Pathophysiology of vascular dysfunction in a rat model of chronic joint inflammation

Abstract

The impact of chronic joint inflammation on articular vascular function in rats was investigated to address whether joint swelling and the associated vascular dysfunction are dependent upon a common prostanoid mechanism. Urinary nitrate/nitrite (NO(x)) and PGE(2) excretion, knee joint diameter and body weight were measured following induction of adjuvant-induced arthritis (AIA). Ten days postinduction of AIA, joint vascular reactivity was assessed by measuring the perfusion response using a laser Doppler imager (LDI) to topical application of acetylcholine (ACh) and sodium nitroprusside (SNP). Four groups were compared: a non-inflamed control group and three AIA groups treated i.p. with vehicle, indomethacin or SC-236 (at equimolar doses). The selective cyclooxygenase-2 (COX-2) inhibitor (SC-236) was used to differentiate between COX-1 and -2-derived prostaglandins. Urinary NO(x) and PGE(2) levels increased substantially during the early phase of AIA but decreased thereafter. Toxicity to indomethacin but not SC-236 was observed, as indicated by a marked decrease in body weight. Joint swelling was similarly attenuated by indomethacin and SC-236 (P= 0.0001 cf. vehicle-treated AIA; n= 5-6 per group), indicating that this is due to COX-2 and not COX-1 inhibition. The AIA-induced changes in urinary NO(x) and PGE(2) were corrected by both COX inhibitors. While vascular reactivity to ACh and SNP was significantly attenuated by AIA (P < 0.002; n= 5-10 per group), the perfusion responses to these vasodilating agents were similar in all three AIA groups, demonstrating that the vascular dysfunction was not corrected by inhibition of either COX-1 or COX-2 enzymes. Furthermore, the attenuation of both ACh and SNP-induced responses in AIA suggest that vascular dysfunction was not exclusively endothelial in nature. In conclusion, the joint swelling and vascular dysfunction associated with AIA appear to be mediated, at least in part, by independent mechanisms. While COX-1/COX-2 inhibition reduced joint swelling, vascular dysfunction in AIA is independent of constitutive or inducible prostanoid mechanisms, and appears not to be solely endothelial-derived, but to involve other components such as the vascular smooth muscle.

Figure 1. Ipsilateral and contralateral knee joint diameters

A, ipsilateral knee joint diameter in AIA. Diameter increased significantly in the vehicle-treated group (•) (P < 0.0001, one-way ANOVA). Both SC-236 (▴) and indomethacin (▪) significantly decreased swelling compared to vehicle-treated animals (P = 0.0001, two-way ANOVA). Knee joint diameter in control non-inflamed animals (○) did not significantly change with time (P = 0.138; n = 5–6; one-way ANOVA). B, contralateral knee joint diameter in AIA. Diameter increased significantly in the vehicle-treated AIA group (•) compared to the control non-inflamed group (P = 0.0001, two-way ANOVA). Both SC-236 (▴) and indomethacin (▪) decreased knee joint diameter by a magnitude proportionate to that of the ipsilateral knee (both P = 0.0001, two-way ANOVA, compared to untreated group). Control non-inflamed values (○) did not significantly change with time (P = 0.5; n = 5; one-way ANOVA).

Figure 2. Body weight

Weights in control non-inflamed animals (○) increased in a time-dependent fashion (P < 0.0001, one-way ANOVA), differing significantly from vehicle-treated AIA (•), SC-236 (▴) and indomethacin (▪) groups (P < 0.0001, 0.0004 and 0.0001, respectively, two-way ANOVA). Animal weights in both vehicle- and SC-236-treated groups did not significantly differ with time, but decreased in the indomethacin-treated group (P < 0.029, one-way ANOVA). Body weights of SC-236 and vehicle groups differed significantly from those of the indomethacin group (P < 0.0001, respectively, two-way ANOVA). Data presented as mean ±s.e.m. (n = 5–6).

Figure 3. Urinary NO_x and PGE₂ changes

A, urinary NO_x changes in AIA. The vehicle-treated AIA group (•) was significantly different from SC-236 (▴), indomethacin (▪) and non-inflamed control (○) groups (P < 0.01, P < 0.005 and P < 0.01, respectively, two-way ANOVA). Both SC-236 and indomethacin reduced NO_x levels to control levels (no significant difference from non-inflamed controls; two-way ANOVA). The non-inflamed controls did not significantly change with time (one-way ANOVA). Data presented as mean ± s.e.m. (n = 5–6). B, urinary PGE₂ changes in AIA. There was an acute increase in PGE₂ levels in the vehicle-treated AIA group (•) after day 1 (just failing to reach significance with time compared to SC-236 (▴), indomethacin (▪) and non-inflamed control (○) groups). The response over the first 3 days was significantly different from non-inflamed controls (P < 0.05, two-way ANOVA). The non-inflamed control group did not significantly change with time (one-way ANOVA) and did not differ significantly from either indomethacin or SC-236 (two-way ANOVA). Data presented as mean ±s.e.m. (n = 4).

Figure 4. Vascular reactivity to ACh and SNP

A, vascular reactivity to ACh in AIA rats. ACh induced a dose-dependent synovial vasodilatation (P = 0.009, two-way ANOVA), which was significantly attenuated in the vehicle (hatched columns; P < 0.002, Bonferroni post hoc), indomethacin (grey columns; P < 0.0001, Bonferroni post hoc) or SC-236-treated (filled columns; P < 0.0001, Bonferroni post hoc) AIA groups compared with the non-inflamed controls (open columns). There was no significant difference between the three AIA-treated groups. Data presented as mean ± s.e.m. (n = 5–10). B, vascular reactivity to SNP in AIA rats. SNP induced a dose-dependent synovial vasodilatation (P = 0.013, two-way ANOVA) which was significantly attenuated in the vehicle (hatched columns; P = 0.013, Bonferroni post hoc), indomethacin (grey columns; P = 0.0045, Bonferroni post hoc) or SC-236-treated (filled columns; P = 0.0002; Bonferroni post hoc) AIA groups compared with the non-inflamed controls (open columns). There was no significant difference between the three AIA-treated groups. Data presented as mean ±s.e.m. (n = 5–6).