Page 170 - Haematologica August 2018

M.T. Georgescu et al.
Messenger ribonucleic acid (mRNA) expression analysis
E16KO mice received HBSS, Dex, FVIII or FVIII+Dex for five consecutive days. Three days after the last injection, spleen and thymus were collected and stabilized in RNA Later (Invitrogen). mRNA was then isolated using a commercial kit (RNeasy Plus Mini Kit, Qiagen) and quantified using the NanoString Mouse Immunology Panel (NanoString). Data was analyzed with nSolver Software (NanoString). Genes that were deemed up/down-regu- lated due to FVIII+Dex treatment had a transcript count ratio ≥2:1 in the same direction when comparing both FVIII+Dex against FVIII and Dex against HBSS.
Statistics
Anti-FVIII IgG and FVIII inhibitors incidence were compared using Fisher’s exact test. Anti-FVIII IgG and Bethesda titers were compared using a Mann-Whitney U test. E17KO/hMHC samples, but not E16KO samples, with titers below the detection limit were excluded from statistical analyses. Percentages of lymphocyte populations were compared using unpaired two-tailed t-tests. Statistical analyses were performed using GraphPad Prism 5.0a (GraphPad Software).
AB
C
P=0.0048
P=0.0485 D P=0.00063
EF
1406
Figure 2. Administration of Dex during initial FVIII exposure reduces the initial anti-FVIII immune response in both E17KO/hMHC and E16KO mice. A. Anti-FVIII IgG incidence and B. anti-FVIII IgG titers in E17KO/hMHC mice five weeks after initial treatment with FVIII or FVIII+Dex. A statistical comparison of the positive anti-FVIII IgG titers across treatment groups could not be performed because fewer than three mice had evidence of antibodies in the FVIII+Dex group. G. Anti-FVIII IgG inci- dence, D. anti-FVIII IgG titers, E. FVIII inhibitor incidence and F. FVIII inhibitory activity in E16KO mice five weeks after initial treatment with FVIII or FVIII+Dex. ND: not detectable; FVIII: factor VIII; Dex: dexamethasone; IgG: immunoglobulin G.
haematologica | 2018; 103(8)