Effects of Decade Long Freezing Storage on Adipose Derived Stem Cells Functionality

Area of Science:

• Regenerative Medicine
• Cell Biology
• Biotechnology

Background:

• Adipose tissue derived stromal/stem cells (ASCs) are crucial for regenerative medicine.
• Optimizing cryopreservation is essential for long-term storage and clinical application of ASCs.
• Previous studies focused on short-term cryopreservation parameters.

Purpose of the Study:

• To evaluate the functionality of ASCs after long-term cryopreservation (over 10 years) compared to short-term (3-7 years) and fresh ASCs.
• To assess post-thaw viability, immunophenotype, osteogenic, and adipogenic differentiation potential.
• To investigate the impact of extended cryopreservation on ASC gene expression.

Main Methods:

• Cryopreservation of ASCs for short-term and long-term storage.
• Post-thaw viability assessment.
• Flow cytometry for immunophenotype analysis (CD29, CD90, CD105, CD44, CD73, CD31, CD34, CD45, CD146).
• Quantitative real-time polymerase chain reaction (qPCR) for osteogenic gene expression.
• In vitro osteogenic and adipogenic differentiation assays.

Main Results:

• No significant difference in post-thaw viability between long-term (78%) and short-term (79%) frozen ASCs.
• Immunophenotype markers remained consistent across all groups (fresh, short-term, long-term frozen).
• Long-term cryopreservation showed a slight negative impact on osteogenic potential, notably decreasing osteopontin expression, while adipogenic potential was unaffected.

Conclusions:

• ASCs can be successfully cryopreserved for over 10 years with preserved viability and immunophenotype.
• Long-term cryopreservation may subtly affect osteogenic differentiation capacity.
• ASCs remain a viable cell source for regenerative medicine applications even after extended storage.