Expression of cartilage‐related genes in bovine synovial tissue

Shintani, Nahoko; Kurth, Tobias B.; Hunziker, Ernst B.

doi:10.1002/jor.20345

Cited by 26 publications

(13 citation statements)

References 29 publications

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“…The following TaqMan ® Gene Expression Assays were used for detection of cartilaginous gene expression: COL2A1 (Bt03251861_m1), COL1A1 (Bt03225322_m1), ACAN (Bt03212186_m1), and SOX9 (custom designed forward primer: ACGCCGAGCTCAGCAAGA; reverse primer: CACGAACGGCCGCTTCT; probe: CGTTCA-GAAGTCTCCAGAGCTTGCCCA). 23 Gene expression values were reported in relative levels to GAPDH (Bt03210913_g1) by the 2 −Δ C t method. 24 All reactions were performed in triplicates.…”

Section: Methodsmentioning

confidence: 99%

Transient hypoxia improves matrix properties in tissue engineered cartilage

Yodmuang

Gadjanski

Chao

et al. 2012

Journal Orthopaedic Research

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Adult articular cartilage is a hypoxic tissue, with oxygen tension ranging from <10% at the cartilage surface to <1% in the deepest layers. In addition to spatial gradients, cartilage development is also associated with temporal changes in oxygen tension. However, a vast majority of cartilage tissue engineering protocols involves cultivation of chondrocytes or their progenitors under ambient oxygen concentration (21% O2), that is, significantly above physiological levels in either developing or adult cartilage. Our study was designed to test the hypothesis that transient hypoxia followed by normoxic conditions results in improved quality of engineered cartilaginous ECM. To this end, we systematically compared the effects of normoxia (21% O2 for 28 days), hypoxia (5% O2 for 28 days) and transient hypoxia—reoxygenation (5% O2 for 7 days and 21% O2 for 21 days) on the matrix composition and expression of the chondrogenic genes in cartilage constructs engineered in vitro. We demonstrated that reoxygenation had the most effect on the expression of cartilaginous genes including COL2A1, ACAN, and SOX9 and increased tissue concentrations of amounts of glycosaminoglycans and type II collagen. The equilibrium Young’s moduli of tissues grown under transient hypoxia (510.01 ± 28.15 kPa) and under normoxic conditions (417.60 ± 68.46 kPa) were significantly higher than those measured under hypoxic conditions (279.61 ± 20.52 kPa). These data suggest that the cultivation protocols utilizing transient hypoxia with reoxygenation have high potential for efficient cartilage tissue engineering, but need further optimization in order to achieve higher mechanical functionality of engineered constructs.

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Section: Methodsmentioning

confidence: 99%

Transient hypoxia improves matrix properties in tissue engineered cartilage

Yodmuang

Gadjanski

Chao

et al. 2012

Journal Orthopaedic Research

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“…Aggregates of synovial and bone-marrow-derived MSCs were prepared by culturing them under micromass conditions, according to a modifi ed version of published protocols (De Bari et al, 2001b;Park et al, 2005;Shintani and Hunziker, 2007). Specifi cally, the fi rst-passage MSCs were suspended at a numerical density of 2x10 7 per mL; 50-μL aliquots (~1 million cells per aggregate culture) were applied to 12-well tissue-culture plates, which were incubated for 4 h at 37 °C in a humidifi ed atmosphere containing 5 % CO 2 .…”

Section: Micromass Culturesmentioning

confidence: 99%

Differential effects of dexamethasone on the chondrogenesis of mesenchymal stromal cells: Influence of microenvironment, tissue origin and growth factor

Shintani¹,

Hunziker²

2011

eCM

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Mesenchymal stromal cells (MSCs), which reside within various tissues, are utilized in the engineering of cartilage tissue. Dexamethasone (DEX) -a synthetic glucocorticoid -is almost invariably applied to potentiate the growthfactor-induced chondrogenesis of MSCs in vitro, albeit that this effect has been experimentally demonstrated only for transforming-growth-factor-beta (TGF-β)-stimulated bone-marrow-derived MSCs. Clinically, systemic glucocorticoid therapy is associated with untoward side effects (e.g., bone loss and increased susceptibility to infection). Hence, the use of these agents should be avoided or limited. We hypothesize that the infl uence of DEX on the chondrogenesis of MSCs depends upon their tissue origin and microenvironment [absence or presence of an extracellular matrix (ECM)], as well as upon the nature of the growth factor. We investigated its effects upon the TGF-β1-and bone-morphogenetic-protein 2 (BMP-2)-induced chondrogenesis of MSCs as a function of tissue source (bone marrow vs. synovium) and microenvironment [cell aggregates (no ECM) vs. explants (presence of a natural ECM)]. In aggregates of bone-marrow-derived MSCs, DEX enhanced TGF-β1-induced chondrogenesis by an up-regulation of cartilaginous genes, but had little infl uence on the BMP-2-induced response. In aggregates of synovial MSCs, DEX exerted no remarkable effect on either TGF-β1-or BMP-2-induced chondrogenesis. In synovial explants, DEX inhibited BMP-2-induced chondrogenesis almost completely, but had little impact on the TGF-β1-induced response. Our data reveal that steroids are not indispensable for the chondrogenesis of MSCs in vitro. Their infl uence is context dependent (tissue source of the MSCs, their microenvironment and the nature of the growth factor). This fi nding has important implications for MSCbased approaches to cartilage repair.

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“…For calibration purposes, cartilage was collected from the metacarpal joints of 4 different animals. RNA was isolated and real-time PCR was performed as previously described (32). The samples were homogenized in QIAzol lysis reagent (Qiagen), and RNA was isolated using the RNeasy Micro Kit (Qiagen).…”

Section: ϫ7mentioning

confidence: 99%

“…Real-time PCR was performed using the ABI Prism 7700 sequence detection system (Applied Biosystems, Foster City, CA). Primers and probes for bovine CI, CII, CIX, CX, and CXI, aggrecan, COMP, SOX9, and 18S ribosomal RNA (rRNA) were generated as previously described (32). Gene expression was normalized to the level of 18S rRNA using the formula 2…”

Section: ϫ7mentioning

confidence: 99%

Chondrogenic differentiation of bovine synovium: Bone morphogenetic proteins 2 and 7 and transforming growth factor β1 induce the formation of different types of cartilaginous tissue

Shintani¹,

Hunziker²

2007

Arthritis & Rheumatism

Self Cite

102

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Objective. To compare the potential of bone morphogenetic proteins 2 and 7 (BMP-2 and BMP-7) and transforming growth factor ␤1 (TGF␤1) to effect the chondrogenic differentiation of synovial explants by analyzing the histologic, biochemical, and gene expression characteristics of the cartilaginous tissues formed.Methods. Synovial explants derived from the metacarpal joints of calves were cultured in agarose. Initially, BMP-2 was used to evaluate the chondrogenic potential of the synovial explants under different culturing conditions. Under appropriate conditions, the chondrogenic effects of BMP-2, BMP-7, and TGF␤1 were then compared. The differentiated tissue was characterized histologically, histomorphometrically, immunohistochemically, biochemically, and at the gene expression level.Results. BMP-2 induced the chondrogenic differentiation of synovial explants in a dose-and timedependent manner under serum-and dexamethasonefree conditions. The expression levels of cartilagerelated genes increased in a time-dependent manner. BMP-7 was more potent than BMP-2 in inducing chondrogenesis, but the properties of the differentiated tissue were similar in each case. The type of cartilaginous tissue formed under the influence of TGF␤1 differed in terms of both cell phenotype and gene expression profiles. Conclusion

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Expression of cartilage‐related genes in bovine synovial tissue

Cited by 26 publications

References 29 publications

Transient hypoxia improves matrix properties in tissue engineered cartilage

Transient hypoxia improves matrix properties in tissue engineered cartilage

Differential effects of dexamethasone on the chondrogenesis of mesenchymal stromal cells: Influence of microenvironment, tissue origin and growth factor

Chondrogenic differentiation of bovine synovium: Bone morphogenetic proteins 2 and 7 and transforming growth factor β1 induce the formation of different types of cartilaginous tissue

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