Improved Cryopreservation of Human Umbilical Vein Endothelial Cells: A Systematic Approach

• Published in: Scientific reports Vol. 6; no. 1; p. 34393
• Main Authors: Sultani, A. Billal, Marquez-Curtis, Leah A., Elliott, Janet A. W., McGann, Locksley E.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 06-10-2016; Nature Publishing Group
• Subjects: 13; 13/106; 13/31; 631/61/51; 639/766/747; Angiogenesis; Cell adhesion & migration; Cooling; Cryoinjury; Cryopreservation; Cryopreservation - methods; Cryoprotective Agents - pharmacology; Dimethyl sulfoxide; Dimethyl Sulfoxide - pharmacology; Drug delivery; Endothelial cells; Freezing; Human Umbilical Vein Endothelial Cells - cytology; Human Umbilical Vein Endothelial Cells - metabolism; Humanities and Social Sciences; Humans; Hydroxyethyl starch; multidisciplinary; Science; Starch; Time Factors; Tissue engineering; Umbilical vein
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep34393

Cryopreservation of human umbilical vein endothelial cells (HUVECs) facilitated their commercial availability for use in vascular biology, tissue engineering and drug delivery research; however, the key variables in HUVEC cryopreservation have not been comprehensively studied. HUVECs are typically cryopreserved by cooling at 1 °C/min in the presence of 10% dimethyl sulfoxide (DMSO). We applied interrupted slow cooling (graded freezing) and interrupted rapid cooling with a hold time (two-step freezing) to identify where in the cooling process cryoinjury to HUVECs occurs. We found that linear cooling at 1 °C/min resulted in higher membrane integrities than linear cooling at 0.2 °C/min or nonlinear two-step freezing. DMSO addition procedures and compositions were also investigated. By combining hydroxyethyl starch with DMSO, HUVEC viability after cryopreservation was improved compared to measured viabilities of commercially available cryopreserved HUVECs and viabilities for HUVEC cryopreservation studies reported in the literature. Furthermore, HUVECs cryopreserved using our improved procedure showed high tube forming capability in a post-thaw angiogenesis assay, a standard indicator of endothelial cell function. As well as presenting superior cryopreservation procedures for HUVECs, the methods developed here can serve as a model to optimize the cryopreservation of other cells.