Cytokine profiles in the joint depend on pathology,but are different between synovial fluid,cartilage tissue and cultured chondrocytes

Introduction

This study aimed to evaluate whether profiles of several soluble mediators in synovial fluid and cartilage tissue are pathology-dependent and how their production is related to in vitro tissue formation by chondrocytes from diseased and healthy tissue.

Methods

Samples were obtained from donors without joint pathology (n = 39), with focal defects (n = 65) and osteoarthritis (n = 61). A multiplex bead assay (Luminex) was performed measuring up to 21 cytokines: Interleukin (IL)-1α, IL-1β, IL-1RA, IL-4, IL-6, IL-6Rα, IL-7, IL-8, IL-10, IL-13, tumor necrosis factor (TNF)α, Interferon (IFN)γ, oncostatin M (OSM), leukemia inhibitory factor (LIF), adiponectin, leptin, monocyte chemotactic factor (MCP)1, RANTES, basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), vascular growth factor (VEGF).

Results

In synovial fluid of patients with cartilage pathology, IL-6, IL-13, IFNγ and OSM levels were higher than in donors without joint pathology (P ≤0.001). IL-13, IFNγ and OSM were also different between donors with cartilage defects and OA (P <0.05). In cartilage tissue from debrided defects, VEGF was higher than in non-pathological or osteoarthritic joints (P ≤0.001). IL-1α, IL-6, TNFα and OSM concentrations (in ng/ml) were markedly higher in cartilage tissue than in synovial fluid (P <0.01). Culture of chondrocytes generally led to a massive induction of most cytokines (P <0.001). Although the release of inflammatory cytokines was also here dependent on the pathological condition (P <0.001) the actual profiles were different from tissue or synovial fluid and between non-expanded and expanded chondrocytes. Cartilage formation was lower by healthy unexpanded chondrocytes than by osteoarthritic or defect chondrocytes.

Conclusions

Several pro-inflammatory, pro-angiogenic and pro-repair cytokines were elevated in joints with symptomatic cartilage defects and/or osteoarthritis, although different cytokines were elevated in synovial fluid compared to tissue or cells. Hence a clear molecular profile was evident dependent on disease status of the joint, which however changed in composition depending on the biological sample analysed. These alterations did not affect in vitro tissue formation with these chondrocytes, as this was at least as effective or even better compared to healthy chondrocytes.     

Sensitive fluorescence in situ hybridization signal detection in maize using directly labeled probes produced by high concentration DNA polymerase nick translation

The signal produced by fluorescence in situ hybridization (FISH) often is inconsistent among cells and sensitivity is low. Small DNA targets on the chromatin are difficult to detect. We report here an improved nick translation procedure for Texas red and Alexa Fluor 488 direct labeling of FISH probes. Brighter probes can be obtained by adding excess DNA polymerase I. Using such probes, a 30 kb yeast transgene, and the rp1, rp3 and zein multigene clusters were clearly detected.     

Restoring Savanna Using Fire: Impact on the Breeding Bird Community

Restoration of many terrestrial plant communities involves the reintroduction of fire. However, there have been few studies of the effects of fire on the avifauna during the restoration process. To study the effects of oak savanna restoration on avian communities, breeding birds were censused and the vegetation structure documented in seven experimental burn units (8–18 ha) that had experienced different frequencies of controlled burns during the past 31 years (0–26 burns). Data were analyzed with both direct and indirect gradient analyses using multivariate techniques. The results showed that, as savanna restoration proceeded, there was a general decline in predominantly insectivorous species, particularly those that feed in the upper canopy region (leaves and air space), and a general increase in omnivorous species, particularly those that feed on the ground and in the lower canopy. Insectivorous bark gleaners (woodpeckers) also increased during restoration and were correlated with the increase in standing dead trees resulting from the fires. Overall, savanna restoration resulted in increases in the abundance of many open country bird species, including many species that have been declining in central and eastern North America, including red‐headed woodpecker, Baltimore oriole, eastern kingbird, vesper sparrow, field sparrow, lark sparrow, brown thrasher, American goldfinch, and brown‐headed cowbird. The shifts in species and guilds were correlated with changes in burn frequency and the macro vegetation structure—tree and shrub density, leaf area index, and relative proportion of standing dead trees. The findings show that savanna restoration can increase bird diversity and provide important habitat for uncommon or declining bird species. These birds are most likely attracted to one or more of the distinctive habitat features of the restored savanna environments, including scattered mature trees, standing dead trees and snags, and presence of both shrubby and grassland vegetation. The findings also suggest that restoration ecologists and wildlife biologists will need to work together to achieve desired goals, since different types of savanna restoration efforts may produce different effects on the breeding bird community.     

Influence of alpha-linked glucose on jejunal sodium-glucose co-transport activity in ruminants.

Eight steers and 12 lambs were used in a completely randomized experimental design to determine the effect of partial alpha-amylase starch hydrolysate (SH) on small intestinal sodium-dependent glucose transport activity. Starch hydrolysate was delivered ruminally or abomasally to steers (960 g/day) and sheep (144 g/day) for 7 days. On day 7, the steers were rendered unconscious, exsanguinated and eviscerated. A 1-m section of jejunum was collected starting at the duodenojejunal flexure. Sheep were anaesthetized with pentobarbital and the second meter of small intestine (jejunum) was collected. Brush-border membrane vesicles were prepared and sodium-dependent glucose uptake activity was measured using the rapid uptake/filtration technique. Alkaline phosphatase and maltase activity was enriched by 8.2+/-0.5- and 8.4+/-1.2-fold in the vesicle preparation, respectively, and was not different between treatments. Abomasal SH increased (P=0.03) the Na/glucose co-transport approximately two-fold in both cattle (47.2-114.0+/-31.5 pmol/mgxsec) and sheep (77.4-152.0+/-25.7 pmol mg(-1) s(-1)). We conclude that Na/glucose co-transport activity by enterocytes responds to luminal alpha-linked glucose (from abomasal infusion) in ruminants, compared with controls. Intestinal maltase-specific activity does not respond to alpha-linked glucose in cattle, and decreases slightly in sheep.     

Cytokine profile of autologous conditioned serum for treatment of osteoarthritis, <Emphasis Type="Italic">in vitro</Emphasis>effects on cartilage metabolism and intra-articular levels after injection

Introduction  

Intraarticular administration of autologous conditioned serum (ACS) recently demonstrated some clinical effectiveness in treatment of osteoarthritis (OA). The current study aims to evaluate the in vitro effects of ACS on cartilage proteoglycan (PG) metabolism, its composition and the effects on synovial fluid (SF) cytokine levels following intraarticular ACS administration.     

Immunolocalization of MMP-19 in the human intervertebral disc: implications for disc aging and degeneration

Matrix metalloproteinases (MMPs) degrade components of the extracellular matrix of the disc, but the presence of MMP-19 has not been explored. In other tissues, MMP-19 is known to act in proteolysis of the insulin-like growth factor (IGF) binding protein-3, thereby exposing this protein to make it available to influence cell behavior. MMP-19 also has been shown to inhibit capillary-like formation and thus play a role in the avascular nature of the disc. Using immunohistochemistry, normal discs from six subjects aged newborn through 10 years and 20 disc specimens from control donors or surgical patients aged 15-76 (mean age 40.2 years) were examined for immunolocalization of MMP-19; six Thompson grade I discs, five Thompson grade II, eight Thompson grade III, five Thompson grade IV, and one Thompson grade V discs were analyzed. The results indicate that in discs from young subjects, MMP-19 was uniformly localized in the outer annulus. In discs from adult donors and surgical patients, outer and inner annulus cells only occasionally showed MMP-19 localization. The greatest expression of MMP-19 was observed in young discs, and little expression was seen in older or degenerating discs. Because MMP-19 has been shown to regulate IGF-mediated proliferation in other tissues, its decline in the aging/degenerating disc may contribute to the age-related decrease in disc cell numbers.