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. Author manuscript; available in PMC: 2016 Oct 18.
Published in final edited form as: J Invest Dermatol. 2016 May 7;136(8):1656–1663. doi: 10.1016/j.jid.2016.04.022

Figure 4. Lce1 genes are transcriptionally upregulated after NRF2 activation and downregulated following blocking of endogenous NRF2 activity.

Figure 4

(a) Keratinocytes from WT newborn mice were isolated, cultured, and incubated with the indicated concentration of tBHQ. Nuclear protein fractions (upper panels) and whole cell lysates (lower panels) were subjected to immunoblotting. tBHQ induced the accumulation of NRF2 in the nucleus (upper panels). There were increased expression levels of LCE1 proteins in a tBHQ dose-dependent manner, similar to those of validated NRF2 downstream targets; NQO1 and SPRR2 (lower panels). − (DMSO alone), + (tBHQ 10 μM), ++ (tBHQ 30 μM). (b) Keratincytes were isolated from 4-day-old LKO mice and cultured along with littermate controls (heterozygous for Lor). After treatment with tBHQ, LKO keratinocytes exhibited significantly increased transcripts for Lce1b, c, e, g, h, and m, compared with littermate controls. − (DMSO alone), + (tBHQ 4 μM), ++ (tBHQ 10 μM). (CTL (Lor±) n = 3, LKO n = 3, **P < 0.01, ***P < 0.005, ****P < 0.001). (c) The transcripts of Lce1b, c, e, g, h, and m are decreased in the back skin of LKO mice expressing the dnNRF2 transgene, compared with littermate LKO controls (WT n = 5, LKO n = 5, LKOdnNRF2 n = 3, ***P < 0.005, ****P < 0.001, LKOdnNRF2 vs LKO). CTL, control; DMSO, dimethyl sulfoxide; dnNRF2, dominant negative Nrf2 transgene; LCE, late cornified envelope; LKO, loricrin knockout; NQO1, NAD(P)H dehydrogenase, quinone 1; NRF2, NF-E2-related factor 2; tBHQ, tert-butylhydroquinone; WT, wild type.