Effects of Platelet Lysate Preparations on the Proliferation of HaCaT Cells

다국어 초록 (Multilingual Abstract)

Background: Standard protocols are lacking for the preparation of platelet lysates (PL) as an alternative to using fetal bovine serum as a cell culture supplement. This study aimed to establish optimum conditions for preparing PL for use in cell cultures.
Methods: Cell density in three pooled platelet concentrates (PC) were adjusted to 1×10 12 / L and 2×10 12 /L. PL was prepared from PC by 1 to 3 freeze-thaw (FT) cycles. HaCaT cells were cultured in media supplemented with 5% or 10% PL. Cell numbers were estimated using a Cell Counting Kit-8 (CCK-8; Dojindo Laboratories, Japan). Growth factors were quantified by using the Luminex 200 system (Luminex Corporation, USA).
Results: Cell proliferation rates in the presence of PLs were similar when prepared from PCs of both cell densities. The rates were higher in media containing 5% PL than 10% PL when prepared by two FT cycles. Concentrations of vascular endothelial growth factor (VEGF), platelet-derived growth factor-AB/BB (PDGF-AB/BB), PDGF-AA, and epidermal growth factor (EGF) were significantly higher in PL prepared from PC with a cell density of 2×10 12 /L than 1×10 12 /L PC. However, only VEGF and PDGF-AA concentrations in PLs were correlated with HaCaT cell counts.
Conclusions: The 5% PL from PC with a cell density of 1×10 12 /L prepared by two FT cy- cles treatment was the most effective condition that supported steady HaCaT cell prolifer- ation. Our finding may be useful for preparing PL-supplemented cell culture media.

참고문헌 (Reference)

1 임영애, "동결해동 활성화 혈소판풍부혈장이 세균 증식에 미치는 효과" 대한수혈학회 22 (22): 221-230, 2011

2 Baeyens W, "The use of platelet concentrates: platelet-rich plasma (PRP) and platelet-rich fibrin (PRF) in bone reconstruction prior to dental implant surgery" 31 : 521-527, 2010

3 Mazzocca AD, "The positive effects of different platelet-rich plasma methods on human muscle, bone, and tendon cells" 40 : 1742-1749, 2012

4 Valeri CR, "Release of platelet-derived growth factors and proliferation of fibroblasts in the releasates from platelets stored in the liquid state at 22 degrees C after stimulation with agonists" 46 : 225-229, 2006

5 Kakudo N, "Proliferation-promoting effect of platelet-rich plasma on human adipose-derived stem cells and human dermal fibroblasts" 122 : 1352-1360, 2008

6 Schallmoser K, "Preparation of pooled human platelet lysate (pHPL) as an efficient supplement for animal serum-free human stem cell cultures" 32 : 1523-, 2009

7 Eppley BL, "Platelet-rich plasma: a review of biology and applications in plastic surgery" 118 : 147-159, 2006

8 Pawitan JA, "Platelet rich plasma in xeno-free stem cell culture: the impact of platelet count and processing method" 7 : 329-335, 2012

9 Mehta S, "Platelet rich concentrate: basic science and current clinical applications" 22 : 432-438, 2008

10 Doucet C, "Platelet lysates promote mesenchymal stem cell expansion: a safety substitute for animal serum in cell-based therapy applications" 205 : 228-236, 2005

11 Sundin M, "No alloantibodies against mesenchymal stromal cells, but presence of anti-fetal calf serum antibodies, after transplantation in allogeneic hematopoietic stem cell recipients" 92 : 1208-1215, 2007

12 조희순, "Individual Variation in Growth Factor Concentrations in Platelet-rich Plasma and Its Influence on Human Mesenchymal Stem Cells" 대한진단검사의학회 31 (31): 212-218, 2011

13 Reinisch A, "Humanized system to propagate cord blood-derived multi potent mesenchymal stromal cells for clinical application" 2 : 371-382, 2007

14 Schallmoser K, "Human platelet lysate can replace fetal bovine serum for clinical-scale expansion of functional mesenchymal stromal cells" 47 : 1436-1446, 2007

15 Bieback K, "Human alternatives to fetal bovine serum for the expansion of mesenchymal stromal cells from bone marrow" 27 : 2331-2341, 2009

16 García-Martínez O, "Effect of platelet-rich plasma on growth and antigenic profile of human osteoblasts and its clinical impact" 70 : 1558-1564, 2012

17 Dohan Ehrenfest DM, "Do the fibrin architecture and leukocyte content influence the growth factor release of platelet concentrates? An evidence-based answer comparing a pure platelet-rich plasma (P-PRP) gel and a leukocyte- and platelet-rich fibrin (L-PRF)" 13 : 1145-1152, 2012

18 Kurita M, "Differential effects of three preparations of human serum on expansion of various types of human cells" 122 : 438-448, 2008

19 Goedecke A, "Differential effect of platelet-rich plasma and fetal calf serum on bone marrow-derived human mesenchymal stromal cells expanded in vitro" 5 : 648-654, 2011

20 Tayapongsak P, "Autologous fibrin adhesive in mandibular reconstruction with particulate cancellous bone and marrow" 52 : 161-165, 1994

21 Roussy Y, "Activation of human platelet-rich plasmas: effect on growth factors release, cell division and in vivo bone formation" 18 : 639-648, 2007

22 Lange C, "Accelerated and safe expansion of human mesenchymal stromal cells in animal serum-free medium for transplantation and regenerative medicine" 213 : 18-26, 2007