Osteoarthritis and Cartilage

Glucosamine modulates chondrocyte proliferation, matrix synthesis, and gene expression

S. Varghese Ph.D., P. Theprungsirikul B.S., S. Sahani B.S., N. Hwang B.S., K.J. Yarema Ph.D. and J.H. Elisseeff Ph.D.

Department of Biomedical Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21218, USA

Summary:

The therapies that currently exist for improving general joint health and for treating the symptoms of the degenerative disease osteoarthritis (OA) include the use of anti-inflammatory and pain relieving drugs. However, it has also been hypothesized that the dietary supplementation of "chondro-protective" components such as glucosamine (GlcN) and chondrotin sulfate (CS) could be used to treat OA by stimulating cartilage regeneration. In this present study, Varghese et al. sought to identify what optimal concentration of GlcN (if one such concentration did exist) had a significant effect on chondrocyte proliferation, matrix production, a gene expression in both monolayer (2D) and three-dimensional (3D) culture conditions. They attempted to accomplish this task by evaluating the effects of various amounts of GlcN on primary bovine articular chondrocytes (BAC) and by demonstrating how the responses of the cells differ between the two culture conditions. Reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR were used to examine the impact of GlcN on gene expression in both culture conditions, specifically aggrecans and collagen type II. Yet in order to accurately evaluate the effects of GlcN on cartilage matrix production in the 3D culture conditions, it was also necessary to perform histology, immunofluorescent staining and biochemical analyses.

Noticeable differences in both cellular morphology and the microenvironment surrounding the cells were observed between the 2D and 3D culture conditions. The authors suggest that these discrete differences in GlcN tolerance may be attributed to the various niches the two conditions provide to the cells. In particular, the 3D hydrogels provided a structural environment that was more similar to native articular cartilage. For instance, incubation of 3D constructs with 2mM GlcN-medium resulted in the highest cartilage specific matrix production, GAG and collagen type II. However, as in 2D culture conditions, a decrease in cell proliferation and adverse effects on chondrocyte matrix production with increasing GlcN concentrations was still observed in the 3D culture conditions. GlcN was also found to upregulate TGF-beta1 mRNA levels in a dose-dependent manner in both the 2D and 3D culture conditions. Moreover, based on their findings, the authors emphasized that chondrocytes needed to be given sufficient time to adhere onto the culture dish (i.e., in monolayer culture conditions) in order to even survive at high GlcN concentrations. TGF-beta1 is known to stimulate the collagen production of articular chondrocytes and is considered to regulate cartilage fracture repair by extracellular matrix production.

Significance:

Various studies that have investigated the effect of agents such as glucosamine on cartilage regeneration report mixed results; some promising, and others not, making this subject highly controversial. The present study, however, provides additional evidence that prolonged exposure of primary chondrocytes to optimal GlcN concentrations increases matrix production. The authors also indicate that this may be the first study of its kind that demonstrates GlcN mediated up-regulation of TGF-beta1 in chondrocytes. The authors propose that the increased production of extracellular matrix can possibly be explained by the GlcN mediated up-regulation of TGF-beta1. By promoting the expression of TGF-beta1, the authors hypothesize that optimal amounts of GlcN can preserve cartilage tissue and promote its repair upon damage. Hence, this study paves the way for the development of better clinical strategies for cartilage repair involving localized and controlled release of GlcN.

Cardiac Tissue Engineering: regeneration of the wounded heart

This paper summarizes recent research on the regeneration of hearts damaged by myocardial infarction. Due to a lack of heart donors and complications associated with immune suppressive treatments, new solutions are needed to regenerate hearts damaged by myocardial infarction. One method for regenerating functional myocardial tissue is cell grafting by syringe injection directly in the ventricular wall or coronary vessels. So far adult stem cells such as bone marrow stem cells have been studied by several teams who claim these cells can develop into cardiomyocytes. Most studies support the notion that cell engraftment in animal models of myocardial infarction can improve contractile function (although the mechanism behind this functional improvement is not clear). The paper goes on to say that cell engraftment is very inefficient, and that 90% of the cell suspension injected is lost and does not engraft. Therefore more effort is going into the development of tissue engineering strategies using biomatrices to successfully engraft new cells into the myocardium. Researchers quickly realized the material had to be elastic to compensate for the heart’s changing shape. Currently several groups are working with scaffold materials composed of natural polymers such as collagen (a major constituent of the cardiac ECM), which have shown promising results. One of the most recent approaches involves the use of materials to create electrically communicating 3D cardiac tissue layers. In order to do this, cells are adhered on tissue culture plates previously coated with poly(N-isopropylacrylamide) (PIPAAm), a temperature sensitive polymer. At high temperatures the polymer is hydrophobic, enabling cell adhesion, but at lower temperatures it is hydrophilic, and becomes inappropriate for cell adhesion due to rapid hydration and swelling of the polymer. Eventually, the matrix can be grown into as many as four layers of synchronously beating cardiomyocytes, which can then be implanted into rats with induced myocardial infarction. The implanted rats show improved myocardial contractility and increased vascularization of the area.