Continuous >95% oxygen (O2; hyperoxia) has been used by research investigators for many years as a prototypical agent to induce ALI in laboratory animals. A screen of a large panel of inbred mice identified C57BL/6J (B) mice as sensitive and 129X1/SvJ (X1) mice as considerably more resistant to the lethal effects of hyperoxia. Genetic analysis of 1,775 recombinant mice, including 840 F2 and 935 backcrosses derived from these strains identified 5 QTLs (i.e. chromosomal regions, designated as Shali1-5) significantly linked to HALI survival time. Mapping results and confidence intervals for the two major loci, Shali1 (chromosome 1) and Shali2 (chromosome 4), are presented in Figure 1. Further analyses revealed that overall survival time also involved decreased penetrance (i.e., the percent of mice that carried the susceptibility gene and expressed the trait) and was significantly affected by sex, cross, and the parent-of-origin.
A closer look at the allelic effects of these two loci in an F2 population derived from X1 and B strain mice revealed opposing actions on survival time. In particular, homozygous X1 alleles were resistant for Shali1, but sensitive for Shali2. Homozygous B alleles were resistant for Shali2, but sensitive for Shali1 (Figure 2). Mice with both sets of sensitive or resistance alleles had significant decreased or increased survival time, providing support for an important gene-gene interaction (Figure 3).
To verify the QTL effects and assess the role of each QTL in HALI survival time, we are constructing reciprocal congenic strains for the 5 Shali regions, a process that involves repeated backcrossing and marker screening of offspring to substitute the QTL region of one strain (donor) with the identical region of another strain (recipient). Figure 4 gives a chromosomal overview of the Shali1 (chromosome 1) and Shali2 (chromosome 4) congenic lines compared to each other and their parental background strains. These congenics have demonstrated dramatic changes in phenotype (Shali1 decreased and Shali2 increased survival time), as compared to the parental X1 strain. Such congenic lines serve as useful models to narrow the region, identify the specific gene(s) involved, and determine the important gene-gene interactions. When refined and validated, these lines will allow detailed mechanistic studies to assess the developing ALI pathology.