A new standardized clinical-grade protocol for banking human umbilical cord tissue cells.
BACKGROUND
Mesenchymal stromal cells (MSCs) isolated from human umbilical cord tissue (UCT) can be considered the perfect candidates for cell-based therapies and regenerative medicine. UCT-derived MSCs can be cryogenically stored in cell banks and expanded as needed for therapeutic uses.


METHODS
We developed a new method for UCT-MSC isolation, cryopreservation, and expansion, following all criteria required by a stem cell bank. UCT-MSCs were isolated either by manual dissociation (MM) or by a semiautomatic dissociation system (SAM). In both protocols UCTs were treated enzymatically using Type IV collagenase good manufacturing practices (GMP) graded and hyaluronidase (medicinal product). Isolated UCT-MSCs were cryopreserved and analyzed after thawing for phenotype; for proliferation rate; and for their osteogenic, adipogenic, and chondrogenic differentiation capabilities.


RESULTS
We found that SAM reduced the time of tissue enzyme exposure and enabled us to obtain a homogeneous single-cell suspension deprived of tissue fragments. The isolated cells in both groups showed high expression of MSC markers CD105, CD73, and CD90 and similar differentiation capabilities, phenotype, and proliferation potential. Moreover, the final yield of MSCs was comparable between the two techniques.


CONCLUSIONS
In this study, we have established a reliable and standardized protocol to isolate UCT-MSCs from UCT for cell banking purposes. Processing the whole umbilical tissue with GMP-graded enzymes using a semiautomatic dissociator allowed us to obtain a single-cell suspension product with a known number of isolated cells that can be cryopreserved right after isolation and thawed as needed for expansion and clinical use.
InScientiaFides Foundation, Republic of San Marino; and.
Raffaella
R
Fazzina
Institute of Gynecology and Obstetrics, Sacred Heart Catholic University.
Andrea
A
Mariotti
Institute of Gynecology and Obstetrics, Sacred Heart Catholic University.
Annabella
A
Procoli
Immunohematology and Transfusion Medicine, San Camillo Forlanini Hospital.
Daniela
D
Fioravanti
Immunohematology and Transfusion Medicine, San Camillo Forlanini Hospital.
Paola
P
Iudicone
Institute of Gynecology and Obstetrics, Sacred Heart Catholic University.
Giovanni
G
Scambia
Immunohematology and Transfusion Medicine, San Camillo Forlanini Hospital.|Department of Experimental Medicine, Sapienza University, Rome, Italy.
Luca
L
Pierelli
Institute of Gynecology and Obstetrics, Sacred Heart Catholic University.
Giuseppina
G
Bonanno
D002469:Cell Separation / Q000379:methods; D002985:Clinical Protocols / Q000592:standards*; D015925:Cryopreservation / Q000379:methods; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology; D014015:Tissue Banks / Q000592:standards*; D014470:Umbilical Cord / Q000166:cytology*