Publikationsdatenbank

New vs. Established: Does an adapted isolation protocol improve the yield of equine umbilical cord-derived mesenchymal stem cells? (2024)

Art

Poster

Autoren

Hollender, A.-C. (WE 17)

Trappe, Susanne (WE 2)

Stage, Hannah J. (WE 17)

Aschenbach, Jörg R. (WE 2)

Gehlen, H. (WE 17)

Kongress

Doctoral Students of Dahlem Research School School of Veterinary Medicine FREIE UNIVERSITÄT BERLIN The 13th PhD and Doctoral Symposium

FREIE UNIVERSITÄT BERLIN, 10.10.2024

Quelle

Sprache

Englisch

Kontakt

Pferdeklinik

Oertzenweg 19 b
14163 Berlin
+49 30 838 62299 / 62300
pferdeklinik@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Introduction: Equine umbilical cord-derived multipotent mesenchymal stem cells (ucMSCs) are widely used for differentiation studies and in regenerative medicine. However, depending on the protocol, the isolation can be lengthy with limited amounts of cell yields. We aimed to improve existing protocols to increase the cell harvest while reducing costs and time commitment.
Materials and Methods: In 32 horses, a standard cell isolation procedure with 3 h of collagenase 1 digestion and 12 h of antimicrobial treatment was compared to a modified collagenase 1 digestion for 24 h and 12 h antimicrobial treatment of the umbilical cord tissue. Every successfully cultured sample was characterized by FACS analysis, RT-qPCR and cell morphology. A representative group of three samples each was subjected to an investigation of trilineage differentiation potential.
Results: 91% of samples were successfully cultured. The reason for unsuccessful cultures was always bacterial or fungal contamination. The average passage interval was 2.5 days shorter (passage zero to passage one) and 7 days shorter (passage one to passage two) with the modified procedure. This faster turnover time was caused by a higher cell yield after the enzymatic digestion for 24 hours. FACS analysis (positive CD29, CD44, CD90; negative CD34, CD45, CD105) and RT-qPCR (OKT4, THY1, MYC, DNMT3B, DNND4A) showed no differences between the groups. Both sample groups showed strong osteogenic differentiation. Investigations on chondrogenic and adipogenic differentiation are ongoing.
Conclusion: A 24-h collagenase digestion improves ucMSC yield and could thereby increase the number of passages possible until senescence. Furthermore, it integrates better into the laboratory schedule by significantly reducing the need for work at night. It also reduces the requirement for collagenase. However, the antimicrobial protocol should be a target of future work to reach an isolation success of 100%.