Proinflammatory signaling by nuclear cytokines in the absence and presence of polyP-70. (A) Cell permeability in response to increasing concentration of HMGB1 (16 hours) was measured by spectrophotometric measurement of the flux of Evans blue-bound albumin across functional endothelial cell monolayers using a modified 2-compartment chamber model. (B-C) The same as (A) except that the barrier disruptive effect of increasing concentrations of either H4 (4 hours) or polyP-70 (4 hours), respectively, were monitored. (D) The same as (A), except that the barrier-disruptive effect of different increasing concentrations of polyP-70 in complex with a fixed concentration of HMGB1 (20 nM) was monitored. (E-F) The same as (D), except that the barrier-disruptive effect of different concentrations of HMGB1 and H4, respectively, was monitored in the presence of a fixed concentration of polyP-70 (2.5 µM). (G-J) The same as other panels, except that the competitive effect of sRAGE on the barrier disruptive effect of HMGB1 (40 nM) and H4 (1.78 µM) was monitored in either the absence (G-H) or presence of polyP-70 (I-J). In the presence of polyP (2.5 µM), the concentrations of nuclear cytokines were reduced to 10 nM for HMGB1 and 0.44 µM for H4 (I-J). All results are shown as mean ± standard deviation of 3 different experiments. OD, optical density. *P < .05; **P < .01; ***P < .001.