Figure 3

The IsdB x C1q bsAb increases clearance of S. aureus in mice with humanized C1q heads. (a) C1q humanized mice were produced by knocking out the mouse C1qA, C1qB and C1qC genes. They were replaced with chimeric versions of the genes in which the N-terminus of the gene encoding the collagen-like tail was mouse, but the C-terminus of the gene encoding the globular head was human. The resulting locus (not to scale) is shown with human sequences in blue and mouse sequences in red. C1q exons are labeled E1, E2, and E3. pA, polyA sequence (top). A representation of the resulting C1 complex is shown below. Human C1q heads are shown in blue, mouse C1q stalks are shown in red, mouse C1r is shown in green and mouse C1s is shown in yellow. (b) Quantitative measurement of the expression of chimeric C1q peptides was assessed via LC-MS/MS. Serum was isolated from individual C1q humanized mice and the concentration of humanized mouse C1q specific peptides was determined. Results are plotted as median and interquartile range. (c) CH50 hemolysis assays were used to determine the activity of serum from humanized C1q mice. Sheep erythrocytes were sensitized to complement lysis by incubation with rabbit anti-sheep antibodies. Dilutions of wild-type and C1q humanized mouse serum, as well as human serum, were then added. Lysis of the red blood cells was used to determine the activity of the sera. (d) C1q humanized mice were infected with 1.5 × 10⁸ CFU S. aureus Newman (MSSA strain). One day after infection, mice were treated with a single dose of the indicated antibody. Weights were recorded each day for four days and percent weight change compared to day 0 is shown (left). Mean weight change for each group of mice is plotted. On day 4, kidneys were harvested, dissociated, and serially diluted. Organ burden was determined by plating (right). The organ burden of each mouse is indicated along with the median value for each group. (e) C1q humanized mice were infected with 1.3 × 10⁸ CFU S. aureus MW2 (MRSA strain) on day 1. One day after infection they were treated with either PBS (no treatment), a single dose of anti-IsdB x anti-C1q antibody, 110 mg/kg vancomycin twice daily, 50 mg/kg daptomycin once daily or 80 mg/kg linezolid twice daily. On day 4, kidneys were harvested, dissociated, and serially diluted. Organ burden was determined by plating. The organ burden of each mouse is indicated along with the median value for each group. (d,e) **P < 0.01, ***P < 0.001, nonparametric Kruskal-Wallis ANOVA with Dunn’s test showing values significantly different from the no treatment samples.