[Experimental study on induction of skin allograft tolerance in mice by heat shock protein 60]

To investigate the role and mechanism of heat shock protein 60 (HSP60) in induction of murine skin allograft tolerance.

At the age of 8-12 weeks, inbred female BALB/C (H-2d) mice (n=45) and CBA/N (H-2k)mice (n=15) were used as transplantation donors and C57BL/6 (H-2b) mice (n=60) as recipients. Recipients C57BL/6 (H-2b) mice were randomized into 4 groups (n=15). In group A, 1 cm x 1 cm Wolfe-Krause skin graft was excised from the back of BALB/C (H-2d) mice and hypoderma was scraped off aseptically, and then transplanted to the back of C57BL/6 (H-2b) mice. The method of skin transplantation in the other 3 groups was the same as to group A. In group B, C57BL/6 (H-2b) mice were treated with incomplete Freund's adjuvant (IFA) administration into the back 2 weeks before transplantation of BALB/C (H-2d) mice skin. In group C, C57BL/6 (H-2b) mice were administered HSP60 emulsified in IFA into the back 2 weeks before transplantation of BALB/C (H-2d) mice skin. In group D, C57BL/6 (H-2b) mice were treated with HSP60 emulsified in IFA into the back and followed by skin transplantation of CBA/N (H-2k) mice 2 weeks later. The delayed type hypersensitivity was determined 7 days after transplantation. One-way mixed lymphocyte reaction, the concentration of cytokines in the mixed lymphocyte reaction culture supernatant was determined 7 days and 25 days after transplantation. The survival time of skin allograft was observed.

The survival time of skin allograft in groups A, B, C and D was 12.4 +/- 0.5, 11.6 +/- 0.8, 29.3 +/- 2.6 and 27.6 +/- 2.1 days, respectively. There was significant difference between groups A, B and groups C, D (P < 0.05), while there was no significant difference between group A and group B as well as between group C and group D (P > 0.05). The counts of per minute impulse (cpm) of mixed lymphocyte reaction 7 days after transplantation in groups A, B, C and D was 12 836 +/- 1357, 11876 +/-1265, 6581 +/- 573 and 6843 +/- 612, respectively. There was significant difference between groups A, B and group C and group D (P < 0.05), while there was no significant difference between group A and group B as well as between group C and group D (P > 0.05). The cpm of mixed lymphocyte reaction at 25 days after transplantation in group A, B, C and D was 13286 +/- 1498, 12960 +/- 1376, 11936 +/- 1265 and 12374 +/- 1269, respectively. There was no significant difference among 4 groups (P > 0.05).The concentration of IL-10 in the mixed lymphocyte reaction culture supernatant in groups C, D were higher than that in groups A, B, and IL-2 and IFN-gamma were lower than that in groups A, B 7 days after transplantation (P < 0.05), while there was no significant difference between group A and group B as well as between group C and group D (P > 0.05). There was no significant difference in cytokines among the 4 groups 25 days after transplantation (P > 0.05). The delayed type hypersensitivity in groups A, B, C and D 7 days after transplantation was 0.84 +/- 0.09, 0.81 +/- 0.07, 0.43 +/- 0.05 and 0.46 +/- 0.03 mm, respectively. There was significant differences between groups A, B and groups C, D (P < 0.05). While there was no significant difference between group A and group B as well as between group C and group D (P > 0.05).

HSP60 may play a role in induction and maintenance of murine skin allograft tolerance.