Modelling, Optimization and Comparable Efficacy of T cells
and HSC Gene Editing for treating HIGM1 (2020)
Figure 7

Figure 7 - CD40LG editing efficiencies achieved in HSPC provide comparable immune rescue as adoptive T cell transfer.

A Schematics of competitive transplant at different ratios of WT (blue) and Cd40lg -/- HSPC (red) (100% WT, 25% WT, 10% WT, 1% WT, 0% WT) into lethally irradiated Cd40lg-/- recipients.

B Total counts of WT donor CD4+ T cells in mice from (A) (n=13 100% WT, 8 25% WT, 14 10% WT, 6 1%, 14 0% WT). Two representative experiments shown out of 3. Mean ± SEM.

C TNP-KLH specific IgG concentration in sera of mice from (A) collected seven days before (pre) and after (post) TNP-KLH vaccination (n=13 100% WT, 13 25% WT, 14 10% WT, 6 1% WT, 14 0% WT). Three independent experiments. Comparisons were performed with an LME model, accounting for multiple experiments, followed by an appropriate post-hoc analysis (Appendix Supplementary Statistical Methods). The reported statistical comparisons refer only to the overall difference among groups (****p<0.0001 in all comparisons). Median ± IQR.

D Percentage of PNA+GL7+ GC B cells within the spleen of mice from (A) (n=16 100% WT, 6 25% WT, 16 10% WT, 11 1% WT, 18 0% WT). Kruskal-Wallis test followed by post-hoc analysis with Dunn's test. P-values were adjusted with Bonferroni's correction to account for multiple comparisons (*p=0.0385 for 25% WT vs 1% WT, *p=0.0129 for 100% WT vs 1% WT, ***p=0.0001 and ****p<0.0001 in both comparisons). Median ± IQR.

E Quantitation of P. murina rRNA in lung homogenate of mice from (A) transplanted with different ratios of WT HSPC cells (n=8 100% WT, 7 25% WT, 7 10% WT, 8 0% WT) and infected with the pathogen. Results are expressed in P.murina/HPRT RNA copies. Kruskal-Wallis test followed by post-hoc analysis with Dunn's test. P-values were adjusted with Bonferroni's correction to account for multiple comparisons. Only the groups 100% WT and 0% WT resulted to be significantly different (**p=0.0028). Median ± IQR.

F Lung weight of mice from (E). Kruskal-Wallis test followed by post-hoc analysis with Dunn's test. P-values were adjusted with Bonferroni's correction to account for multiple comparisons. Only the groups 100% WT and 0% WT resulted to be significantly different (*p=0.0254). Median ± IQR.

G Left: Representative pictures of lung tissue sections stained immunohistochemically with a rabbit primary antibody (upper left: negative sample; lower left: positive sample). Brown areas represent intra-alveolar aggregates of P. murina organisms. Right: Representative pictures of P. murina organisms detected by immunofluorescence in lungs homogenate (upper right: negative sample; lower right: positive sample).

H Quantitation of P. Murina rRNA in lung homogenate of mice adoptively transferred (n=9 CD40LG + T cells) or not (n=8 CD40LG) with in vivo primed CD4+ T cells in absence of conditioning and infected with the pathogen. Results are expressed in P.Murina/HPRT RNA copies. Mann-Whitney test. ***P-value of the comparison = 0.0003. Median ± IQR.

I Lung weight of experimental mice from (H). Mann-Whitney test. **P-value of the comparison = 0.0055. Median ± IQR.

EXPANDED VIEW FIGURE TITLES AND LEGENDS