(A) Immunoblotting for STAT phosphorylation after stimulation for 15 min with IFN-α (100 IU/mL) of HC and DS hTERT-immortalized fibroblasts previously stimulated (Primed) or not (Naive) with a primary stimulus of IFN-α (10 IU/mL) for 12 h, washed, and allowed to rest for 36 h. Result representative of fibroblasts derived from n=3 HCs and n=3 individuals with DS.
(B) Immunoblotting for STAT phosphorylation after stimulation for 15 min with IFN-α (100 IU/mL) of IFNAR2 knockouts complemented with doxycycline-inducible IFNAR2, treated with indicated amounts of doxycycline and previously stimulated (Primed) or not (Naïve) with a primary stimulus of IFN-α (10 or 100 IU/mL). Results representative of n=3 independent experiments.
(C) Immunoblotting for STAT phosphorylation after stimulation for 15 min with IFN-α (100 IU/mL) of IFNAR2 knockouts complemented with doxycycline-inducible IFNAR2, treated with indicated amounts of doxycycline and previously stimulated (Primed) or not (Naïve) with a primary stimulus of IFN-α (10 or 100 IU/mL).
(D) Immunoblotting for STAT phosphorylation after stimulation for 15 min with IFN-α (100 IU/mL) of IFNAR2 knockouts complemented with doxycycline-inducible IFNAR2, treated with increasing concentrations of doxycycline and Primed (with 10 IU/mL IFN-α) or not (Naïve), in the presence (parental line) or absence (USP18 KO Clones 1 and 2) of endogenous USP18.
(E) Diagram of hyper- and hypo-responses to IFN-I over time with subsequent stimulations in HC and DS individuals.
See also Figure S3.