This study examined effects of intermittent hydrostatic pressure (IHP) and a chondrogenic growth factor, bone morphogenetic protein-2 (BMP-2), on anabolic, catabolic, and other metabolic markers in human osteoarthritic (OA) chondrocytes in vitro.
Articular chondrocytes, isolated from femoral OA cartilage and maintained in high-density monolayer culture, were examined for effects of BMP-2 and IHP on gene expression of matrix-associated proteins (aggrecan, type II collagen, and SOX9) and catabolic matrix metalloproteinases (MMP-2 and MMP-3) and culture medium levels of the metabolic markers MMP-2, nitric oxide (NO), and glycosaminoglycan (GAG). The results were analyzed using a mixed linear regression model to investigate the effects of load and growth factor concentration.
IHP and BMP-2 modulated OA chondrocyte metabolism in accordance with growth factor concentration independently, without evidence of synergism or antagonism. Each type of stimulus acted independently on anabolic matrix gene expression. Type II collagen and SOX9 gene expression were stimulated by both IHP and BMP-2 whereas aggrecan was increased only by BMP-2. IHP exhibited a trend to decrease MMP-2 gene expression as a catabolic marker whereas BMP-2 did not. NO production was increased by addition of BMP-2 and IHP exhibited a trend for increased levels. GAG production was increased by BMP-2.
This study confirmed the hypothesis that human OA chondrocytes respond to a specific type of mechanical load, IHP, through enhanced articular cartilage macromolecule gene expression and that IHP, in combination with a chondrogenic growth factor BMP-2, additively enhanced matrix gene expression without interactive effects.
"Conversely, insufficient mechanical stimulus, such as that due to joint immobilization, has also been associated with cartilage destruction . On the other hand, moderate (physiological) mechanical stimulus has been confirmed not only to promote articular cartilage anabolism  but also to inhibit catabolism [5,6]. "
[Show abstract][Hide abstract] ABSTRACT: Although the individual effects of hyperbaric oxygen (HBO) and low-intensity pulsed ultrasound (LIPUS) on osteoarthritic (OA) chondrocytes have been reported, the effects of HBO combined with LIPUS treatment are unknown.
OA chondrocytes were obtained from patients undergoing knee replacement surgery. RNA was isolated for real-time polymerase chain reaction (PCR) analysis of inducible nitric oxide synthase (iNOS), type-II collagen, and aggrecan gene expression. The protein levels of MMP-3 and TIMP-1 were quantified by enzyme-linked immunosorbent assay (ELISA) after LIPUS or HBO treatment. The data are given as mean +/- standard deviation (SD) of the results from three independent experiments. A p value less than 0.05 was defined as statistically significant.
Our data suggested that ultrasound and HBO treatment increased cell bioactivity of OA chondrocytes. Real-time PCR analysis showed that HBO treatment increased the mRNA of type-II collagen, aggrecan, and TIMP-1 but suppressed the iNOS expression of OA chondrocytes. LIPUS treatment increased the type-II collagen and iNOS expression of OA chondrocytes. ELISA data showed that HBO or LIPUS treatment increased TIMP-1 production of OA chondrocyte. MMP-3 production was suppressed by HBO treatment. HBO combined with LIPUS treatments resulted in additive effect in TIMP-1 production and compensatory effect in iNOS expression.
HBO combined with LIPUS treatment-induced increase of the anabolic factor (TIMP-1)/catabolic factor (MMP-3) ratio may provide an additive therapeutic approach to slow the course of OA degeneration.
Journal of Orthopaedic Surgery and Research 02/2014; 9(1):5. DOI:10.1186/1749-799X-9-5 · 1.39 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This study examined the effects of intermittent hydrostatic pressure (IHP) and transforming growth factor-beta 3 on chondrogenesis of adult human mesenchymal stem cells (hMSCs) in vitro. Chondrogenic gene expression was determined by quantifying mRNA signal levels for SOX9, a transcription factor critical for cartilage development and the cartilage matrix proteins, aggrecan and type II collagen. Extracellular matrix production was determined by weight and histology. IHP was applied to hMSCs in pellet culture at a level of 10 MPa and a frequency of 1 Hz for 4 h per day for periods of 3, 7, and 14 days. hMSCs responded to addition of TGF-beta 3 (10 ng/mL) with a greater than 10-fold increase (p < 0.01) in mRNA levels for each, SOX9, type II collagen, and aggrecan during a 14-day culture period. Applying IHP in the presence of TGF-beta 3 further increased the mRNA levels for these proteins by 1.9-, 3.3-, and 1.6-fold, respectively, by day 14. Chondrogenic mRNA levels were increased with just exposure to IHP. Extracellular matrix deposition of type II collagen and aggrecan increased in the pellets as a function of treatment conditions and time of culture. This study demonstrated adjunctive effects of IHP on TGF-beta 3-induced chondrogenesis and suggests that mechanical loading can facilitate articular cartilage tissue engineering.
[Show abstract][Hide abstract] ABSTRACT: This study was designed to evaluate the combined effects of intermittent hydrostatic pressure (IHP) and TGF-β1 or TNF-α on proteoglycan4 (PRG4) expression in rat temporomandibular synovial fibroblasts (SFs).
Rat SFs were isolated and expanded in monolayer cultures and subjected to IHP in the presence of TGF-β1 or TNF-α. Quantitative real-time RT-PCR was applied to analyze the PRG4 expression levels. Enzyme-linked immunosorbent assay was also used for the quantification of PRG4 accumulation in the culture medium while immunofluorescence staining was used to detect intracellular PRG4 protein expression.
The combination of IHP and TGF-β1 induced greater PRG4 expression than either stimulus alone. In contrast, TNF-α inhibited PRG4 expression, and this was partially alleviated by IHP.
Our study demonstrates a beneficial role of IHP, which can be used successfully in combination with TGF-β1 to enhance PRG4 production, and can partially counteract TNF-α-induced PRG4 inhibition in isolated rat SFs.
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.