Role of tachykinin 1 and 4 gene-derived neuropeptides and the neurokinin 1 receptor in adjuvant-induced chronic arthritis of the mouse

Abstract

Objective: Substance P, encoded by the Tac1 gene, is involved in neurogenic inflammation and hyperalgesia via neurokinin 1 (NK1) receptor activation. Its non-neuronal counterpart, hemokinin-1, which is derived from the Tac4 gene, is also a potent NK1 agonist. Although hemokinin-1 has been described as a tachykinin of distinct origin and function compared to SP, its role in inflammatory and pain processes has not yet been elucidated in such detail. In this study, we analysed the involvement of tachykinins derived from the Tac1 and Tac4 genes, as well as the NK1 receptor in chronic arthritis of the mouse.

Methods: Complete Freund's Adjuvant was injected intraplantarly and into the tail of Tac1(-/-), Tac4(-/-), Tacr1(-/-) (NK1 receptor deficient) and Tac1(-/-/)Tac4(-/-) mice. Paw volume was measured by plethysmometry and mechanosensitivity using dynamic plantar aesthesiometry over a time period of 21 days. Semiquantitative histopathological scoring and ELISA measurement of IL-1β concentrations of the tibiotarsal joints were performed.

Results: Mechanical hyperalgesia was significantly reduced from day 11 in Tac4(-/-) and Tacr1(-/-) animals, while paw swelling was not altered in any strain. Inflammatory histopathological alterations (synovial swelling, leukocyte infiltration, cartilage destruction, bone damage) and IL-1β concentration in the joint homogenates were significantly smaller in Tac4(-/-) and Tac1(-/-/)Tac4(-/-) mice.

Conclusions: Hemokinin-1, but not substance P increases inflammation and hyperalgesia in the late phase of adjuvant-induced arthritis. While NK1 receptors mediate its antihyperalgesic actions, the involvement of another receptor in histopathological changes and IL-1β production is suggested.

Figure 1. Adjuvant-induced mechanical hyperalgesia throughout the 21-day experimental period.

Each data point represents the mean ± SEM of the percentage decrease of the mechanonociceptive threshold of (A) Tac1^−/−, (B) Tac4^−/−, (C) Tacr1^−/−, (D) Tac1^−/−/Tac4^−/− mice compared to the initial control values (n = 9–24 mice per group; *p<0.05, **p<0.01, ***p<0.001 vs. C57Bl/6; two-way ANOVA followed by Bonferroni’s modified t-test).

Figure 2. Adjuvant-induced oedema throughout the 21-day experimental period.

Each data point represents the mean ± SEM of the percentage increase of the paw volume of (A) Tac1^−/−, (B) Tac4^−/−, (C) Tacr1^−/−, (D) Tac1^−/−/Tac4^−/− mice compared to the initial control values (n = 9–24 mice per group, two-way ANOVA followed by Bonferroni’s modified t-test).

Figure 3. Histopathological changes of the paws on day 21.

Panel A shows a representative histopathological picture of an intact tibiotarsal joint and panel B demonstrates the joint structure of an adjuvant-treated C57Bl/6 wildtype mouse with remarkable synovial swelling, leukocyte infiltration, cartilage damage, and bone destruction. The lower panes demonstrate the joint structures of adjuvant-treated (C) Tac1^−/−, (D) Tacr1^−/−, (E) Tac4^−/−, and (F) Tac1^−/−/Tac4^−/− mice, decreased inflammatory parameters can be observed in the latter two groups. Hematoxylin-eosin staining, 40x magnification (ti: tibia, ta: tarsus, s: synovium). (G) Semiquantitative histopathological scoring on the basis of inflammatory cell accumulation, synovial enlargement, cartilage destruction and bone erosion. Box plots represent the composite scores (n = 4–12 mice per group,⁺⁺⁺p<0.001 vs. intact C57Bl/6, *p<0.05, ***p<0.001 vs. C57Bl/6 CFA-treated, Kruskal-Wallis followed by Dunn’s post test).

Figure 4. Concentrations of the inflammatory cytokine interleukin-1β (IL-1β) in the joint homogenates on day 21.

Each bar represents the mean ± SEM of each group (n = 5–12 mice per group, *p<0.05 vs. C57Bl/6; Kruskal-Wallis followed by Dunn’s post test).