TABLE 1.

Effects of various signal transduction inhibitors on TNF-α− or LPS-induced Stx2 cytotoxicity^a

Inhibitor	Source	Target	LPS	TNF-α	Positive control	Reference(s)
Wortmannin	EMD Biosciences	Phosphatidylinositol 3-kinase	−	−	pAkt	33
PKCζ pseudosubstrate	Biosource	PKCζ	50%	−	ELISA, IL-6	14, 44
PKCι siRNA	Dharmacon	PKCι	−	−	PKCι	25
Bay11-7082	EMD Biosciences	IκB kinase	−	−	Cell-based ELISA, ICAM-1	43
SN50	EMD Biosciences	NF-κB	−	−	Cell-based ELISA, ICAM-1	35
SB202190	EMD Biosciences	p38	+	+	NA	19
SB203580	EMD Biosciences	p38	+	+	NA	19
Ebselen	EMD Biosciences	Traf2/ASK1	ND	−	Toxic	66
SP600125	EMD Biosciences	JNK	−	−	p-c-Jun	30
Jak inhibitor 1 (pyridone 6)	EMD Biosciences	JAK	−	−	IRF-1	34, 57
PD98059	Cell Signaling Technology	MEK1/2	−	−	pERK1/2	42
U0126	EMD Biosciences	MEK1/2	−	−	pERK1/2	17
Cycloheximide	Sigma Aldrich	Protein synthesis	+	+	NA	60

^aHUVEC were incubated with the indicated inhibitors for up to 1 h (see Materials and Methods), treated with 200 U/ml TNF-α or 10 μg/ml LPS with the inhibitor for 24 h, and then treated with 1 nM Stx2 for 24 h. CCK-8 or neutral red cell viability assays were performed to determine if an inhibitor protected against TNF-α- or LPS-induced Stx2 cytotoxicity. +, protection; −, inhibitor had no effect; 50%, LPS-induced Stx2 cytotoxicity was reduced by one-half; ND, not determined. The data are representative of at least three independent experiments. Positive controls indicate cellular responses that were modified by identical concentrations of the inhibitors in HUVEC. Supporting data are shown in Fig. S1 in the supplemental material. NA, not applicable.