Posology and Serum-/Xeno-Free Engineered Adipose Stromal Cells Cell Sheets

Area of Science:

• Regenerative Medicine
• Biomaterials Science
• Cell Biology

Background:

• Adipose stem cells (ASCs) are crucial for regenerative medicine.
• Cell sheet engineering requires well-defined culture media for translational applications.
• Standard cell culture often relies on serum-containing or animal-derived components, posing risks.

Purpose of the Study:

• To develop and engineer multilayer adipose stem cell sheets (ASCCSs) using chemically defined, serum-free, and xeno-free culture media.
• To differentiate ASCCSs into osteoblasts and chondrocytes.
• To establish a non-invasive method for estimating cell numbers within engineered cell sheets.

Main Methods:

• Engineered undifferentiated ASCCSs in 10 days using serum-free/xeno-free media.
• Differentiated ASCCSs into osteoblasts and chondrocytes in 14 days using chemically defined, animal-free media.
• Utilized cell sheet transmittance to quantify cell numbers and assessed the impact of ROCK inhibitor on chondrocyte differentiation.

Main Results:

• Successfully engineered undifferentiated ASCCSs expressing specific surface markers (CD29+, CD73+, CD90+, CD105+, HLA-A+, HLA-DR-).
• Engineered differentiated ASCCSs (osteoblasts and chondrocytes) within 14 days, with ROCK inhibitor improving chondrocyte sheets.
• Observed higher transmittance decrease in osteoblast sheets due to Ca2+ accumulation, enabling cell number estimation.

Conclusions:

• Developed three types of ASCCSs (undifferentiated, osteoblast, chondrocyte) using serum-free, xeno-free media.
• Demonstrated a non-invasive transmittance-based method to estimate cell numbers per sheet, crucial for cell sheet posology.
• The engineered ASCCSs hold potential for translational medicine applications.