Autoimmune cardiomyopathy and heart block develop spontaneously in HLA-DQ8 transgenic IAbeta knockout NOD mice - PubMed (original) (raw)

Fig. 3.

(A) Identical Western blots made with heart muscle proteins and probed with sera from two different DQ8+/+,_IA_β-/- NOD animals (Left, 6-week-old animal without heart block, 1:10,000 dilution; Right, 28-week-old animal with complete AV block, 1:50,000 dilution). Lanes contain muscle proteins extracted from: 1, mouse atria; 2, mouse ventricles; 3, mouse skeletal muscle or purchased commercially from Sigma; 4, purified porcine skeletal myosin; and 5, purified porcine cardiac myosin. (B) Titration curves for anticardiac myosin antibodies present in the sera of DQ8+/+,_IA_β-/- NOD mice of various ages. Sera were pooled from animals ages 2.5 weeks (n = 9; ▴), 5 weeks (n = 7; ⋄), 10 weeks (n = 7; •), and 20 weeks (n = 6; ○). As a control, values for pooled sera from DQ8+/+,_IA_βg7+/+ NOD mice (n = 5; □) are also shown. A number of other control strains (NOD/Lt, IAβ-/-NOD, C57BL/6) were similarly negative. Total serum IgG concentrations in older preterminal DQ8+/+,_IA_β-/- NOD animals were somewhat higher (up to 4-fold increased) compared with those found in control strains. ELISA plates were coated with porcine cardiac myosin, incubated with various dilutions of pooled antisera and then secondary antibodies, and developed with horseradish peroxidase. The data are expressed as mean of pooled sera tested in triplicate. (Inset) Data replotted as absorbance versus age for pooled sera at the 1:100,000 dilution. (C) Incidence of heart block in irradiated animals that received adoptive transfer of splenocytes, cells (n = 12), serum (n = 12), both agents (n = 17), or PBS only (n = 20). All animals were irradiated at 5 weeks of age, injected as indicated, and then followed with weekly ECGs. The percentage of animals showing first-degree or complete heart block at various weeks posttransfer are indicated. The serum and PBS groups were not statistically different, nor were the cells vs. cells plus serum groups. At 11 and 12 weeks, the cells group statistically differed from the PBS group with P = 0.03. Similarly, at 11 and 12 weeks, the cells plus serum group differed from the PBS group with P = 0.001. Fisher's exact test was used to calculate two-tailed P values. (D) Heart block in animals receiving CsA injections vs. PBS. Beginning at weaning, animals were given daily injections of CsA (10 mg/kg s.c) or PBS (n = 10 animals in each group). Values indicate the percentage of animals with first-degree or complete heart block. CsA injections were discontinued at 15 weeks of age (indicated by arrow). At 14 and 15 weeks of age, the two groups were statistically different (P = 0.023) by Fisher's exact test.