title: Chitosan-plasmid DNA nanoparticles encoding small hairpin RNA targeting MMP-3 and -13 to inhibit the expression of dedifferentiation related genes in expanded chondrocytes.
abstract: Overexpression of matrix metalloproteinase (MMP)-3 and -13 can lead to the dedifferentiation of expanded chondrocytes. After implanting dedifferentiated cells for cartilage defect repair, graft failure may occur. Short hairpin RNA (shRNA) is a powerful genetic tool to reduce the expression of target genes. This study investigated the effects of chitosan-plasmid DNA (pDNA) nanoparticles encoding shRNA targeting MMP-3 and -13 on the dedifferentiation of expanded chondrocytes. The objective was to optimize the parameters of chitosan-pDNA formulation for achieving higher efficiency of pDNA delivery and gene silencing. The chitosan-pDNA nanoparticles were prepared using a complex coacervation process. Then the characteristics including size, shape, stability, and transfection efficiency were compared in different groups. The results indicated that chitosan of 800 kDa at N/P ratio of 4 and pH 7.0 was optimal to prepare chitosan-pDNA nanoparticles. These nanoparticles showed high DNA loading efficiency (95.8 ± 1.5%) and high gene transfection efficiency (24.5 ± 1.6%). After the expanded chondrocytes were transfected by chitosan-pDNA nanoparticles, MMP-3-610 and MMP-13-2024 groups showed greater suppression in mRNA and protein levels. The results indicated that chitosan-pDNA nanoparticles encoding shRNA targeting MMP-3 and -13 had great potential in silencing the dedifferentiation-related genes for regenerating prolonged and endurable cartilage.
authors.affiliation: Department of Sports Medicine, Xi-Jing Hospital, the Fourth Military Medical University, Xi'an 710032, Shaanxi Province, China; Department of Orthopaedic Surgery, the 82nd Hospital of PLA, Huai'an 223001, Jiangsu Province, China.
authors.forename: Jingxin
authors.initials: J
authors.lastname: Zhao
authors.forename: Xiangli
authors.initials: X
authors.lastname: Fan
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhang
authors.forename: Fangfei
authors.initials: F
authors.lastname: Sun
authors.forename: Xiaojian
authors.initials: X
authors.lastname: Li
authors.forename: Chuan
authors.initials: C
authors.lastname: Xiong
authors.forename: Chunli
authors.initials: C
authors.lastname: Zhang
authors.forename: Hongbin
authors.initials: H
authors.lastname: Fan
mesh_terms: D000818:Animals; D054337:Cell Dedifferentiation; D002478:Cells, Cultured; D048271:Chitosan / Q000737:chemistry*; D019902:Chondrocytes / Q000166:cytology / Q000201:enzymology*; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D018390:Gene Targeting / Q000379:methods*; D053509:Matrix Metalloproteinase 13 / Q000096:biosynthesis* / Q000235:genetics; D019278:Matrix Metalloproteinase 3 / Q000096:biosynthesis* / Q000235:genetics; D053758:Nanoparticles / Q000737:chemistry*; D010957:Plasmids / Q000737:chemistry* / Q000235:genetics; D034741:RNA, Small Interfering / Q000096:biosynthesis* / Q000235:genetics; D011817:Rabbits
chemical_list: D034741:RNA, Small Interfering; D004247:DNA; D048271:Chitosan; D053509:Matrix Metalloproteinase 13; D019278:Matrix Metalloproteinase 3
keywords: MMP; chitosan; chondrocyte; nanoparticle; short hairpin RNA