Reversal of the antiinflammatory effects of methotrexate by the nonselective adenosine receptor antagonists theophylline and caffeine: evidence that the antiinflammatory effects of methotrexate are mediated via multiple adenosine receptors in rat adjuvant arthritis.

OBJECTIVE: Weekly low-dose methotrexate (MTX) remains the mainstay of second-line therapy for rheumatoid arthritis (RA). We have previously reported that adenosine, acting at specific receptors on inflammatory cells, mediates the antiinflammatory effects of MTX in both in vitro and in vivo models of acute inflammation, but the mechanism by which MTX suppresses the chronic inflammation of arthritis remains controversial. The present study was undertaken to further investigate the means by which adenosine mediates the antiinflammatory effects of MTX. METHODS: The effects of 2 nonselective adenosine receptor antagonists, theophylline and caffeine, were examined, using the rat adjuvant arthritis model of RA. These agents were given alone and in conjunction with MTX, and arthritis severity was assessed clinically, radiologically, and histologically. Since rodent adenosine A3 receptors are not blocked by theophylline, selective A1, A2A, and A2B receptor antagonists were tested as well. RESULTS: Control animals developed severe arthritis, which was markedly attenuated by weekly treatment with MTX (0.75 mg/kg/week). Neither theophylline alone nor caffeine alone (each at 10 mg/kg/day) significantly affected the severity of the arthritis, but both agents markedly reversed the effect of MTX as measured by a severity index, hindpaw swelling, and hindpaw ankylosis. Radiographic and histologic analyses confirmed these observations. Neither A1, A2A, nor A2B receptor antagonists affected the capacity of MTX to ameliorate inflammation in adjuvant arthritis. CONCLUSION: These results provide strong evidence that adenosine mediates the antiinflammatory effects of MTX in this model of RA. Moreover, the findings suggest that abstinence from caffeine, a ubiquitous food additive and medication, may enhance the therapeutic effects of MTX in RA.

Peroxisome proliferator-activated receptor gamma is induced during differentiation of colon epithelium cells.

Peroxisome proliferator-activated receptor gamma (PPARgamma), a fatty acid-activated nuclear receptor, is implicated in adipocyte differentiation and insulin sensitisation. In view of the association of dietary fat intake and bowel disease, the expression of PPARgamma in rodent and human intestine was studied. Expression of PPARgamma mRNA was examined by Northern blot hybridisation, RNase protection, and/or competitive RT-PCR assays, whereas PPARgamma protein levels were evaluated by immunoblotting and immunohistochemistry. PPARgamma mRNA and protein were abundantly expressed in colon relative to the small intestine both in rodents and in man. Interestingly, expression of PPARgamma was primarily localised in the more differentiated epithelial cells in the colon. The level of expression of PPARgamma in colon was similar to the levels seen in adipose tissue. Expression of PPARgamma increased from proximal to distal segments of the colon in man. In Caco-2 and HT-29 human adenocarcinoma cells, PPARgamma expression increased upon differentiation, consistent with PPARgamma being associated with a differentiated epithelial phenotype. High-level expression of PPARgamma was observed in the colon, but not in the small intestine, suggesting a potential role of this nuclear receptor in the colon.

Role of skeletal muscle in thiazolidinedione insulin sensitizer (PPARgamma agonist) action.

Thiazolidinedione (TZD) insulin sensitizers are specific agonists of peroxisome proliferator activated receptor (PPAR)gamma. However, their mechanism of action and the in vivo target tissue(s) that mediate insulin sensitization remain poorly defined. Although PPARgamma messenger RNA expression has been reported in skeletal muscle, the expression of PPARgamma within myocytes in intact muscle tissue has not been examined. An antipeptide PPARgamma antibody was generated; immunohistochemistry was then used to demonstrate that PPARgamma is present within nuclei of myocytes [in both skeletal (white and red fibers) and cardiac tissue (rodent and human)]. The effect of insulin sensitizer treatment on muscle insulin action was studied using ob/ob mice after 4 days dosing with a potent (6 nM PPARgamma Kd) TZD (10 mg/kg x day). 2-deoxyglucose (2-DOG) uptake was then assessed in freshly isolated soleus muscles from lean vs. ob/ob vs. TZD-treated ob/ob mice. In lean mouse muscles, 2-DOG uptake was stimulated by 82%, 95%, 165% (with 25, 100, 2000 microU/ml insulin); muscles from ob/ob were severely insulin resistant (<80% stimulation with 2000 microU/ml insulin). Muscles from TZD-treated ob/ob displayed a normal insulin response with 100 (71%) or 2000 (158%) microU/ml insulin. Additional studies were performed using ZDF rats treated with/without TZD for 7 days. In vivo 2-DOG glucose uptake into soleus, gastrocnemius, and diaphragm muscles was measured during euglycemic-hyperinsulinemic clamp. Compared with lean rats, muscle 2-DOG uptake in ZDF was reduced by 52% (soleus) or 71% (diaphragm). Partial (40-60%) normalization of the reduced 2-DOG uptake was evident in TZD-treated ZDF rats. In contrast to the effect of in vivo treatment on muscle insulin action, preincubation of isolated soleus muscles from naive lean or ob/ob mice for 5 h with 100 nM TZD did not affect insulin-stimulated 2-DOG uptake. We conclude: 1) PPARgamma is expressed in myocytes within skeletal and cardiac muscle. 2) In vivo activation of PPARgamma by treatment of insulin-resistant mice/rats with a potent TZD corrects impaired muscle insulin action. 3) The lack of a direct effect on muscle after 5 h in vitro TZD incubation suggests that changes in insulin action may require a longer duration of PPARgamma activation or that improved muscle insulin sensitivity may result from an indirect in vivo effect of PPARgamma activation (e.g. changes in systemic lipid metabolism).

Financial resources, parent psychological functioning, parent co-caregiving, and early adolescent competence in rural two-parent African-American families.

We proposed a family process model that links family financial resources to academic competence and socioemotional adjustment during early adolescence. The sample included 90 9-12-year old African-American youths and their married parents who lived in the rural South. The theoretical constructs in the model were measured via a multimethod, multi-informant design. Rural African-American community members participated in the development of the self-report instruments and observational research methods. The results largely supported the hypotheses. Lack of family financial resources led to greater depression and less optimism in mothers and fathers, which in turn were linked with co-caregiving support and conflict. The associations among the co-caregiving processes and youth academic and socioemotional competence were mediated by the development of youth self-regulations. Disruptions in parental co-caregiving interfered with the development of self-regulation. This interference negatively influenced youths' academic competence and socioemotional adjustment.