Patrick Netter

Obesity and osteoarthritis: more complex than predicted!

Dysregulation of lipid homeostasis is one of the mechanisms leading to osteoarthritis Osteoarthritis is usually considered to be a joint disorder the central pathological feature of which is cartilage destruction. However, this concept has evolved, and today osteoarthritis is generally regarded as a disease that may affect the whole joint (bone, muscles, ligaments and synovium). Although the aetiology of osteoarthritis is not established, the main risk factors are well known and commonly include mechanical, biochemical and genetic factors. Of these risk factors, obesity is beyond doubt considered a prominent one. The overload effect on joint cartilage may explain part of the increased risk of osteoarthritis, at least for osteoarthritis of the knee, in overweight people. A recent discovery in the discipline of cartilage biology is the presence of mechanoreceptors at the surface of chondrocytes, which are sensitive to pressure and link extracellular environment to intracellular signalling cascades. Three types of mechanoreceptors have been described on chondrocytes: the stretch-activated channels, the α-5β1 integrin and CD44. Compression and stretch stimulate integrins and stretch-activated channels leading to the activation of signalling pathways (mitogen-activated protein kinase, NF-κB), as well as the release of second messengers (calcium, Inositol triphosphate and Adenosine monophosphate cyclic). 1 After mechanoreceptor activation, cytokines, growth factors and metalloproteinases may be expressed, and mediators such as prostaglandins or nitric oxide may be produced.2 As experimental studies have shown that under specific conditions overload may trigger both inhibition of matrix synthesis and cartilage degradation, we can speculate that obesity may induce cartilage damage through activation of these mechanoreceptors. In the same manner, the mechanoreceptors expressed on osteoblasts3,4 may also be involved in the impaired response of chondrocytes to the obesity-induced overload. Even if it is usually accepted that mechanical loading contributes to joint cartilage destruction in overweight patients, recent advances …

Mutations in ANKH Cause Chondrocalcinosis

Chondrocalcinosis (CC) is a common cause of joint pain and arthritis that is caused by the deposition of calcium-containing crystals within articular cartilage. Although most cases are sporadic, rare familial forms have been linked to human chromosomes 8 (CCAL1) or 5p (CCAL2) (Baldwin et al. 1995; Hughes et al. 1995; Andrew et al. 1999). Here, we show that two previously described families with CCAL2 have mutations in the human homolog of the mouse progressive ankylosis gene (ANKH). One of the human mutations results in the substitution of a highly conserved amino acid residue within a predicted transmembrane segment. The other creates a new ATG start site that adds four additional residues to the ANKH protein. Both mutations segregate completely with disease status and are not found in control subjects. In addition, 1 of 95 U.K. patients with sporadic CC showed a deletion of a single codon in the ANKH gene. The same change was found in a sister who had bilateral knee replacement for osteoarthritis. Each of the three human mutations was reconstructed in a full-length ANK expression construct previously shown to regulate pyrophosphate levels in cultured cells in vitro. All three of the human mutations showed significantly more activity than a previously described nonsense mutation that causes severe hydroxyapatite mineral deposition and widespread joint ankylosis in mice. These results suggest that small sequence changes in ANKH are one cause of CC and joint disease in humans. Increased ANK activity may explain the different types of crystals commonly deposited in human CCAL2 families and mutant mice and may provide a useful pharmacological target for treating some forms of human CC.

Cyclo-oxygenase isoenzymes. How recent findings affect thinking about nonsteroidal anti-inflammatory drugs.

SummaryThe discovery of at least 2 cyclo-oxygenase (COX) isoenzymes, referred to as COX-1 and COX-2, has updated our knowledge of nonsteroidal anti-inflammatory drugs (NSAIDs). This has lead investigators to reconsider what can be awaited from this class of drugs. The 2 COX isoenzymes share structural and enzymatic similarities, but are specifically regulated at the molecular level and may be distinguished apart in their functions, although some physiological overlap between them does occur.The major goal in developing selective COX inhibitors is to improve NSAID tolerability. Classic NSAIDs preferentially inhibit COX-1 in vitro, but it appears hazardous to judge their gastrointestinal (GI) safety profile from these data. New compounds with a high selectivity for COX-2, especially those that are non-acidic, may be better tolerated in the GI tract.While these compounds also might have a potential use in various diseases such as colorectal cancer and neurodegenerative diseases of the Alzheimer type, the possible appearance of adverse effects, perhaps renally-related, must be taken into consideration. Finally, well-designed large clinical trials are required to adequately estimate both the promising therapeutic advantages that may be offered by highly selective NSAIDs, and the potential drawbacks that may be inherent with prolonged COX-2 inhibition.

Interleukin-1β down-regulates the expression of glucuronosyltransferase I, a key enzyme priming glycosaminoglycan biosynthesis influence of glucosamine on interleukin-1β-mediated effects in rat chondrocytes

OBJECTIVE To assess the variations of galactose-beta-1,3-glucuronosyltransferase I (GlcAT-I) expression related to the decrease in proteoglycan synthesis mediated by interleukin-1beta (IL-1beta) in rat chondrocytes, and to evaluate the influence of glucosamine on the effects elicited by this proinflammatory cytokine. METHODS Rat articular chondrocytes in primary monolayer cultures or encapsulated into alginate beads were treated with recombinant IL-1beta in the absence or presence (1.0-4.5 gm/liter) of glucosamine. Variations of GlcAT-I and expression of stromelysin 1 (matrix metalloproteinase 3 [MMP-3]) messenger RNA (mRNA) were evaluated by quantitative multistandard reverse transcriptase-polymerase chain reaction. In vitro enzymatic activity of GlcAT-I was measured by thin-layer chromatography, with radiolabeled UDP-glucuronic acid and a digalactoside derivative as substrates. Proteoglycan synthesis was determined by ex vivo incorporation of Na2-35SO4. Nitric oxide synthase and cyclooxygenase activities were monitored by the evaluation of nitrite (NO2-) and prostaglandin E2 (PGE2) produced in the culture medium, respectively. RESULTS IL-1beta treatment resulted in a marked inhibition of GlcAT-I mRNA expression and in vitro catalytic activity, together with a decrease in proteoglycan synthesis. In addition, glucosamine was able to prevent, in a dose-dependent manner, the inhibitory effects of IL-1beta. In the same way, the amino sugar reduced NO2- and PGE2 production induced by IL-1beta. Finally, the up-regulation of stromelysin 1 (MMP-3) mRNA expression by IL-1beta was fully prevented by glucosamine. CONCLUSION The results of this study suggest that the deleterious effect of IL-1beta on the anabolism of proteoglycan could involve the repression of GlcAT-I, a key enzyme in the biosynthesis of glycosaminoglycan. Glucosamine was highly effective in preventing these IL-1beta-mediated suppressive effects. The amino sugar also prevented the production of inflammatory mediators induced by the cytokine. This action could account for a possible beneficial effect of glucosamine on osteoarthritic articular cartilage.

Glucosamine modulates IL-1-induced activation of rat chondrocytes at a receptor level, and by inhibiting the NF-κB pathway

We recently reported that glucosamine reversed the decrease in proteoglycan synthesis and in UDP‐glucuronosyltransferase I mRNA expression induced by interleukin‐1β (IL‐1β) [Arthritis Rheum. 44 (2001) 351–360]. In the present work, we show that glucosamine does not exert the same effects when chondrocytes were stimulated with reactive oxygen species (ROS). In order to better understand its mechanism of action, we determined if glucosamine could prevent the binding of IL‐1β to its cellular receptors or could interfere with its signaling pathway at a post‐receptor level. Addition of glucosamine to rat chondrocytes treated with IL‐1β or with ROS decreased the activation of the nuclear factor κB, but not the activator protein‐1. After treatment with IL‐1β, glucosamine increased the expression of mRNA encoding the type II IL‐1β receptor. These results emphasize the potential role of two regulating proteins of the IL‐1β signaling pathway that could account for the beneficial effect of glucosamine in osteoarthritis.

Cytocompatibility of Ti-6Al-4V and Ti-5Al-2.5Fe alloys according to three surface treatments, using human fibroblasts and osteoblasts.

Titanium alloys are well known for their superior mechanical properties as well as for their good biocompatibility, making them desirable as surgical implant materials. However, these alloys have been proven to behave poorly in friction since wear particles were often detected in tissues and organs associated with titanium implants. In this paper, three surface treatments were investigated in order to improve the wear resistance and the hardness of Ti-6Al-4V and Ti-5Al-2.5Fe: (a) glow discharge nitrogen implantation (10(17) atoms cm-2), (b) plasma nitriding by plasma diffusion treatment (PDT) and (c) deposition of TiN layer by plasma-assisted chemical vapour deposition (PACVD) additionally to PDT. Surface characterization after the different treatments showed considerable improvement in surface hardness, especially after the two nitriding processes. Moreover, the good corrosion resistance of untreated alloys was maintained. A cell culture model using human cells was chosen to study the effect of such treatments on the cytocompatibility of these materials. The results showed that Ti-5Al-2.5Fe alloy was as cytocompatible as the Ti-6Al-4V alloy and the same surface treatment led to identical biological consequences on both alloys. Nitrogen implantation did not modify at all the cellular behaviour observed on untreated samples. After the two nitriding treatments, cell proliferation and viability appeared to be significantly reduced and the scanning electron microscopy study revealed somewhat irregular surface states. However, osteoblast phenotype expression and protein synthesis capacity were not affected. PDT and PACVD may be interesting alternatives to the physical vapour deposition technique.

Evaluation of acoustical parameter sensitivity to age-related and osteoarthritic changes in articular cartilage using 50-MHZ ultrasound

The current study reports the sensitivity of acoustical parameters estimated at high frequency to the osteoarthritic morphological and structural changes in patellar cartilage in rat knees. Osteoarthritis (OA) was induced by a single intra-articular injection of mono-iodo-acetic acid in right knees. OA patellas and their contralateral controls were excised at regular intervals after injection and were examined in vitro with a scanning acoustical microscope operating with a poly(vinylidene di-fluoride) (PVDF) 80-MHz focused transducer. Cartilage thickness was estimated using B-scan images. The quantitative analysis of the radiofrequency signal backscattered by the cartilage was performed using integrated reflection coefficient (IRC) and apparent integrated backscatter (AIB), which were estimated in the 20-60-MHz frequency range. One week after injection, a cartilage thickness decrease was detected (-6%, on average) that preceded the significant hypertrophy (20.1%) that occurred 2 weeks after injection and could be due to tissue repair. From 1 week to 3 weeks after injection, the IRC of OA patellas was significantly lower than that of control patellas. The IRC difference increased with time from -3.3 +/- 2.4 dB at 1 week to -8.4 +/- 1.7 dB at 3 weeks. An AIB decrease was observed with time for both OA and control patellas (-2.9 to -4.2 dB per week). An AIB difference between OA and control patellas was detected from 1 week to 3 weeks after injection. This difference decreased with time. IRC variation reflects a change in acoustical impedance of the superficial layer of the cartilage and could be linked to a change in constituent content and/or to a disruption of fibers of the collagen network that led to the fibrillation of the cartilage surface. AIB variation reflects a change in shape, size and/or density of the scatterers and could be related to changes in the constituent content and in the organization of the matrix in the internal layer of the cartilage. IRC and AIB could provide information about the structural modifications of the cartilage due to osteoarthritis or to cartilage maturation.

Assessment of articular cartilage and subchondral bone : Subtle and progressive changes in experimental osteoarthritis using 50 MHz echography in vitro

The main objectives of this work were to demonstrate the potential of 50 MHz echography for assessing initial and progressive morphological and structural changes of articular cartilage and bone developed in an experimental model of osteoarthritis (OA). Degenerative lesions were induced in rat knees by the unilateral intra‐articular injection of a 3 mg dose of mono‐iodo‐acetic acid. To assess the lesion progression, the animals (n = 30) were sacrificed at different time intervals up to 8 weeks after the injection. Three‐dimensional echographic data were acquired in vitro on patellar cartilage and bone at various stages of the remodeling process using a scanning ultrasound microscope. Changes involving the OA cartilage characteristics are discussed relative to those of the contralateral control joint which received a placebo. Images of control cartilage showed a smooth hyperechoic articular surface and an echoic matrix. The cartilage thickness was 266 ± 44 μm (mean ± SD) in the central region of the tissue. The precision of ultrasonic thickness measurements was better than 1.3%. First changes in cartilage internal structure and subchondral bone appeared on ultrasound images 3 days after the injection and were even more evident by day 7. They resulted in a slight thinning of the cartilage, a 30% increase of its internal structure echogenicity, and the appearance of echoic zones in subchondral bone. Histologic findings confirmed chondrocyte depletion and degeneration, decrease of matrix proteoglycans, and fibrovascular connective tissue proliferation at the subchondral plate. Progressive and severe lesions at both bone and cartilage surface and internal structure were assessed and correlated to histologic features. These results show that high resolution echography is sensitive to subtle and progressive osteochondral remodeling. This technique has the potential to be used for intra‐articular quantitative imaging and assessment of early changes in bone and cartilage structure associated with natural human disease.

Evaluation of the effect of three surface treatments on the biocompatibility of 316L stainless steel using human differentiated cells

AISI 316L stainless steel (SS) is widely used in orthopaedic implantology, although biological complications may result from its insufficient mechanical and tribological properties. In order to improve the wear and corrosion resistance as well as the hardness of 316L SS, three surface treatments, derived from those applied in mechanical engineering industries, were investigated: (1) glow discharge nitrogen implantation, (2) carbon-doped stainless steel coating sputtering and (3) low temperature plasma nitriding. Surface characterization according to the different heat treatments showed that corrosion and wear resistance were strongly improved, especially by ion implantation or carbon-doped SS coating sputtering. In the same way, microhardness was significantly increased after the three treatments. The effect of such treatments on the biocompatibility of 316L SS was studied with human osteoblast and fibroblast cultures. Basic and specific features of the cells showed that ion-implanted and carbon-doped stainless steels were biocompatible, whereas dramatic cellular reactions were noted when contacted with nitrided stainless steel. A hypothesis is given to explain this observation but further experiments are needed to optimize the nitriding process. Nitrogen implantation and carbon-doped layer deposition could be efficient means for improving the physical properties of stainless steel without affecting its biocompatibility. Such surface treatments may have relevance for increasing the life time of 316L biomedical devices.

15-Deoxy-Δ12,14-PGJ2, but not troglitazone, modulates IL-1β effects in human chondrocytes by inhibiting NF-κB and AP-1 activation pathways

The activation of peroxisome proliferator‐activated receptor γ (PPARγ) has been shown to inhibit the production and the effects of proinflammatory cytokines. Since interleukin‐1β (IL‐1β) directly mediates cartilage degradation in osteoarthritis, we investigated the capability of PPARγ ligands to modulate IL‐1β effects on human chondrocytes. RT‐PCR and Western blot analysis revealed that PPARγ expression was decreased by IL‐1β. 15‐Deoxy‐Δ12,14‐prostaglandin J2 (15d‐PGJ2), in contrast to troglitazone, was highly potent to counteract IL‐1β‐induced cyclooxygenase‐2 and inductible nitric oxide synthase expression, NO production and the decrease in proteoglycan synthesis. Western blot and gel‐shift analyses demonstrated that 15d‐PGJ2 inhibited NF‐κB activation, while troglitazone was ineffective. Although 15d‐PGJ2 attenuated activator protein‐1 binding on the DNA, it potentiated c‐jun migration in the nucleus. The absence or the low effect of troglitazone suggests that 15d‐PGJ2 action in human chondrocytes is mainly PPARγ‐independent.