Culture of rat corpus cavernosal endothelial cells using modified immunomagnetic beads and cloning.
- Author:
Fan-Bo ZHANG
1
;
Rui JIANG
1
Author Information
1. Department of Urology, The Affiliated Hospital of Southwest Medical University, Luzhou, Sichuan 646000, China.
- Publication Type:Journal Article
- Keywords:
cloning;
corpus cavernosum;
endothelial cell;
magnetic bead;
rat
- MeSH:
Animals;
Cell Culture Techniques;
Cell Movement;
Cell Proliferation;
Cells, Cultured;
Endothelial Cells;
chemistry;
cytology;
physiology;
Erectile Dysfunction;
pathology;
Flow Cytometry;
Humans;
Immunomagnetic Separation;
Male;
Penis;
cytology;
Platelet Endothelial Cell Adhesion Molecule-1;
analysis;
Rats;
Rats, Sprague-Dawley;
Sincalide;
analysis;
von Willebrand Factor;
analysis
- From:
National Journal of Andrology
2017;23(6):503-509
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To search for the methods of isolating, purifying and culturing corpus cavernosal endothelial cells (CCECs) from SD rats, observe their growth characteristics, and providing seed cells for the study of erectile dysfunction (ED).
METHODS:The corpus cavernosal tissue from the SD rat was digested with 0.1% elastase, followed by purification of CCECs with immunomagnetic beads. After further amplification, monoclonal CCECs were sorted out with the cloning cylinder and their morphological and proliferative characteristics were observed. The von Willebrand factor (VWF) in the CCECs was identified by immunofluorescence staining, the CD31 molecule detected by immumohistochemistry, the purity of the CCECs determined by flow cytometry, and the proliferation of the cells measured with CCK-8 and growth curves.
RESULTS:After 7 days of purification and culture, the CCECs were fused into a monolayer under the inverted phase-contrast microscope, arranged like flagstones. The growth curves showed that the CCECs were in latency with a low growth rate at 1－2 days, in the logarithmic growth phase with a rapid rate at 3－4 days, and into the platform phase around the 6th day. VWF was positively expressed in the CCECs with much green fluorescence, and so was CD31 with a large number of brownish particles. The positive rate of the CCECs which were labelled with the VWF purified with magnetic beads combined with cloning cylinders was up to (91.9±3.75)%.
CONCLUSIONS:High-purity rat CCECs can be cultured successfully using immunomagnetic beads combined with cloning cylinders, with stable proliferation and passage in the endothelial cell medium.
