Adipose tissue mesenchymal stem cell expansion in animal serum-free medium supplemented with autologous human platelet lysate.
BACKGROUND
Mesenchymal stem cells (MSCs) have been considered for human regenerative therapy applications, and safe culture and expansion protocols are needed especially in the context of interspecies contamination. Human platelet lysate (PL) has been proposed as animal serum substitute during in vitro MSC expansion. In this work, a simplified and efficient method to obtain autologous PL to replace animal serum in cell culture applications is described.


METHODS
PL obtained by freezing and centrifugation procedures was tested as medium supplement for human adipose mesenchymal stem cell (hASC) culture. Differential proliferation, immunophenotypic changes, and differentiation under PL or fetal bovine serum (FBS) were assessed.


RESULTS
In contrast to 10% FBS supplementation, cell population doubling time was significantly lower when hASCs were cultured with the same concentration of PL (PL 22.9 +/- 1.5 hr vs. FBS 106.7 +/- 6.5 hr, t test, p < 0.05). Furthermore, hASCs maintained with 2.5% PL supplementation also showed satisfactory results. Immunophenotypic analysis revealed no differences between hASCs cultivated with PL or FBS supplementation and both cultures retained the potential to differentiate into adipose cells. These results demonstrate that autologous PL obtained from the same donor can be used as animal serum substitute in hASC culture.


CONCLUSIONS
Taken together, evidence is provided that platelets provided by a single donor are sufficient to obtain PL for hASC propagation for clinical-scale applications mitigating the potential untoward side effects associated with the use of animal-derived reagents.
Blande
Ivan Souza
IS
Heart Institute (InCor), University of Sao Paulo Medical School, Sao Paulo, Brazil.
Bassaneze
Vinicius
V
Lavini-Ramos
Carolina
C
Fae
Kellen Cristhina
KC
Kalil
Jorge
J
Miyakawa
Ayumi Aurea
AA
Schettert
Isolmar Tadeu
IT
Krieger
Jose Eduardo
JE
D017667:Adipocytes / Q000166:cytology; D000273:Adipose Tissue / Q000166:cytology*; D000818:Animals; D001792:Blood Platelets; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002455:Cell Division; D002457:Cell Extracts / Q000494:pharmacology*; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D006801:Humans; D016130:Immunophenotyping; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D010011:Osteocytes / Q000166:cytology; D044967:Serum
D002457:Cell Extracts; D016895:Culture Media, Serum-Free