Abstract
Aim: Endothelial injury results in vascular disorders. Culture of endothelial cells (ECs) is important to investigate the pathogenesis of vascular diseases. ECs of rat aorta are widely applied in these researches. However, there are many problems to isolate them, which make it hard to culture primary ECs. In the present study, we established a simple and convenient method to isolate and culture rat aorta ECs. Methods: Two 5ml centrifuge tubes were prepared. An opening (3 mm × 25 mm) was made in one tube wall while the other was cut in half longitudinally. After the rat aorta was dissected along one side and attached to the wall of the half tube, the tube with an opening and the half tube with aorta were mated and secured by a rotund metal clip. Then, ECs were digested via the opening by 0.2% collagenase I at 37℃ for 1 h and identified by immunofluorescence of von Willebrand factor (vWF) and smooth muscle actin (SMA). Results: A simple method for isolating ECs were established. Cultured ECs exhibited typical “cobblestone” morphology at confluence. vWF was positively expressed in the ECs, and neither the negative stain of vWF nor the positive stain of SMA was found. Conclusion: This simple and convenient method to isolate and culture ECs of rat aorta can be implemented with readily available 5ml centrifuge tubes and will greatly benefit the research of vascular diseases.
Highlights
Vascular endothelial cells (ECs) cover the internal surface of the vessel and comprise the largest homogenous surface of the body [1]
From (Figure 4), we found that all of the smooth muscle cells (SMCs) presented the positive stain of smooth muscle actin (SMA); there was no positive stain of SMA in ECs
The rat aorta has lot of small branches, and the enzyme would leak out of the lumen if the aorta was directly perfused by digestive solution as human umbilical vein
Summary
Vascular endothelial cells (ECs) cover the internal surface of the vessel and comprise the largest homogenous surface of the body [1]. They play important roles in homeostatic mechanisms, such as regulation of vascular tone, maintenance of a non-thrombotic environment, and mediating the immune defense [2]. Human umbilical vein endothelial cell (HUVEC) is the most commonly used to investigate vascular disorders because it is easy to obtain and culture [7]. For cultured by explants of endothelium, ECs are easy to be contaminated by SMCs even if the explants were constantly removed after ECs grew from them This is time-consuming and inconvenient [11]. The procedure does not require special equipment such as magnetic beads or FACS, and a large number of ECs can be obtained in a short span of time by means of the method
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