Peptide-induced nasal tolerance for a mycobacterial heat shock protein 60 T cell epitope in rats suppresses both adjuvant arthritis and nonmicrobially induced experimental arthritis

Abstract

Adjuvant arthritis (AA) can be induced in Lewis rats by immunization with mycobacterial antigens. Passive transfer of a T cell clone recognizing the 180-188 amino acid sequence in mycobacterial heat shock protein 60 (hsp60) was found to induce AA. In the present study, we investigated whether tolerance was obtained for this AA-associated T cell epitope after intranasal or s.c. administration of a peptide containing this epitope. Two 15-mer peptides containing the mycobacterial hsp60 sequences 176-190 and 211-225 were used; 176-190 contained the T cell epitope 180-188, which was recognized by the arthritogenic T cell clone A2b and was the immunodominant hsp60 T cell epitope after induction of AA, and 211-225 contained a T cell epitope that was recognized both after induction of arthritis with whole Mycobacterium tuberculosis and after immunization with mycobacterial hsp60. In rats treated intranasally or subcutaneously with 176-190 and immunized with mycobacterial hsp60, proliferative responses to 176-190 were reduced. Proliferative responses to 211-225 and to whole mycobacterial hsp60 were not affected. AA was inhibited intranasally in the 176-190-treated rats but not in the 211-225-treated rats. Moreover, intranasal 176-190 led to similar arthritis-protective effects in a nonmicrobially induced experimental arthritis (avridine-induced arthritis). Therefore, tolerance for a disease-triggering, microbial cartilage-mimicking epitope may cause resistance to arthritis irrespective of the actual trigger leading to development of the disease.

Figure 1

Modulation of the proliferative response to peptide 176–190 after i.n. administration of PBS (controls) or peptide 176–190. Rats received 100 μg of 176–190 dissolved in 10 μl of PBS or PBS alone i.n. at days −15, −10, and −5. At day 0, rats were immunized with 50 μg of mycobacterial hsp60 in DDA in the hind footpads. PLNC were isolated 14 days after immunization and tested for responses to peptide 176–190 and 211–225.

Figure 2

DTH after i.n. administration or after immunization in DDA with peptide 176–190 or 211–225. Four groups were tested: i.n. administration of 176–190 and 211–225 (100 μg of peptide in 10 μl of PBS) and immunization in the hind footpads with 100 μg of the same peptides in DDA (three rats in each group). Shown are DTH responses in rats after i.n. administration at days −15, −10, and −5 and after immunization at day −10. DTH was measured by injecting 10 μg of peptide in 10 μl of PBS s.c. in the left ear and 10 μl of PBS in the other ear. The difference in thickness between left and right ears was measured (using a micrometer calipers for each animal 24 (black bars) and 48 h (shaded bars) after the challenge. The results were expressed as the mean for each experimental group (mm/100) ± SEM.

Figure 3

Modulation of AA development after i.n. administration of peptide 176–190 and 211–225. Rats received 100 μg of peptide (176–190 or 211–225) dissolved in 10 μl of PBS or PBS alone i.n. at days −15, −10, and −5. At day 0, rats were immunized with 0.5 mg of Mt in 100 μl of incomplete Freund’s adjuvant to induce AA. Six rats were in each treatment group. Arthritis scores were assessed every other day after Mt immunization. On the Y axis are shown the mean arthritis scores ± SEM (error bars).

Figure 4

Mean change in body weight in grams after i.n. administration of peptide 176–190 and 211–225 in AA. Experimental details as in Fig. 2. Body weight was measured every other day. Y axis shows the mean changes of body weight from day 10 after Mt immunization.

Figure 5

Modulation of avridine-induced arthritis after i.n. administration of 176–190 and 211–225. Rats received 100 μg of peptide (176–190 or 211–225) dissolved in 10 μl of PBS or PBS alone i.n. at days −15, −10, and −5. At day 0, rats were immunized with 2 mg of avridine in 100 μl of mineral oil injected intradermally at the base of the tail to induce arthritis. Six rats were in each treatment group. Arthritis scores were assessed daily from day 8 after avridine immunization. On the Y axis are shown the mean arthritis scores ± SEM (error bars).

Figure 6

Lymphocyte proliferative responses on primed lymph node cells from pooled popliteal and inguinal lymph nodes after the induction of AA. Peptide concentration 20 μg/ml; One rat in the 176–190-treated group suffered from severe arthritis (maximum arthritis score 12 at day 21); PLNC derived from this animal showed clear proliferative responses to 176–190 (SI 3.2, marked with an asterisk).