Effects of testosterone, estrogen and progesterone on TNF-α mediated cellular damage in rat arthritic synovial fibroblasts.

Sexual dimorphism is a well-established phenomenon in rheumatoid arthritis, with women exhibiting higher disease severity. Understanding the role of sex hormones using in vivo animal models is limited due to the systemic effects as well as the difficulty in exploring different dose combinations of the hormones simultaneously. However, cell culture systems pose ideal systems for exploring different combinations and concentrations of the hormones simultaneously. In this study, the procedure for isolation of arthritic fibroblasts was standardized using a combination of collagenase and trypsin based on maximal yield and viability after employing different enzymatic disaggregation procedures. The cultured synovial fibroblasts from arthritic rats did not differ significantly from normal rat fibroblasts in terms of proliferation or secretion of inflammatory mediators. Stimulation of fibroblasts with TNF-α was standardized and TNF-α stimulated rat arthritic synovial fibroblasts exhibited an ideal in vitro system for screening antiinflammatory molecules. The effects of physiological and pharmacological concentrations of testosterone, estrogen and progesterone were studied on TNF-α induced cellular damage in rat arthritic synovial fibroblasts. The results showed that estrogen and testosterone exerted antiinflammatory effects on rat arthritic synovial fibroblasts at physiological and pharmacological concentrations. However, there was no significant difference in the effects between physiological and pharmacological concentrations. Progesterone independently did not show any protective effects. In combination with physiological concentrations of estrogen, progesterone abrogated estrogen's protective effect but it exhibited protection in combination with pharmacological concentrations of estrogen.

Low frequency pulsed electromagnetic field--a viable alternative therapy for arthritis.

Arthritis refers to more than 100 disorders of the musculoskeletal system. The existing pharmacological interventions for arthritis offer only symptomatic relief and they are not definitive and curative. Magnetic healing has been known from antiquity and it is evolved to the present times with the advent of electromagnetism. The original basis for the trial of this form of therapy is the interaction between the biological systems with the natural magnetic fields. Optimization of the physical window comprising the electromagnetic field generator and signal properties (frequency, intensity, duration, waveform) with the biological window, inclusive of the experimental model, age and stimulus has helped in achieving consistent beneficial results. Low frequency pulsed electromagnetic field (PEMF) can provide noninvasive, safe and easy to apply method to treat pain, inflammation and dysfunctions associated with rheumatoid arthritis (RA) and osteoarthritis (OA) and PEMF has a long term record of safety. This review focusses on the therapeutic application of PEMF in the treatment of these forms of arthritis. The analysis of various studies (animal models of arthritis, cell culture systems and clinical trials) reporting the use of PEMF for arthritis cure has conclusively shown that PEMF not only alleviates the pain in the arthritis condition but it also affords chondroprotection, exerts antiinflammatory action and helps in bone remodeling and this could be developed as a viable alternative for arthritis therapy.

Comparative studies on the interplay of testosterone, estrogen and progesterone in collagen induced arthritis in rats.

Rheumatoid arthritis (RA) is a sexually dimorphic autoimmune disorder exhibiting a higher disease prevalence and severity among females. This study was carried out to understand the role of the major sex hormones viz., testosterone, estrogen and progesterone on the severity in arthritis. The interplay of the sex hormones was studied in a rat model of collagen induced arthritis (CIA). The parameters used for analyzing the disease severity included paw volume, radiology, histopathology of joint, markers for bone turnover, cytokine profile, levels of pain mediator (prostaglandin E(2)) and immune response to type II collagen. Arthritis induction to castrated and ovariectomised rats resulted in enhanced inflammation thereby indicating the importance of sex hormones. Treatment with physiological doses of dihydrotestosterone and estrogen attenuated the inflammation, with estrogen exhibiting higher potency. Progesterone was not shown to have any significance in disease modification; however, when progesterone was administered in combination with estrogen, the protective effects of estrogen were noticed to decrease.

Estrogen and testosterone attenuate extracellular matrix loss in collagen-induced arthritis in rats.

Rheumatoid arthritis (RA) is a sexually dimorphic, autoimmune inflammatory disorder affecting the joints. Joint disability in RA results primarily from loss of matrix components (collagen and glycosoaminoglycan) in the cartilage and synovium. This study was carried out to understand the effect of physiological levels of testosterone, estrogen, and progesterone on oxidative stress-induced changes in matrix composition in rat synovium in arthritis. Arthritis induction in castrated and ovariectomized rats resulted in enhanced oxidative stress and this was assessed by lipid peroxidation levels and depletion of antioxidants. This, in turn, led to significantly (p < 0.01) increased levels of TNF-alpha and matrix metalloproteinase-2 (MMP-2), subsequently resulting in loss of collagen, elastin, and glycosoaminoglycan (GAG) and disorganization of reticulin as evidenced by biochemical quantitation and also by staining for collagen, reticulin, and elastin. Treatment with physiological doses of dihydrotestosterone (25 mg topically) and estrogen (5 microg/0.1 ml subcutaneously) restored the antioxidant levels significantly (p < 0.05) and reduced the levels of TNF-alpha and MMP-2, with estrogen exhibiting a higher potency. This, in turn, attenuated the damage to reticulin organization as well as the loss of collagen and GAG in the articular tissues. However, elastin loss could not be attenuated by either treatment. Progesterone (2 mg/0.1 ml subcutaneously) was not shown to have any significance in disease modification, and on the contrary, it inhibited the protective effects of estrogen. However, progesterone contributed to increased collagen levels in the tissues.

Low frequency and low intensity pulsed electromagnetic field exerts its antiinflammatory effect through restoration of plasma membrane calcium ATPase activity.

Rheumatoid arthritis (RA) is a chronic inflammatory disorder affecting 1% of the population worldwide. Pulsed electromagnetic field (PEMF) has a number of well-documented physiological effects on cells and tissues including antiinflammatory effect. This study aims to explore the antiinflammatory effect of PEMF and its possible mechanism of action in amelioration of adjuvant induced arthritis (AIA). Arthritis was induced by a single intradermal injection of heat killed Mycobacterium tuberculosis at a concentration of 500 microg in 0.1 ml of paraffin oil into the right hind paw of rats. The arthritic animals showed a biphasic response regarding changes in the paw edema volume. During the chronic phase of the disease, arthritic animals showed an elevated level of lipid peroxides and depletion of antioxidant enzymes with significant radiological and histological changes. Besides, plasma membrane Ca(2+) ATPase (PMCA) activity was inhibited while intracellular Ca(2+) level as well as prostaglandin E(2) levels was noticed to be elevated in blood lymphocytes of arthritic rats. Exposure of arthritic rats to PEMF at 5 Hzx4 microT x 90 min, produced significant antiexudative effect resulting in the restoration of the altered parameters. The antiinflammatory effect could be partially mediated through the stabilizing action of PEMF on membranes as reflected by the restoration of PMCA and intracellular Ca(2+) levels in blood lymphocytes subsequently inhibiting PGE(2) biosynthesis. The results of this study indicated that PEMF could be developed as a potential therapy for RA in human beings.

Gender differences and protective effects of testosterone in collagen induced arthritis in rats.

To understand the gender differences noticed in autoimmune disorders, particularly rheumatoid arthritis, we used a rat model of collagen induced arthritis (CIA). This study was carried out in two parts. In the first study, severity of inflammation was compared between male and female rats with respect to radiology, histology, activities of lysosomal enzymes, lipid peroxidation, immune response to type II collagen and the level of prostaglandin, a major inflammatory mediator. Since female rats developed severe inflammation, this study was extended to confirm if testosterone at physiological concentration had protective effect against CIA. Hence, studies were carried out on the effect of testosterone application on castrated arthritic rats. Female arthritic rats were also treated with testosterone to find out the effectiveness of the androgen in the presence of female hormones. Results of this study conclusively showed that testosterone possessed significant anti-inflammatory effects at physiological concentration and exerted its action in a gender nonspecific manner.

Myocardial ischemia and reperfusion injury in rats: lysosomal hydrolases and matrix metalloproteinases mediated cellular damage.

The aim of this study was to evaluate the time course events of cellular damage during myocardial ischemia and reperfusion injury in rats and to find out a correlation between the structural alterations with respect to the biochemical changes. Cardiac biomarkers and lysosomal enzymes viz. cathepsin D, acid phosphatase and beta-glucuronidase and matrix metalloproteinases (MMPs) were evaluated at different time points, in response to ischemia-reperfusion induced oxidative stress in an isolated rat heart model perfused in Langendorff mode. Microscopically, changes in myocardial architecture, myofibrillar degradation, and collagen (COL) integrity were studied using hematoxylin-eosin, Masson's trichrome and toluidine blue staining techniques. A three-fold increase in the level of myoglobin was observed after 30 min of ischemia followed by 120 min of reperfusion as compared to 15 min ischemia, 120 min reperfusion. Similarly, a significant increase (P<0.05) in the levels of lipid peroxides and superoxide anion coupled with a decrease in enzymatic and nonenzymatic antioxidant levels were observed. A concomitant increase in the activity of cathepsin D (24.07+/-0.95) and a higher expression of MMPs after 120 min of reperfusion following 30 min ischemia were shown to correlate with the myocardial damage as shown by histopathology, suggesting that free radical induced activation of cathepsin D and MMPs could mediate early damage during myocardial ischemia and reperfusion.