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. 2021 Dec 17;14(12):1322. doi: 10.3390/ph14121322

Figure 4.

Figure 4

The eNAMPT-neutralizing ALT-100 mAb inhibits NFκB phosphorylation in PCa cells and xenografts. DU145 cells were exposed to vehicle (PBS), denatured eNAMPT (den-eNAMPT), anti-eNAMPT mAb (mAb), eNAMPT, or mAb mixed with eNAMPT. Immunoblot for phosphor-NFkB p65 (p-NFkB), total NFkB, and internal control beta-actin showed that mAb-treatment significantly decreased eNAMPT-induced NFκB phosphorylation in DU145 cells (A) and PC3 cells (B). Immunoblot density measurement showed that mAb treatment significantly decreased eNAMPT-induced NFκB phosphorylation level in DU145 cells (C) and PC3 cells (D) (* p < 0.05). IHC for phosphor-NFκB p65 (p-NFκB) showed high levels of NFκB phosphorylation (brown) in vehicle-treated xenografts, whereas the level was markedly decreased in mAb-treated DU145 (E) and PC3 xenografts (F). Measurement of IHC density showed that phosphor-NFκB p65 (p-NFκB) was significantly lower in mAb-treated DU145 (G) and PC3 xenografts (H) compared to vehicle IgG-treated mice (* p < 0.05). 100x. Immunoblots for phosphor-ERK1/2 (p-ERK1/2), total ERK1/2, and internal control beta-actin showed that mAb-treatment significantly decreased eNAMPT-induced ERK1/2 phosphorylation in DU145 (I,K) and PC3 cells (J,L) quantified by immunoblot densitometric measurements (*,** p < 0.05).