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Fig. 6
Inhibitors of TLR3, activin, and complement partially rescued Rps19-deficient zebrafish embryos. (A) TLR3 inhibitor decreased expression of interferon mediators irf7 and stat1b, decreased expression of inflammatory markers il6 and ptgs2/cox2 and acute phase response gene fos. It also downregulated p21 responsible for cell cycle arrest and pro-apoptotic inflammatory gene bax, 40 hpf, RT-qPCR. Bars represent the mean of three replicates ± sd. Asterisk indicates significant difference in comparison to controls, p < 0.05. (B) TLR3 inhibitor improved hematopoiesis in Rps19 morphants, day 3, o-dianizidine staining. 50 embryos per group, representative staining is shown. (C) Embryos were injected with rps19 morpholino, MO group was left untreated, other groups were treated with TLR3 inhibitor (TLRinh), SB431542 activin inhibitor (SB43) or SB290157 complement inhibitor (SB29). Treatments decreased morphological defects and improved survival in Rps19-deficient zebrafish embryos. Embryos classified as abnormal had strong defects such as tail kinks, curved body, strongly shortened body, arrest at early developmental stage, and underdeveloped eyes. Embryos with mild defects had smaller size, smaller eyes and heads. 24 hpf. The experiment was performed in triplicates, 60 embryos per group; bars represent the mean of three replicates. Number of dead/abnormal/mild/normal embryos for MO 10 ± 0.6/15 ± 0.6/30 ± 1.2/5 ± 1; for TLR3 inhibitor 2 ± 1/6 ± 1/37 ± 1/15 ± 0.6; for SB431542 inhibitor 0 ± 0/4 ± 1/38 ± 2.5/18 ± 1.5; for SB2990157 3 ± 1/7 ± 0.5/35 ± 1.2/15 ± 0.7.