title: Effect of cryopreservation on in vitro clonal growth of cordonal blood cells.
abstract: Cordonal blood (CB) is today recognized as a potentially important source of hematopoietic stem cells (SCs) for allogeneic transplantation, and to this task it would be of extreme importance to have the possibility of using cryopreserved CB units. Hence we investigated whether freezing and thawing alter the viability of CB hematopoietic cells. Mononuclear cells, recovered from fresh CB units by density-gradient centrifugation, were partly frozen and then thawed and partly immediately utilized for clonogenic tests. The cells were cultured in H4330 (C group) or H4434 (H4330 added with SC clonogenic growth factors) (C+ group) semisolid media added or not with serum-free Dulbecco modified Eagle medium (DMEM/sf). Cells seeded on H4330 were exposed to the conditioned supernatants from two human-embryo liver cell lines, which have been previously found to stimulate clonal growth of fresh CB hematopoietic cells. As expected, the number of colony-forming units (CFU) was higher in C+ than C group, and was not influenced by the addition of DMEM/sf. CFU number was higher in cryopreserved than fresh cells in both C and C+ culture groups. Conditioned supernatants from both cell lines stimulated clonal growth in both fresh and cryopreserved cell cultures. These findings indicate that cryopreservation and thawing do not alter the viability of CB SCs, but, on the contrary, improve their basal and cytokine-stimulated clonal growth, probably by negatively selecting SCs among the mononuclear CB cell population.
authors.affiliation: Department of Pharmaceutical Sciences (School of Pharmacy), University of Padua, I-35121 Padua, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Del Pup
authors.forename: Sergio
authors.initials: S
authors.lastname: De Angeli
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Conconi
authors.forename: Claudio
authors.initials: C
authors.lastname: Grandi
authors.forename: Pier Giorgio
authors.initials: PG
authors.lastname: Gamba
authors.forename: Pier Paolo
authors.initials: PP
authors.lastname: Parnigotto
authors.forename: Gastone G
authors.initials: GG
authors.lastname: Nussdorfer
mesh_terms: D002470:Cell Survival; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D002999:Clone Cells / Q000187:drug effects / Q000378:metabolism*; D003114:Colony-Forming Units Assay; D015925:Cryopreservation / Q000379:methods*; D017077:Culture Media, Conditioned; D016895:Culture Media, Serum-Free; D005312:Fetal Blood / Q000166:cytology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007231:Infant, Newborn; D007377:Interleukin-3 / Q000494:pharmacology; D008099:Liver / Q000166:cytology; D008979:Moloney murine leukemia virus / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D013539:Simian virus 40 / Q000235:genetics; D019089:Stem Cell Factor / Q000494:pharmacology
chemical_list: D017077:Culture Media, Conditioned; D016895:Culture Media, Serum-Free; D007377:Interleukin-3; D019089:Stem Cell Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor