Interleukin 22 is a candidate gene for Tmevp3, a locus controlling Theiler's virus-induced neurological diseases

Abstract

After intracerebral inoculation, Theiler's virus induces in its natural host, the mouse, an acute encephalomyelitis followed, in susceptible animals, by chronic inflammation and primary demyelination. Susceptibility to demyelination among strains of laboratory mice is explained by the capacity of the immune system to control viral load during persistence. Also, differences of susceptibility to viral load between the susceptible SJL strain and the resistant B10.S strain are mainly due to two loci, Tmevp2 and Tmevp3, located close to the Ifng locus on chromosome 10. In this article, we show that the Tmevp3 locus controls both mortality during the acute encephalomyelitis and viral load during persistence. Most probably, two genes located in the Tmevp3 interval control these two different phenotypes with efficiencies that depend on the age of the mouse at inoculation. Il22, a member of the IL-10 cytokine family, is a candidate gene for the control of mortality during the acute encephalomyelitis.

Figure 1.—

Genotype of the B10S.Tmevp3^SJL and SJL.Tmevp3^B10S strains on chromosome 10. Microsatellite markers are listed on the left. Distances are arbitrary and do not reflect physical distances. Solid squares, B10.S genome interval; open squares, SJL genome interval; shaded squares, interval in which the crossing over occurs.

Figure 2.—

Phylogenetic tree of 14 inbred strains of mice. From each 11 strains of laboratory mice and 3 wild-derived strains, MAI, MBT, and STF, sequences from 11 DNA segments covering the ∼400-kb region were concatenated in a 3.9 kb long sequence. These concanated sequences were aligned according to CLUSTALW program. A phylogenetic tree was constructed using the TREE-PUZZLE program with the Hasegawa–Kishino–Yano model of substitution. Parameters were estimated from the data. The tree was drawn by the DRAWTREE program. Italics, strains of laboratory mice in which viral load is predicted by the H2 haplotype; underlined type, laboratory mouse strain in which viral load is not predicted by the H2 haplotype; boldface type, wild-derived strains.

Figure 3.—

Viral load in the spinal cords of the four groups of mice inoculated at 3–4 weeks of age, at 45 days post-inoculation. SQRT, square root; rectangle, 90% interval of viral load around median; error bar, 95% interval around median. Viral load among the four groups of mice is significantly different (P < 0.01). Effect of the Tmevp3 locus, the background, and the interaction between both factors are shown below. Significant differences between two groups of mice are shown. P = NS, nonsignificant difference; *P < 0.05; **P < 0.01; ***P < 0.001. n, number of mice.

Figure 4.—

Mortality during the acute encephalomyelitis of bone marrow chimeras. Cumulative survival curve for B10S.Tmevp3^SJL recipient strain (A) and B10.S recipient strain (B) are obtained using the Kaplan-Meier model stratified according to the donor strain (circle, B10.S; square, B10S.Tmevp3^SJL; triangle, SJL.Tmevp3^B10S; diamond, SJL). n, number of mice. The strain before the arrow is the donor strain and that after is the recipient strain.

Figure 5.—

Viral load (A) and expression of Il22 gene (B) in the four groups of mice at 21 days p.i. SQRT, square root; rectangle, 90% interval of viral load around median; error bar, 95% interval around median. Viral load is not significantly different between each parental strain and the congenic strain of the same background. Expression of Il22 gene among the four groups of mice is significantly different (P < 0.01). Effect of the Tmevp3 locus, the background, and the interaction between both factors on Il22 expression are shown below. Significant differences of Il22 expression between two groups of mice are shown. P = NS, nonsignificant difference; *P < 0.05; **P < 0.01; ***P < 0.001. n, number of mice.

Figure 6.—

Allelic expression of Il22 gene in (B10S.Tmevp3^SJL × B10.S)F₁ animals at 21 days p.i. (A) and in bone marrow chimeras at 45 days p.i. (B). For each sample, fluorescence (in arbitrary unit) of the SJL allele (x-axis) and that of the B10.S allele (y-axis) are measured using an allelic-specific probe after 40 cycles of amplification by Taqman RT–PCR. The strain before the arrow is the donor strain and the strain after is the recipient strain. Samples of spinal cord at 21 or 45 days p.i. from B10.S and SJL mice with very different level of Il22 expression were chosen as positive controls. Each sample of a B10.S mouse was matched with one of an SJL one according to their level of Il22 expression. In all positive control, the PCR amplification reached a plateau and the specificity of detection of each Il22 allele was always verified.