title: Induction of chondrogenesis of adipose-derived stem cells by novel recombinant TGF-β3 fusion protein.
abstract: A new type of TGF-β3 fusion protein with targeted therapy function was constructed, and its feasibility and target specificity of inducing chondrogenesis were investigated by transfecting LAP-MMP-mTGF-β3 gene into adipose-derived stem cells (ADSCs). The recombinant pIRES-EGFP-MMP was constructed by inserting the sense and antisense DNA of encoding the amino acid of the synthetic MMP enzyme cutting site into the eukaryotic expression vector pIRES-EGFP. LAP and mTGF-β3 fragments were obtained by using RT-PCR and inserted into the upstream and downstream of MMP from pIRES-EGFP-MMP respectively, and the recombinant plasmid of pIRES-EGFP-LAP-MMP-mTGF-β3 was constructed, which was transferred to ADSCs. The ADSCs were cultured and divided in three groups: experimental group (MMP group), negative control group (no MMP) and non-transfection group. The morphological changes were observed microscopically, and the expression of proteoglycan and type II collagen (ColII) was detected by using Alcian blue staining and immunohistochemistry staining at 7th, 14th and 21st day after culture. The recombinant plasmid of pIRES-EGFP-LAP-MMP-mTGF-β3 was correctly constructed by methods of enzyme cutting and sequencing analysis. The mTGF-β3 fusion protein was successfully expressed after transfection, and in the presence of the MMP, active protein mTGF-β3 was generated, which significantly promoted differentiation of ADSCs into chondrocytes and the expression of cartilage matrix. The novel fusion protein LAP-MMP-mTGF-β3 can targetedly induce differentiation of ADSCs into chondrocytes, which would open up prospects for target therapy of cartilage damage repair in future.
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Dong
authors.initials: D
authors.lastname: Zheng
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Dan
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Huang
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China. Xiatian@sina.com.
authors.forename: Tian
authors.initials: T
authors.lastname: Xia
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Shu-Hua
authors.initials: SH
authors.lastname: Yang
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Wei-Hua
authors.initials: WH
authors.lastname: Xu
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Cao
authors.initials: C
authors.lastname: Yang
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Guo-Hui
authors.initials: GH
authors.lastname: Liu
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Xian-Zhe
authors.initials: XZ
authors.lastname: Liu
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Feng
mesh_terms: D000273:Adipose Tissue / Q000378:metabolism*; D000818:Animals; D020219:Chondrogenesis / Q000235:genetics*; D005260:Female; D008297:Male; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism*; D013234:Stem Cells / Q000378:metabolism*; D053782:Transforming Growth Factor beta3 / Q000235:genetics* / Q000378:metabolism*
chemical_list: D011993:Recombinant Fusion Proteins; C509046:Tgfb3 protein, rat; D053782:Transforming Growth Factor beta3
references.citation: Biorheology. 2004;41(3-4):389-99
references.pmid: 15299271
references.citation: Nat Clin Pract Rheumatol. 2007 Dec;3(12):716-24
references.pmid: 18037931
references.citation: J Biochem. 1998 Mar;123(3):431-9
references.pmid: 9538225
references.citation: Osteoarthritis Cartilage. 1998 Sep;6(5):306-17
references.pmid: 10197165
references.citation: Cytokine Growth Factor Rev. 2002 Jun;13(3):241-57
references.pmid: 12486877
references.citation: Biopolymers. 2005;80(6):736-46
references.pmid: 15880774
references.citation: Dev Dyn. 2008 Aug;237(8):2013-20
references.pmid: 18330928
references.citation: Arthritis Rheum. 2005 Aug;52(8):2386-95
references.pmid: 16052587
references.citation: Methods Mol Biol. 2011;698:253-78
references.pmid: 21431525
references.citation: J Orthop Res. 2002 May;20(3):535-44
references.pmid: 12038628
references.citation: Biomaterials. 2010 May;31(14):3858-67
references.pmid: 20153043
references.citation: J Cell Physiol. 2005 May;203(2):345-52
references.pmid: 15468069
references.citation: Ann Rheum Dis. 2007 Jul;66(7):916-20
references.pmid: 17314121
references.citation: Arthritis Rheum. 1993 Jan;36(1):44-50
references.pmid: 8424835
references.citation: Arthritis Rheum. 2010 Aug;62(8):2347-52
references.pmid: 20506137
references.citation: J Biol Chem. 2004 Sep 3;279(36):38032-9
references.pmid: 15208302
references.citation: Tissue Cell. 2003 Feb;35(1):69-77
references.pmid: 12589731
references.citation: J Biomed Mater Res A. 2006 Jun 15;77(4):785-94
references.pmid: 16575912
references.citation: Tissue Eng. 2001 Apr;7(2):211-28
references.pmid: 11304456
references.citation: Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2005 May;21(3):349-52
references.pmid: 15862156
references.citation: Hum Gene Ther. 1997 Apr 10;8(6):729-38
references.pmid: 9113512
references.citation: Rheumatol Int. 2008 Nov;29(1):31-6
references.pmid: 18597092
references.citation: Clin Orthop Relat Res. 2004 Oct;(427 Suppl):S27-36
references.pmid: 15480070
references.citation: Circ Res. 2007 May 11;100(9):1249-60
references.pmid: 17495232