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Publication Number
US-10513689-B2
Publication Date
2019-12-24
Expiration Date
Abstract
A culture medium for human adipose tissue derived mesenchymal stem cells includes a keratinocyte-SFM basal medium, L-Cysteine, FGF and SDF-1 is disclosed. The culture medium is effective for growing mesenchymal stem cells at a high proliferation rate while maintaining the purity, characterization and undifferentiated state of the cells.
Core Innovation
The invention provides a defined stem cell growth medium for human adipose-derived mesenchymal stem cells (human Ad-MSCs) that uses a keratinocyte-SFM basal medium. The medium is formulated with L-cysteine or glutathione plus selected growth and maintenance factors including FGF and stromal-derived factor 1 (SDF-1), together with a serum-containing balance and additional supplements selected from epidermal growth factor, hydrocortisone, calcium chloride, insulin, selenium, sodium pyruvate, transferrin, and bovine pituitary extract (BPE).
The described medium is presented as enabling high MSC proliferation while maintaining purity, an undifferentiated state, and retained stem characteristics. The stem/MSC characteristics are reported using stem cell markers, including CD73/CD90/CD105 positive and CD34/CD45/HLA-DR negative immunophenotype. The retained functional capacity is also reported as multilineage differentiation potential, including adipogenic, chondrogenic, and osteogenic differentiation potential.
An additional embodiment presents the medium in an attachment-medium context, and the document reports comparisons of the claimed medium with commercial media. The comparisons and marker-based characterizations are used to support the stated performance of the defined culture system for human Ad-MSCs.
Claims Coverage
The independent claim identified in the provided material is directed to a stem cell growth medium defined by multiple inventive features, including specified serum content plus a defined set of growth factors, hormones, salts, and supplements balanced to a serum-free medium volume to 100%.
Defined stem cell growth medium with serum-balanced volume
A stem cell growth medium comprising serum about 1% to about 10% by volume and serum free medium balance to 100% volume.
Combined growth factor, hormone, and maintenance component ranges
A stem cell growth medium comprising fibroblast growth factor 1 to about 10 ng/ml and epidermal growth factor 1 to about 10 ng/ml; hydrocortisone about 10 to about 100 ng/ml; calcium chloride 0.01 to about 0.1 mM; and insulin about 0.5 to about 5 mg/ml.
Bovine pituitary extract and redox component inclusion
A stem cell growth medium comprising bovine pituitary extract about 10 to about 100 μg/ml and L-cysteine or glutathione about 0.5 to about 5 mM.
SDF-1 plus nutritional supplements and transport protein
A stem cell growth medium comprising selenium about 0.1 to about 1 μg/ml; stromal-derived factor 1 to about 10 ng/ml; sodium pyruvate 2 to about 20 mg/ml; and transferrin 0.1 to about 1 mg/ml.
Overall, the independent claim is covered by the medium’s defined combination of serum level, multiple specified concentration ranges for FGF, EGF, hydrocortisone, calcium chloride, insulin, bovine pituitary extract, L-cysteine or glutathione, selenium, stromal-derived factor 1, sodium pyruvate, and transferrin, with the serum-free medium serving as the balance to 100% volume. Dependent claim refinements further specify serum type, recombinant-source constraints, selectable named serum-free basal media, and immunophenotype-defined multipotent stem cells using CD marker expression and absence.
Stated Advantages
High MSC proliferation while maintaining purity.
Maintenance of an undifferentiated state and retained stem characteristics.
Retained multilineage differentiation potential (adipogenic, chondrogenic, osteogenic).
Documented Applications
Culturing human adipose-derived mesenchymal stem cells (human Ad-MSCs) in a defined culture medium, with characterization by stem/MSC markers and differentiation potential.
An attachment-medium embodiment for use of the medium in an attachment-medium context.
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