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Primary cultivation and identification of human placental microvascular endothelial cells / 中国组织工程研究
Chinese Journal of Tissue Engineering Research ; (53): 1706-1711, 2014.
Article in Chinese | WPRIM | ID: wpr-446482
ABSTRACT

BACKGROUND:

Establishment of in vitro culture system of human placental microvascular endothelial cel s with high purity is very important. In recent studies, some scholars have successful y obtained a large number of placental microvascular endothelial cel s by three-stepenzyme digestion and magnetic separation method, but the procedures were extremely complex and it had great damage to the cel s. Therefore, how to separate human placental microvascular endothelial cel s easily and obtain high-purified cel s has become a research hotspot.

OBJECTIVE:

To investigate an efficient method to isolate and purify human placental microvascular endothelial cel s from early vil us microvessels, observe the cel growth and identify the cel s.

METHODS:

The vil i from normal early pregnancies (6-8 weeks) after artificial abortion were col ected aseptical y. Using two-step digestion procedure and discontinuous Percol density gradient centrifugation method, human placental microvascular endothelial cel s were obtained. Then the cel s were identified by trypsin digestion method and repeated adherence method. RESULTS AND

CONCLUSION:

Human placental microvascular endothelial cel s were isolated successful y from early vil i. The primary cel s adhered to the wal s after inoculated for 24 hours and entered logarithmic phase at 10 days. 80%of the cel s achieved a confluence at 12-13 days after inoculating. The subculture cel s grew swiftly with the typical cobblestone appearance. Immunofluorescence staining showed that, cultured human placental microvascular endothelial cel s demonstrated a strong positive reaction to von Wil ebrand factor antigen and CD31, accounting for 100%. MTT assay results showed that, human placental microvascular endothelial cel s at passage 5 exhibited an S-shaped growth curve. High-purity human placental microvascular endothelial cel s can be obtained by proteolytic enzymes digestion and discontinuous Percol density gradient centrifugation method, and the purity is detected by trypsin digestion method and repeated adherence method.

Full text: Available Index: WPRIM (Western Pacific) Type of study: Diagnostic study / Prognostic study Language: Chinese Journal: Chinese Journal of Tissue Engineering Research Year: 2014 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Type of study: Diagnostic study / Prognostic study Language: Chinese Journal: Chinese Journal of Tissue Engineering Research Year: 2014 Type: Article