A combination of catalase and trehalose as additives to conventional freezing medium results in improved cryoprotection of human hematopoietic cells with reference to in vitro migration and adhesion properties.
BACKGROUND
Cryopreservation of hematopoietic cells from cord blood is an essential component in unrelated transplant settings. Cell damage during freezing is caused by multiple factors, of which membrane damage and oxygen free radical generation are two major factors. It was reported earlier that a combination of catalase and trehalose as additives in freezing medium affords better cryoprotection in terms of long-term culture assays.


METHODS
Mononuclear and CD34+ cells isolated from cord blood were used as a source of hematopoietic cells. KG1a cell line was used as a model system in adhesion assays. The cells were frozen in a programmable freezer and stored at -196 degrees C. Various homing-related assays were carried out on the frozen cells.


RESULTS
Herein it is reported that these two additives afford better preservation of adhesion- and migration-related properties of the frozen cells. The test cells frozen with additives resulted in improved migration toward stromal-derived factor-1alpha and showed higher expression of its receptor CXCR4. Colony-forming unit assay of migrated test cells showed that these cells are early progenitors having capacity to give rise to all types of myeloid colonies. Test cells also show increased expression of FLT3R and improved responsiveness to FLT3 ligand, the homing-related cytokine. Adhesion to stroma and extracellular matrix was better in test cells as compared to control cells.


CONCLUSIONS
The present data provide evidence that addition of catalase and trehalose to the conventional freezing medium preserves migration- and adhesion-related properties of the hematopoietic graft.
Sasnoor
Lalita M
LM
National Center for Cell Science, Ganeshkhind, Pune, India.
Kale
Vaijayanti P
VP
Limaye
Lalita S
LS
D000906:Antibodies / Q000494:pharmacology; D018952:Antigens, CD34 / Q000378:metabolism; D001793:Blood Preservation / Q000379:methods; D002374:Catalase / Q000494:pharmacology*; D002448:Cell Adhesion / Q000187:drug effects; D002450:Cell Communication; D002465:Cell Movement / Q000187:drug effects; D054377:Chemokine CXCL12; D019743:Chemokines, CXC / Q000494:pharmacology; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology*; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D011518:Proto-Oncogene Proteins / Q000494:pharmacology; D020794:Receptor Protein-Tyrosine Kinases / Q000494:pharmacology; D019718:Receptors, CXCR4 / Q000276:immunology / Q000378:metabolism; D017154:Stromal Cells / Q000166:cytology; D014199:Trehalose / Q000494:pharmacology*; D051941:fms-Like Tyrosine Kinase 3
D000906:Antibodies; D018952:Antigens, CD34; C515255:CXCL12 protein, human; D054377:Chemokine CXCL12; D019743:Chemokines, CXC; D003451:Cryoprotective Agents; D011518:Proto-Oncogene Proteins; D019718:Receptors, CXCR4; D014199:Trehalose; D002374:Catalase; C497970:FLT3 protein, human; D020794:Receptor Protein-Tyrosine Kinases; D051941:fms-Like Tyrosine Kinase 3