Cilvēka dermas mezenhimālo šūnu ilgtermiņa kultūru raksturošana un ilgtermiņa uzglabāšanas optimizēšana
Date
2011
Authors
Kungs, Jānis
Journal Title
Journal ISSN
Volume Title
Publisher
Latvijas Universitāte
Abstract
No cilvēka ādas izdalītas mezenhimālās cilmes šūnas (MCŠ) ir multipotentas šūnas ar potenciālu pielietojumam klīniskajos pētījumos. Lai novērtētu ilgtermiņa in vitro pavairošanas ietekmi uz dermālo MCŠ kultūrām, tika pārbaudīta kariotipa stabilitāte, TERT, diferenciācijas marķieru un ar apoptozi saistītu gēnu ekspresijas izmaiņas pēc 5, 25 un 50 populācijas dalīšanās reizēm (PD). Papildus tika pilnveidota laboratorijā izmantotā ilgtermiņa krioprezervācijas metode.
Pēc PD50 šūnas saglabāja normālu kariotipu, neuzsāka TERT ekspresiju, bet pakāpeniski palielinājās apoptozes līmenis kultūrās. Diferenciācijas marķieru ekspresijas panelī tika novērotas ar ar atsevišķām MCŠ kultūrām (iegūtas no atšķirīgiem indivīdiem) saistītas variācijas. Glicerīns tika atzīts par nepiemērotu krioprotektantu ilgtermiņa krioprezervācijā.
Human skin-derived mesenchymal stem cells (MSC) are multipotent cells with a potential for applications in clinical trials. To evaluate the influence of long-term in vitro expansion on dermal MSC cultures, karyotype stability, TERT, differentiation marker and apoptosis-related gene expression changes were tested after 5, 25 and 50 population doublings (PD). In addition long-term cryopreservation method used in the laboratory was improved. After 50 PDs cells maintained normal karyotype, did not express TERT gene, however apoptosis level of the cultures steadily increased. Individual MSC culture (established from different individuals) related differences were observed in differentiation marker expression panel. It was concluded that glycerol is unsuitable cryoprotectant for long-term cryopreservation.
Human skin-derived mesenchymal stem cells (MSC) are multipotent cells with a potential for applications in clinical trials. To evaluate the influence of long-term in vitro expansion on dermal MSC cultures, karyotype stability, TERT, differentiation marker and apoptosis-related gene expression changes were tested after 5, 25 and 50 population doublings (PD). In addition long-term cryopreservation method used in the laboratory was improved. After 50 PDs cells maintained normal karyotype, did not express TERT gene, however apoptosis level of the cultures steadily increased. Individual MSC culture (established from different individuals) related differences were observed in differentiation marker expression panel. It was concluded that glycerol is unsuitable cryoprotectant for long-term cryopreservation.
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Bioloģija