conclusions: These results indicate that the isolated cells were RMPs showing both morphologic and functional characteristics of pericytes. Rat RMPs can be obtained easily by our method. Here we establish a simple and convenient method for the primary culture of rat RMPs, which would be more helpful to identify their roles the pathogenesis of various retinal angiogenic diseases such as diabetic retinopathy.

Keywords: 694 retinal culture • 691 retina: proximal (bipolar, amacrine, and ganglion cells) • 698 retinal development  
×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×