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. 2020 Sep 7;9:e55732. doi: 10.7554/eLife.55732

Figure 1. C1q induces intracellular signaling activation in hNSC.

(A) Phosphoarray analysis quantification of hNSC exposed to C1q [0.1 nM], [1.0 nM], or [200 nM] concentrations for 15, 30, or 60 min. Data show mean ± SEM (N = 3 biological replicates per condition) for quantified optical intensity, normalized to control (dashed line = 1). In addition to statistical analysis, using one-sample t-test (*p≤0.05, **p≤0.01, ***p≤0.001), a minimum threshold of 1.5-fold activation (dashed line) was set for statistical significance. (B) ERK and Akt activation was verified using western blot in lysates from hNSC exposed to C1q [0.1 nM], [1.0 nM], or [200 nM] concentrations for 60 min. p-ERK and p-Akt band intensities were normalized to β-actin. Data show mean ± SEM (N = 3 biological replicates per condition) for quantified optical intensity normalized to control (dashed line). Statistical analysis using one-sample t-test (NS, not significant; *p≤0.05, **p≤0.01, ***p≤0.001).

Figure 1.

Figure 1—figure supplement 1. C1q induces intracellular signaling activation in hNSC.

Figure 1—figure supplement 1.

(A) Representative images and numerical scheme of phosphoarray analysis of hNSC exposed to control media or C1q [0.1 nM; 1.0 nM; 200 nM] for 30 min. Array positions indicated for p-ERK (blue), p-Akt thr308 (orange), and ser473 (red). (B) Orientation and positions of the intracellular signaling proteins (in duplicate) assayed by phosphoarray: number 3 (blue, p-ERK); number 6 (orange, p-Akt thr308); and number 7 (red, p-Akt ser473). (C) Complete list of targets, intracellular proteins, and modifications tested by phosphoarray. Phosphoarray by PathScan Intracellular signaling array kit as described under Methods (Catalog #7744, Cell Signaling Technology).