Vitreous Cryopreservation of Human Umbilical Vein Endothelial Cells with Low Concentration of Cryoprotective Agents for Vascular Tissue Engineering.

Cryopreservation of human umbilical vein endothelial cells (HUVECs) is important to tissue engineering applications and the study of the role of endothelial cells in cardiovascular and cerebrovascular diseases. The traditional methods for cryopreservation by vitrification (cooling samples to a cryogenic temperature without apparent freezing) using high concentration of cryoprotective agents (CPAs) and slow freezing are suboptimal due to the severe toxicity of high concentration of CPAs and ice formation-induced cryoinjuries, respectively. In this study, we developed a method to cryopreserve HUVECs by vitrification with low concentration of CPAs. This is achieved by optimizing the CPAs and using highly thermally conductive quartz capillary (QC) to contain samples for vitrification. The latter minimizes the thermal mass to create ultra-fast cooling/warming rates. Our data demonstrate that HUVECs can be vitrified in the QC using 1.4 mol/L ethylene glycol and 1.1 mol/L dimethyl sulfoxide with more than 90% viability. Moreover, this method significantly improves the attachment efficiency of the cryopreserved HUVECs. The attached cells post-cryopreservation proliferate similarly to fresh cells. Therefore, this study may provide an effective vitrification technique to bank HUVECs for vascular tissue engineering and other applications.

[1]  M. Gong,et al.  Superparamagnetic core/shell GoldMag nanoparticles: size-, concentration- and time-dependent cellular nanotoxicity on human umbilical vein endothelial cells and the suitable conditions for magnetic resonance imaging , 2015, Journal of Nanobiotechnology.

[2]  L. Bonewald,et al.  Prostaglandin E2 promotes proliferation of skeletal muscle myoblasts via EP4 receptor activation , 2015, Cell cycle.

[3]  Gang Zhao,et al.  Nanoparticle-mediated intracellular delivery enables cryopreservation of human adipose-derived stem cells using trehalose as the sole cryoprotectant. , 2015, ACS applied materials & interfaces.

[4]  G. Church,et al.  Multi-kilobase homozygous targeted gene replacement in human induced pluripotent stem cells , 2014, Nucleic acids research.

[5]  Y. Sotomaru,et al.  A study on cryoprotectant solution suitable for vitrification of rat two-cell stage embryos. , 2014, Cryobiology.

[6]  Ketan Kotwal,et al.  Results and Discussions , 2013 .

[7]  Hiroshi Mizumoto,et al.  An approach for formation of vascularized liver tissue by endothelial cell-covered hepatocyte spheroid integration. , 2009, Tissue engineering. Part A.

[8]  M. Toner,et al.  Vitrification by ultra-fast cooling at a low concentration of cryoprotectants in a quartz micro-capillary: a study using murine embryonic stem cells. , 2008, Cryobiology.

[9]  M. Toner,et al.  Thermal performance of quartz capillaries for vitrification. , 2007, Cryobiology.

[10]  P. Mazur,et al.  Kinetics of water loss and the likelihood of intracellular freezing in mouse ova , 1984, Cell Biophysics.

[11]  M. Kuwayama,et al.  Highly efficient vitrification for cryopreservation of human oocytes and embryos: the Cryotop method. , 2007, Theriogenology.

[12]  Michel Vaubourdolle,et al.  A protocol for isolation and culture of human umbilical vein endothelial cells , 2007, Nature Protocols.

[13]  V. Jacobs,et al.  Identification and reduction of cryoinjury in endothelial cells: a first step toward establishing a cell bank for vascular tissue engineering. , 2006, Tissue engineering.

[14]  Jeroen Rouwkema,et al.  Endothelial cells assemble into a 3-dimensional prevascular network in a bone tissue engineering construct. , 2006, Tissue engineering.

[15]  I. Katkov,et al.  Low- and high-temperature vitrification as a new approach to biostabilization of reproductive and progenitor cells , 2006 .

[16]  R. Thorne,et al.  Effects of cryoprotectant concentration and cooling rate on vitrification of aqueous solutions , 2006, physics/0604109.

[17]  Mehmet Toner,et al.  Cryo‐Injury and Biopreservation , 2005, Annals of the New York Academy of Sciences.

[18]  G. Vajta,et al.  Highly efficient vitrification method for cryopreservation of human oocytes. , 2005, Reproductive biomedicine online.

[19]  Daniel L. Riddle,et al.  Volume of Ambulatory Care Visits and Patterns of Care for Patients Diagnosed with Plantar Fasciitis: A National Study of Medical Doctors , 2004, Foot & ankle international.

[20]  Dai Fukumura,et al.  Tissue engineering: Creation of long-lasting blood vessels , 2004, Nature.

[21]  A. Dinnyés,et al.  Cryopreservation of goat oocytes and in vivo derived 2- to 4-cell embryos using the cryoloop (CLV) and solid-surface vitrification (SSV) methods. , 2003, Theriogenology.

[22]  Y. S. Yang,et al.  Vitrification of mouse oocytes using closed pulled straws (CPS) achieves a high survival and preserves good patterns of meiotic spindles, compared with conventional straws, open pulled straws (OPS) and grids. , 2001, Human reproduction.

[23]  S. Nakamura,et al.  Successful birth after transfer of vitrified human blastocysts with use of a cryoloop containerless technique. , 2001, Fertility and sterility.

[24]  G. Vajta,et al.  Full-term development of nuclear transfer calves produced from open-pulled straw (OPS) vitrified cytoplasts: work in progress. , 1999, Theriogenology.

[25]  M. Kasai,et al.  Vitrification of human embryos based on the assessment of suitable conditions for 8-cell mouse embryos. , 1998, Human reproduction.

[26]  F. Dignat-George,et al.  Immunologic phenotype of cultured endothelial cells: quantitative analysis of cell surface molecules. , 1997, Tissue antigens.

[27]  S. Leibo,et al.  Effect of chilling bovine oocytes on their developmental competence , 1996, Molecular reproduction and development.

[28]  A. Karow,et al.  Cryopreservation of cells for tissue engineering. , 1996, Tissue engineering.

[29]  P. Solli,et al.  Pulmonary Endothelial Cell Modifications after Storage in Solid-Organ Preservation Solutions , 1995, The Journal of international medical research.

[30]  D. Wiedbrauk,et al.  CRYOPRESERVATION OF CELLS , 1992 .

[31]  G. Fahy,et al.  Some emerging principles underlying the physical properties, biological actions, and utility of vitrification solutions. , 1987, Cryobiology.

[32]  P. Mazur.  Freezing of living cells: mechanisms and implications. , 1984, The American journal of physiology.

[33]  G M Fahy,et al.  Vitrification as an approach to cryopreservation. , 1984, Cryobiology.