Human telomerase reverse transcriptase and glucose-regulated protein 78 increase the life span of articular chondrocytes and their repair potential

Department of Orthopaedic Surgery, Surgical Science, Tokai University School of Medicine, Isehara, Kanagawa, Japan.
BMC Musculoskeletal Disorders (Impact Factor: 1.72). 04/2012; 13:51. DOI: 10.1186/1471-2474-13-51
Source: PubMed

ABSTRACT Like all mammalian cells, normal adult chondrocytes have a limited replicative life span, which decreases with age. To facilitate the therapeutic use of chondrocytes from older donors, a method is needed to prolong their life span.
We transfected chondrocytes with hTERT or GRP78 and cultured them in a 3-dimensional atelocollagen honeycomb-shaped scaffold with a membrane seal. Then, we measured the amount of nuclear DNA and glycosaminoglycans (GAGs) and the expression level of type II collagen as markers of cell proliferation and extracellular matrix formation, respectively, in these cultures. In addition, we allografted this tissue-engineered cartilage into osteochondral defects in old rabbits to assess their repair activity in vivo.
Our results showed different degrees of differentiation in terms of GAG content between chondrocytes from old and young rabbits. Chondrocytes that were cotransfected with hTERT and GRP78 showed higher cellular proliferation and expression of type II collagen than those of nontransfected chondrocytes, regardless of the age of the cartilage donor. In addition, the in vitro growth rates of hTERT- or GRP78-transfected chondrocytes were higher than those of nontransfected chondrocytes, regardless of donor age. In vivo, the tissue-engineered cartilage implants exhibited strong repairing activity, maintained a chondrocyte-specific phenotype, and produced extracellular matrix components.
Focal gene delivery to aged articular chondrocytes exhibited strong repairing activity and may be therapeutically useful for articular cartilage regeneration.

Download full-text


Available from: Masato Sato, Sep 26, 2015
16 Reads
  • Source
    • "Human telomerase is a ribonucleoprotein that adds TTAGGG repeats to telomere ends of linear chromosomes in eukaryotes (Sato et al., 2012). Telomerase activity is not evident in normal cells, with elimination of germ cells and regeneration tissues (Buseman et al., 2012). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Background: Diosgenin, a steroidal saponin from a therapeutic herb, fenugreek (Trigonellafoenum-graceum L.), has been recognized to have anticancer properties. Telomerase activity is not detected in typical healthy cells, while in cancer cell telomerase expression is reactivated, therefore providing a promising cancer therapeutic target. Materials and methods: We studied the inhibitory effect of diosgenin on human telomerase reverse transcriptase gene (hTERT) expression which is critical for telomerase activity. MTT- assays and qRT-PCR analysis were conducted to assess cytotoxicity and hTERT gene expression inhibition effects, respectively. Results: MTT results showed that IC50 values for 24, 48 and 72h after treatment were 47, 44 and 43μM, respectively. Culturing cells with diosgenin treatment caused down-regulation of hTERT expression. Discussion: These results show that diosgenin inhibits telomerase activity by down-regulation of hTERT gene expression in the A549 lung cancer cell line.
    Asian Pacific journal of cancer prevention: APJCP 11/2013; 14(11):6945-8. DOI:10.7314/APJCP.2013.14.11.6945 · 2.51 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Heat-shock proteins (HSPs) are molecular chaperones that are highly conserved between species. In recent decades it has become clear that these proteins play an important role in the pathogenesis of inflammatory and degenerative joint diseases by (dys)regulating the immune system and by direct effects on the stromal tissues of the joint. In this review we discuss current insights into the expression pattern of HSPs in connective tissues, the direct biological role of HSPs in stromal tissues and the potential clinical applications.
    Rheumatology (Oxford, England) 08/2013; 53(2). DOI:10.1093/rheumatology/ket277 · 4.48 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In the last few years, great progress has been made to validate tissue engineering strategies in preclinical studies and clinical trials on the regeneration of osteochondral defects. In the preclinical studies, one of the dominant strategies comprises the development of biomimetic/bioactive scaffolds, which are used alone or incorporated with growth factors and/or stem cells. Many new trends are emerging for modulation of stem cell fate towards osteogenic and chondrogenic differentiations, but bone/cartilage interface regeneration and physical stimulus have been showing great promise. Besides the matrix-associated autologous chondrocyte implantation (MACI) procedure, the matrix-associated stem cells implantation (MASI) and layered scaffolds in acellular or cellular strategy are also applied in clinic. This review outlines the progresses at preclinical and clinical levels, and identifies the new challenges in osteochondral tissue engineering. Future perspectives are provided, e.g., the applications of extracellular matrix-like biomaterials, computer-aided design/manufacture of osteochondral implant and reprogrammed cells for osteochondral regeneration
    02/2015; 1(4):150220124046001. DOI:10.1021/ab500038y