Efficient recovery of undifferentiated human embryonic stem cell cryopreserved with hydroxyethyl starch, dimethyl sulphoxide and serum replacement

Autor(es)

Orellana, Maristela Delgado ^{[1, 2]} ; De Santis, Gil Cunha ^[1] ; Abraham, Kuruvilla Joseph ^[3] ; Fontes, Aparecida Maria ^[4] ; Rosa Magalhaes, Danielle Aparecida ^[1] ; Oliveira, Viviane de Cassia ^[5] ; Oliveira Costa, Everton de Brito ^[1] ; Bonini Palma, Patricia Vianna ^[1] ; Covas, Dimas Tadeu ^{[1, 2]}

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Ribeirao Preto Med Sch, Hemotherapy Ctr Ribeirao Preto, BR-14051140 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Ribeirao Preto Med Sch, Dept Clin Med, BR-14051140 Ribeirao Preto, SP - Brazil

^[3] Univ Sao Paulo, Ribeirao Preto Med Sch, Dept Pediat, BR-14051140 Ribeirao Preto, SP - Brazil

^[4] Univ Sao Paulo, Ribeirao Preto Med Sch, Dept Genet, BR-14051140 Ribeirao Preto, SP - Brazil

^[5] Univ Sao Paulo, Sch Dent Ribeirao Preto, Dept Dent Mat & Prosthodont, BR-14051140 Ribeirao Preto, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: CRYOBIOLOGY; v. 71, n. 1, p. 151-160, AUG 2015.
• Citações Web of Science: 4

Resumo

Background: The therapeutic use of human embryonic stem cells (hESCs) is dependent on an efficient cryopreservation protocol for long-term storage. The aim of this study was to determine whether the combination of three cryoprotecting reagents using two freezing systems might improve hESC recovery rates with maintenance of hESC pluripotency properties for potential cell therapy application. Methods: Recovery rates of hESC colonies which were frozen in three cryoprotective solutions: Me2SO/HES/SR medium, Defined-medium (R) and Me2SO/SFB in medium solution were evaluated in ultra-slow programmable freezing system (USPF) and a slow-rate freezing system (SRF). The hESC pluripotency properties after freezing-thawing were evaluated. Results: We estimated the distribution frequency of survival colonies and observed that independent of the freezing system used (USPF or SRF) the best results were obtained with Me2SO/HES/SR as cryopreservation medium. We showed a significant hESC recovery colonies rate after thawing in Me2SO/HES/SR medium were 3.88 and 2.9 in USPF and SRF, respectively. The recovery colonies rate with Defined-medium (R) were 1.05 and 1.07 however in classical Me2SO medium were 0.5 and 0.86 in USPF and SRF, respectively. We showed significant difference between Me2SO/HES/SR medium x Defined-medium (R) and between Me2SO/HES/SR medium x Me2SO medium, for two cryopreservation systems (P < 0.05). Conclusion: We developed an in house protocol using the combination of Me2SO/HES/SR medium and ultra-slow programmable freezing system which resulted in hESC colonies that remain undifferentiated, maintain their in vitro and in vivo pluripotency properties and genetic stability. This approach may be suitable for cell therapy studies. (C) 2015 Elsevier Inc. All rights reserved. (AU)

• Processo FAPESP: 13/08135-2 - CTC - Centro de Terapia Celular
• Beneficiário: Dimas Tadeu Covas
• Modalidade de apoio: Auxílio à Pesquisa - Centros de Pesquisa, Inovação e Difusão - CEPIDs