title: Surface markers and gene expression to characterize the differentiation of monolayer expanded human articular chondrocytes.
abstract: Autologous chondrocyte implantation (ACI) is a method of cartilage repair. To improve the quality of regenerated tissue by ACI, it is essential to identify surface marker expression correlated with the differentiation status of monolayer expanded human articular chondrocytes and to define the index for discriminating dedifferentiated cells from monolayer expanded human articular chondrocytes. Normal human articular chondrocytes were cultured in monolayer until passage 4. At each passage, mRNA expression of collagen type I, II, and X and aggrecan was analyzed by real-time quantitative PCR, and the surface marker expression of CD14, CD26, CD44, CD49a, CD49c, CD54, and CD151 was analyzed by fluorescence-activated cell sorting (FACS). The ratios of mRNA levels of collagen type II to I (Col II/Col I) represented the differentiation status of chondrocytes more appropriately during monolayer culture. The surface marker expression of CD44, CD49c, and CD151 was upregulated according to the dedifferentiation status, whereas that of CD14, CD49a, and CD54 was downregulated. The most appropriate combination of the ratio of Col II/Col I was CD54 and CD44. Cell sorting was performed using a magnetic cell sorting system (MACS) according to CD54 and CD44, and real-time quantitative PCR was performed for the cell subpopulations before and after cell sorting. The expression of collagen type II and aggrecan of the chondrocytes after MACS was higher than that before sorting, but not significantly. The mean fluorescence intensity (MFI) ratio of CD54 to CD44 could be an adequate candidate as the index of the differentiation status.
authors.affiliation: Department of Orthopaedic Surgery, Nagoya University Graduate School of Medicine, Nagoya, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Hamada
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Sakai
authors.forename: Hideki
authors.initials: H
authors.lastname: Hiraiwa
authors.forename: Motoshige
authors.initials: M
authors.lastname: Nakashima
authors.forename: Yohei
authors.initials: Y
authors.lastname: Ono
authors.forename: Hirohito
authors.initials: H
authors.lastname: Mitsuyama
authors.forename: Naoki
authors.initials: N
authors.lastname: Ishiguro
mesh_terms: D000293:Adolescent; D000328:Adult; D053674:Aggrecans / Q000235:genetics / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism*; D015415:Biomarkers / Q000378:metabolism; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism*; D002454:Cell Differentiation / Q000235:genetics; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D016130:Immunophenotyping; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D008297:Male; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055815:Young Adult
chemical_list: C508319:ACAN protein, human; D053674:Aggrecans; D015703:Antigens, CD; D015415:Biomarkers; C497483:CD44 protein, human; D018960:Hyaluronan Receptors; D012333:RNA, Messenger; D018799:Intercellular Adhesion Molecule-1; D003094:Collagen
references.citation: J Cell Physiol. 2005 Mar;202(3):731-42
references.pmid: 15389573
references.citation: Clin Orthop Relat Res. 2004 Oct;(427 Suppl):S152-62
references.pmid: 15480059
references.citation: Eur J Radiol. 2006 Jan;57(1):24-31
references.pmid: 16188417
references.citation: Arthroscopy. 1989;5(1):25-32
references.pmid: 2706048
references.citation: Tissue Eng. 2007 Jul;13(7):1583-92
references.pmid: 17630901
references.citation: Clin Orthop Relat Res. 1984 Jan-Feb;(182):200-5
references.pmid: 6692614
references.citation: J Immunol. 1986 Jul 1;137(1):245-54
references.pmid: 3086451
references.citation: Connect Tissue Res. 1991;26(3):207-16
references.pmid: 1722744
references.citation: Osteoarthritis Cartilage. 2002 Jan;10(1):62-70
references.pmid: 11795984
references.citation: J Orthop Res. 1990 Mar;8(2):266-75
references.pmid: 2303960
references.citation: Cell. 1982 Aug;30(1):215-24
references.pmid: 7127471
references.citation: Clin Orthop Relat Res. 1999 Oct;(367 Suppl):S147-55
references.pmid: 10546643
references.citation: Eur J Immunol. 2011 May;41(5):1365-75
references.pmid: 21469097
references.citation: Nature. 1990 Aug 2;346(6283):425-34
references.pmid: 1974032
references.citation: Biochem Biophys Res Commun. 2001 Jan 26;280(3):806-12
references.pmid: 11162592
references.citation: Blood. 2003 Feb 1;101(3):869-76
references.pmid: 12393715
references.citation: Arthroscopy. 2003 May-Jun;19(5):477-84
references.pmid: 12724676
references.citation: J Orthop Res. 1995 Jan;13(1):4-12
references.pmid: 7853102
references.citation: J Bone Joint Surg Am. 2003;85-A Suppl 3:109-15
references.pmid: 12925617
references.citation: Osteoarthritis Cartilage. 2004 Jul;12(7):525-35
references.pmid: 15219567
references.citation: J Orthop Res. 1998 Mar;16(2):207-16
references.pmid: 9621895
references.citation: Int J Mol Med. 2009 Feb;23(2):253-9
references.pmid: 19148550
references.citation: Biotechnol Bioeng. 2009 Mar 1;102(4):1259-67
references.pmid: 18949749
references.citation: Arch Oral Biol. 2008 Apr;53(4):330-6
references.pmid: 18160062
references.citation: Circulation. 2001 Jun 19;103(24):2885-90
references.pmid: 11413075
references.citation: Instr Course Lect. 1998;47:487-504
references.pmid: 9571450
references.citation: J Cell Physiol. 2010 Jan;222(1):248-53
references.pmid: 19795395
references.citation: Arthritis Rheum. 2007 Feb;56(2):586-95
references.pmid: 17265493
references.citation: Cell. 1990 Jun 29;61(7):1303-13
references.pmid: 1694723
references.citation: Adv Drug Deliv Rev. 2004 Apr 19;56(6):763-78
references.pmid: 15063588
references.citation: J Pathol. 1996 Aug;179(4):396-402
references.pmid: 8869287
references.citation: J Orthop Res. 2008 Sep;26(9):1230-7
references.pmid: 18404652
references.citation: Osteoarthritis Cartilage. 2007 Dec;15(12):1339-47
references.pmid: 17629514
references.citation: Arthritis Rheum. 2001 Aug;44(8):1800-7
references.pmid: 11508432
references.citation: Clin Orthop Relat Res. 2001 Oct;(391 Suppl):S26-33
references.pmid: 11603710
references.citation: J Bone Joint Surg Br. 1967 May;49(2):211-28
references.pmid: 6026508
references.citation: Lasers Surg Med. 1989;9(6):595-601
references.pmid: 2601554