Marcia L. Bliven

Exercise protects against articular cartilage degeneration in the hamster

OBJECTIVE It has been reported that osteoarthritis can occur in hamsters. The present study was undertaken to determine the effects of exercise on the composition of articular cartilage and synovial fluid and on the development of cartilage degeneration in these animals. METHODS Young (2.5-month-old) group-housed hamsters were compared with 5.5-month-old hamsters that had undergone 3 months of daily wheel running exercise (6-12 km/day) or 3 months of sedentary, individually housed living. The condition of the femoral condyles was determined by scanning electron microscopy in 12 exercising hamsters, 12 sedentary hamsters, and 6 of the young controls. The content of proteoglycan, hyaluronic acid, hydroxyproline, and proline in synovial fluid and patellar cartilage was measured. RESULTS By scanning electron microscopy, the femoral articular cartilage was smooth and undulating in young controls and older exercising hamsters. In contrast, the femoral condyles were fibrillated in all 12 of the sedentary hamsters. There was no difference in the patellar cartilage collagen content between the 3 groups, but proteoglycan content and synthesis were lower in the patellar cartilage of the sedentary group. Synovial fluid volume was also decreased in the sedentary group compared with the young controls or the older exercising hamsters. CONCLUSION A sedentary lifestyle in the hamster leads to a lower proteoglycan content in the cartilage and a lower synovial fluid volume. These changes are associated with cartilage fibrillation, pitting, and fissuring. Daily exercise prevents early cartilage degeneration and maintains normal articular cartilage.

DETECTION OF COLLAGENASE-INDUCED DAMAGE OF COLLAGEN BY 9A4, A MONOCLONAL C-TERMINAL NEOEPITOPE ANTIBODY

To determine whether the collagen network is compromised by collagenase during acute inflammation, a monoclonal antibody (9A4) was developed with specificity for the C-terminal neoepitope sequence generated by collagenase-cleavage of type II collagen (Gly-Pro-Pro-Gly-Pro-Gln-Gly-COOH). 9A4 was shown to detect the collagen collagenase-cleavage neoepitope with a K = 1.7 x 10(-7) M (type II) and K = 2 x 10(-6) M (type I). It does not recognize uncleaved native or denatured collagen. Articular cartilage from control animals is unstained by 9A4. During acute inflammation elicited in hamsters by intra-articular LPS, positive staining for the 9A4 neoepitope indicated the collagen was damaged. Wheel running exercise was used to apply stress to control cartilage and cartilage from animals with damaged collagen. After 6 months of running, the cartilage from normal animals was unaffected. By contrast, in the group with damaged collagen, the cartilage was fibrillated in all animals and in half of those, the cartilage failed and bony eburnation resulted.

The pharmacologic regulation of interleukin-1 production: The role of prostaglandins

The role of prostaglandins in the regulation of lipopolysaccharide (LPS)-induced interleukin-1 (IL-1) production by murine C3H/HeN resident peritoneal macrophages was studied. IL-1 production was initially studied in the presence of piroxicam and indomethacin, both inhibitors of prostaglandin biosynthesis. IL-1 was assayed using the IL-1-dependent proliferative response of C3H/HeJ thymocytes. LPS stimulation resulted in 15 to 20 ng/ml of prostaglandin E2 (PGE2) produced in the first hour of culture. IL-1-containing supernatants from drug-treated macrophages at dilutions of up to 1:32 resulted in enhanced thymocyte proliferation compared to control, non-drug-treated cultures and contained less than 2 ng/ml of PGE2. Similar enhancement of proliferation could be obtained by incubating non-drug-treated supernatants with monoclonal anti-PGE2 but not anti-thromboxane B2 (TxB2) antibody. Further dilutions of the drug-treated supernatants gave thymocyte proliferation responses which were indistinguishable from control cultures and, correspondingly, had identical values for IL-1 production. The absence of an effect on IL-1 production was confirmed by quantitation of intracellular IL-1 alpha using goat anti-IL-1 alpha antibody and by quantitation of supernatant IL-1 receptor competition assay. Exogenous PGE2, in the concentration range produced in macrophage supernatants (10-20 ng/ml), directly inhibited IL-1-stimulated thymocyte proliferation. Finally, when macrophages were stimulated with LPS for 24 hr in the presence of added PGE2, thymocyte proliferation was inhibited at the lowest supernatant dilutions, but as the IL-1-containing supernatants were diluted out, the assay curves were indistinguishable from non-PGE2-treated control. Thus, in this system, PGE2 has no effect on IL-1 synthesis, but rather has a direct inhibitory effect on thymocyte proliferation. Nonsteroidal anti-inflammatory drugs are not stimulating IL-1 production but are, in fact, relieving inhibition of the thymocyte IL-1 assay caused by the presence of prostaglandins.

Inhibition of interleukin 1 synthesis by tenidap: A new drug for arthritis

Tenidap is a new antiarthritic drug of novel chemical structure. This study shows the effects of tenidap on the in vitro synthesis of interleukin 1 (IL-1). IL-1 production by murine peritoneal macrophages was induced either by stimulation with lipopolysaccharide (LPS) or by phagocytosis of zymosan. With either stimulus, tenidap inhibited IL-1 production as measured by a quantitative competitive IL-1 receptor binding assay. Approximately 20 ng/mL of IL-1 was produced by 10(6) macrophages in response to LPS and about half that amount was produced in response to zymosan. Fifty percent inhibition of IL-1 production by tenidap was found at 3 microM for both stimuli. Using goat anti-IL-1 alpha and Western blot analysis, the appearance of intracellular 34 kDa pro-IL-1 alpha was inhibited by tenidap down to 3 microM. Tenidap decreased [35S]Met incorporation into cellular protein at 30 microM but not at 10 or 3 microM, indicating selectivity for IL-1 inhibition relative to total protein synthesis. Because tenidap inhibited IL-1 induction by both zymosan and LPS, it must act subsequently to receptor triggering. As the appearance of IL-1 was inhibited both intracellularly and extracellularly, the primary drug effect cannot be on secretion.

The effect of interleukin-1 on adhesion formation. in the rat***

The potential role of interleukin-1 in postoperative adhesion formation was examined. Cecal abrasion gave a consistently higher adhesion score when compared with sham laparotomy, on the basis of adhesion number, density, and vascularity, and so was chosen for use in further studies. The extent of serosal bleeding during cecal abrasion did not affect adhesion scores. Intraperitoneal injection of 10 micrograms murine recombinant interleukin-1 alpha in cecally abraded animals on the day of surgery and on the following 4 days resulted in a significant increase in adhesion scores when compared with those of cecally abraded animals injected with vehicle alone. Adhesions enhanced with murine recombinant interleukin-1 alpha, which were thicker and more vascular, were equivalently enhanced at doses from 10 to 10,000 ng, implying maximal response over that range. Rats not operated on and receiving recombinant interleukin-1 alpha 2 weeks after injury had increased adhesion formation. These results demonstrate that interleukin-1 alpha may be an important short-term mediator of postsurgical adhesion formation.

Anti-arthritic effect of bee venom.

Bee venom, administered subcutaneously, suppressed the development of carrageenan-induced paw edema and adjuvant arthritis in the rat in a dose-related manner. A single dose of bee venom administered subcutaneously the day before or on the day of injection of complte Freunds adjuvant (CFA) effectively suppressed the development of polyarthritis. This suppressive effect decreased progressively as dosing was delayed. Bee venom was found to be most effective when mixed and injected (sub-plantar) together with CFA, the disease-inducing agent. Similarly, antigens such as egg albumin, when incorporated into CFA, and injected into the hind paw, prevented the development of arthritis. These results suggest that at least two mechanisms are involved in the anti-arthritic action of bee venom: (1) alteration of the immune response, probably via antigen competition; and (2) an anti-inflammatory action via corticosteroids or through an as yet undetermined mechanism.

Effects of exercise on synovium and cartilage from normal and inflamed knees

The effect of running activity on normal and inflamed knees was determined by light microscopic (LM) and scanning electron microscopic (SEM) observations on hamster articular cartilage. Animals were split into two groups; one housed in standard cages and one given free access to running wheels. Twenty-one days prior to analysis, half of each group was given an intraarticular injection of lipopolysaccharide (LPS) to cause an inflammation, the other half were uninjected. No remarkable changes were observed by LM in either the control running or nonrunning groups. In contrast, cartilage proteoglycan depletion, and pannus and synovial hyperplasia were equally observed in both groups of LPS-injected animals. SEM observations on the patellae from control animals found them to be free from damage to the articular cartilage. The joints of both the LPS nonrunning and running animals contained synovial hypertrophy with villus projection from the synovial lining. However, only the LPS-injected running hamsters had cartilage fraying over large areas of the articular surface, as well as areas in which the villus projections had been flattened. These results demonstrated that mechanical stress applied to a proteoglycan-depleted cartilage enhances the breakdown of the collagen matrix as judged by fibrillation, and may aggravate the inflammation by crushing the swollen synovial lining where it encroaches on the joint space.

Some factors affecting inhibition and restoration of mobility after induction of an acute arthritis in the hamster

Mobility is impaired during arthritis. In order to study the causes of the mobility impairment, we have examined hamsters with a LPS arthritis in which running is inhibited over a 4–5 day period. Parameters have been examined to determine which correlate with running impairment. Knee diameter, as well as cell infiltration into the surrounding synovial tissue and the accompanying edema, failed to resolve by the time normal mobility was reestablished. Other factors must be primary determinants of mobility.

Interaction of levamisole and mercaptans during thymocyte proliferation induced by concanavalin A

Abstract Levamisole enhances[ 3 H]thymidine uptake of murine thymocytes stimulated by concanavalin A (Con A). The proliferative response of thymocytes to Con A can also be enhanced by addition of mercaptans. Six different mercaptans were examined for this effect; three of them, 2-mercaptoethanol, cysteamine, and l -cysteine, stimulated the Con A response. Addition of levamisole to an optimal stimulatory dose of 2-mercaptoethanol or cysteamine resulted in complete inhibition of cell proliferation. Three other mercaptans, penicillamine, d -cysteine, and glutathione, failed to enhance the Con A response and, in fact, were mildly inhibitory. Levamisole gave only slightly less than normal stimulation in the presence of these mercaptans. In the absence of Con A neither levamisole nor the mercaptans stimulated cell proliferation. Oxidized 2-mercaptoethanol reacted analogously to reduced 2-mercaptoethanol both in the presence and absence of levamisole. We have interpreted these results as suggesting that the effect of levamisole is dependent upon the state of activation of the lymphocyte.

Limitation of activity in an acute model of arthritis: Effect of drug treatment

Abstract.Objective and Design: The limitation of activity and its modification by therapy in an experimental arthritis was studied.¶Subjects: Female hamsters in groups of six per treatment were used.¶Treatment: An acute arthritis was induced by intraarticular injection of 0.1 μg lipopolysaccharide (LPS) in hamsters with free access to running wheels. Tenidap at 100 mg%, and piroxicam and indomethacin at 30 mg% were administered in the hamsters normal diet.¶Methods: Activity was monitored and analysed by computer. Plasma blood levels of drugs were determined by high pressure liquid chromatographic (HPLC) analysis.¶Results: Hamsters normally run 10–15 km/day. That distance was reduced to less than 2 km/day after arthritis induction. Speed of movement, essentially the equivalent of walking time, was reduced 40% by the arthritis. However, the time spent in movement (activity time) was more severely affected by arthritic disease. Therapy gave a modest 1.3-fold increase in speed of movement, but a highly significant 2-fold increase in activity time.¶Conclusions: The effects of arthritis on activity in this animal model suggest that time spent in movement (activity time) should be considered as an outcome measure in clinical studies. These observations may also help explain why the modest disease improvements obtained with cyclooxygenase inhibition are valued. From a patient perspective, a doubling of activity time is a highly significant improvement in quality-of-life.