Human Platelet Lysate as a Replacement for Fetal Bovine Serum in Limbal Stem Cell Therapy

Kunal Suri, Hwee K. Gong, Ching Yuan, Stephen C. Kaufman

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Purpose: To evaluate the use of human platelet lysate (HPL) as an alternative supplement for limbal explant culture.Methods: Culture media were prepared using either 10% pooled HPL (PHPL), single donor HPL, or fetal bovine serum (FBS). Limbal tissues, obtained from the Minnesota Lions Eye Bank, were cultured in each medium on plastic plates or on denuded amniotic membrane (AM). Immunofluorescence staining was performed for ABCG2, tumor protein p63α, and cytokeratin 3 (K3). Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to evaluate the expression of ABCG2 and p63. Limbal explants grown in each medium were labeled with bromodeoxyuridine (BrdU) to assess the proliferative capacity in each medium. Concentration of growth factors including epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), transforming growth factor-β (TGF-β), and platelet derived growth factor (PDGF) in HPL and PHPL was compared to that in human serum (HS).Results: Immunofluorescence staining on AM showed prominent expression of ABCG2, p63α but sparse expression of K3 in HPL and PHPL supplemented medium. Real time-PCR showed 1.7 fold higher expression of ABCG2 in PHPL supplemented medium (p = 0.03), and similar expression of p63 in HPL and PHPL supplemented medium compared to FBS medium. The proliferation assay showed that LSCs retained their proliferative potential in HPL supplemented medium. Higher concentration of growth factors were found in HPL, compared to HS.Conclusions: Human platelet lysate has higher concentration of grown factors and is effective in maintaining growth and stem cell phenotype of corneal limbal explant cultures.

Original languageEnglish (US)
Pages (from-to)1266-1273
Number of pages8
JournalCurrent Eye Research
Volume41
Issue number10
DOIs
StatePublished - Oct 2 2016

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Cell- and Tissue-Based Therapy
Stem Cells
Blood Platelets
Serum
Amnion
Fluorescent Antibody Technique
Intercellular Signaling Peptides and Proteins
Keratin-3
Eye Banks
Lions
Staining and Labeling
Platelet-Derived Growth Factor
Transforming Growth Factors
Bromodeoxyuridine
Reverse Transcriptase Polymerase Chain Reaction
Epidermal Growth Factor
Vascular Endothelial Growth Factor A
Plastics
Culture Media
Real-Time Polymerase Chain Reaction

Keywords

  • Cornea
  • limbal stem cell deficiency
  • limbus
  • platelet lysate
  • stem cells

Cite this

Human Platelet Lysate as a Replacement for Fetal Bovine Serum in Limbal Stem Cell Therapy. / Suri, Kunal; Gong, Hwee K.; Yuan, Ching; Kaufman, Stephen C.

In: Current Eye Research, Vol. 41, No. 10, 02.10.2016, p. 1266-1273.

Research output: Contribution to journalArticle

Suri, Kunal ; Gong, Hwee K. ; Yuan, Ching ; Kaufman, Stephen C. / Human Platelet Lysate as a Replacement for Fetal Bovine Serum in Limbal Stem Cell Therapy. In: Current Eye Research. 2016 ; Vol. 41, No. 10. pp. 1266-1273.
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AB - Purpose: To evaluate the use of human platelet lysate (HPL) as an alternative supplement for limbal explant culture.Methods: Culture media were prepared using either 10% pooled HPL (PHPL), single donor HPL, or fetal bovine serum (FBS). Limbal tissues, obtained from the Minnesota Lions Eye Bank, were cultured in each medium on plastic plates or on denuded amniotic membrane (AM). Immunofluorescence staining was performed for ABCG2, tumor protein p63α, and cytokeratin 3 (K3). Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to evaluate the expression of ABCG2 and p63. Limbal explants grown in each medium were labeled with bromodeoxyuridine (BrdU) to assess the proliferative capacity in each medium. Concentration of growth factors including epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), transforming growth factor-β (TGF-β), and platelet derived growth factor (PDGF) in HPL and PHPL was compared to that in human serum (HS).Results: Immunofluorescence staining on AM showed prominent expression of ABCG2, p63α but sparse expression of K3 in HPL and PHPL supplemented medium. Real time-PCR showed 1.7 fold higher expression of ABCG2 in PHPL supplemented medium (p = 0.03), and similar expression of p63 in HPL and PHPL supplemented medium compared to FBS medium. The proliferation assay showed that LSCs retained their proliferative potential in HPL supplemented medium. Higher concentration of growth factors were found in HPL, compared to HS.Conclusions: Human platelet lysate has higher concentration of grown factors and is effective in maintaining growth and stem cell phenotype of corneal limbal explant cultures.

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KW - stem cells

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