Effects of the differentiated keratinocyte phenotype on expression levels of CYP1–4 family genes in human skin cells

Du, Liping; Neis, Mark M.; Ladd, Patricia A.; Lanza, Diane L.; Yost, Garold S.; Keeney, Diane S.

doi:10.1016/j.taap.2005.10.003

Cited by 49 publications

(41 citation statements)

References 49 publications

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“…We found that CYP450 that are important in the bioactivation of these drugs in the liver do not appear to play a significant role in their bioactivation in NHEKs. It should be noted, however, that recent data suggests the level of CYP450 expression may differ as keratinocytes differentiate (Du et al, 2006). Nevertheless, we observed significant protein haptenation in NHEKs in the absence of evidence for involvement of CYP450.…”

Section: Discussioncontrasting

confidence: 86%

Enzyme-Mediated Protein Haptenation of Dapsone and Sulfamethoxazole in Human Keratinocytes: I. Expression and Role of Cytochromes P450

Vyas

Roychowdhury²,

Khan³

et al. 2006

J Pharmacol Exp Ther

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Cutaneous drug reactions (CDRs) are among the most common adverse drug reactions and are responsible for numerous minor to life-threatening complications. Several arylamine drugs, such as sulfamethoxazole (SMX) and dapsone (DDS), undergo bioactivation, resulting in adduction to cellular proteins. These adducted proteins may initiate the immune response that ultimately results in a CDR. Recent studies have demonstrated that normal human epidermal keratinocytes (NHEKs) can bioactivate these drugs, resulting in protein haptenation. We sought to identify the enzyme(s) responsible for this bioactivation in NHEKs. Using immunofluorescence confocal microscopy and an adduct-specific enzyme-linked immunosorbent assay (ELISA), we found that N-acetylation of the primary amine of SMX and DDS markedly reduced the level of protein haptenation in NHEKs. Detection of mRNA and/or protein confirmed the presence of CYP3A4, CYP3A5, and CYP2E1 in NHEKs. In contrast, although a faint band suggestive of CYP2C9 protein was detected in one NHEK sample, a CYP2C9 message was not detectable. We also examined the ability of chemical inhibitors of cytochromes P450 (aminobenzotriazole and 1-dichloroethylene) and cyclooxygenase (indomethacin) to reduce protein haptenation when NHEKs were incubated with SMX or DDS by either confocal microscopy or ELISA. These inhibitors did not significantly attenuate protein adduction with either SMX or DDS, indicating that cytochromes P450 and cyclooxygenase do not play important roles in the bioactivation of these xenobiotics in NHEKs and thus suggesting the importance of other enzymes in these cells.

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Section: Discussioncontrasting

confidence: 86%

Enzyme-Mediated Protein Haptenation of Dapsone and Sulfamethoxazole in Human Keratinocytes: I. Expression and Role of Cytochromes P450

Vyas

Roychowdhury²,

Khan³

et al. 2006

J Pharmacol Exp Ther

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“…Keratinocytes isolated from discarded, human newborn foreskins were expanded to passage 3 in Complete medium 154-CF (Cascade, Portland, OR; supplemented with kit S-0001-K), adjusted to 0.05 mM Ca 2ϩ as described previously (Du et al, 2006). At Ϸ75% confluence, proliferating cultures were either passaged or media calcium was adjusted to 1.4 mM to initiate cellular differentiation (day 0).…”

Section: Methodsmentioning

confidence: 99%

“…Liver and skin express many of the same P450s, but an important difference is the most abundant hepatic P450s tend to be expressed at relatively low levels in cutaneous tissues and vice versa (Du et al, 2006). This tissue-specificity suggests unique adaptation to different environmental exposures and different P450 protein…”

Section: Introductionmentioning

confidence: 99%

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Differentiation-Specific Factors Modulate Epidermal CYP1–4 Gene Expression in Human Skin in Response to Retinoic Acid and Classic Aryl Hydrocarbon Receptor Ligands

Neis

Ladd

et al. 2006

J Pharmacol Exp Ther

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Human epidermal keratinocytes express subsets of cytochromes P450 (P450) (CYP gene products) that are strongly up-regulated, not regulated, or down-regulated by differentiation-specific factors. We investigated how drug exposure affects epidermal expression of CYP1-4 genes, which encode many drug-metabolizing P450s. Real-time polymerase chain reaction (PCR) assays measured CYP1-4 mRNA levels in epidermal keratinocytes differentiated in vitro in the presence of drug or vehicle for 6 days. We confirmed the spinous phenotype at day 6 by changes in cellular morphology and upregulation of cytokeratin 10 and transglutaminase (TGM)1 mRNA in the differentiating keratinocytes. Effects of drug exposure depended on the influence of differentiation-specific factors in controlling epidermal CYP1-4 expression. CYP2C18, 2C19, 2C9, 2W1, 3A4, and 4B1 are up-regulated by cellular differentiation; mRNA levels for these CYP genes were inhibited in differentiating keratinocytes exposed to retinoic acid and aryl hydrocarbon receptor (AhR) ligands. These same drugs effected Յ2-fold change or even augmented mRNA levels for CYP genes that are not regulated by differentiation (CYP2S1, 2J2, 1B1, 1A1, 1A2, 2E1, and 2D6) and for CYP2U1, which is expressed at highest levels in undifferentiated keratinocytes. The clinically relevant drugs miconazole, dexamethasone, rifampicin, and dapsone had little effect on CYP1-4 mRNA levels under assay conditions. The AhR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin also up-regulated keratinocyte TGM1 mRNA in a concentration-and time-dependent manner. This effect was blocked by the AhR antagonist resveratrol. These findings implicate AhR-dependent up-regulation of TGM1 mRNA in differentiating keratinocytes as one mechanism contributing toward chloracne in humans exposed to toxic levels of dioxin.Structural and biochemical features of skin are specialized toward its major protective functions as a barrier to the host environment. In addition to regulating water loss and body temperature, skin has mechanisms for self-renewal and for surveillance and repair of cell damage due to biological, physical, and chemical exposures. Epidermal keratinocytes constitute the major reservoir of cutaneous cytochromes P450 enzymes (CYP gene products) involved in oxidative metabolism and disposition of endogenous and foreign compounds, including environmental toxins, natural products, and drugs. Most drug-metabolizing cytochromes P450 are members of the CYP1-4 gene families, which number 35 (61%) of the 57 putatively functional CYP genes in humans (Nelson et al., 2004).Major detoxifying organs such as the liver rank among tissues containing the highest concentrations of cytochromes P450; much lower concentrations are found in peripheral tissues such as skin. Liver and skin express many of the same P450s, but an important difference is the most abundant hepatic P450s tend to be expressed at relatively low levels in cutaneous tissues and vice versa (Du et al., 2006). This tissue-specificity suggests unique adaptation to ...

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“…(Ding et al, 1995;Koskela et al, 1999) HepG2 (Yamano et al, 1990), COS-7 (Ding et al, 1995), COS-1 (Oscarson et al, 2002) a 2S1 19q13.1 Skin, liver (also trachea, lung, stomach, small intestine, spleen, colon, urinary tract, nasal cavity, bronchi, bronchioli, tumors) (Rylander et al, 2001;Smith et al, 2003;Saarikoski et al, 2005a,b) S. cerevisiae (Smith et al, 2003), E. coli Wu et al, 2006b;Yoshioka et al, 2006) 4QN (Nishida et al, 2010), several eicosanoid peroxides (Bui et al, 2010) (retinoic acid, carcinogens?) (see also Wu et al, 2006b; b 2U1 4q25 Thymus, brain (Chuang et al, 2004;Karlgren et al, 2004) (also keratinocytes, kidney, heart, ovarian cancer) (Downie et al, 2005;Du et al, 2006) Sf9 insect cells (Chuang et al, 2004) Fatty acids (, -1) (Chuang et al, 2004) 2W1 7q22.3 Tumors (Karlgren et al, 2005(Karlgren et al, , 2006 HEK293 cells (Karlgren et al, 2005), E. coli (Wu et al, 2006b) Several carcinogens (Wu et al, 2006b), 4QN (Nishida et al, 2010) 3A43 7q21 Liver, brain (also kidney, pancreas, prostate) (Domanski et al, 2001;Westlind et al, 2001) E. coli (Domanski et al, 2001), COS-1 cells (Agarwal et al, 2008) Alprazolam (Agarwal et al, 2008), testosterone (Domanski et al, 2001) 4A22 1q33 Liver (Savas et al, 2003) 4F11 19q13.1-2 Liver (Kalsotra et al, 2004), also kidney, heart, skeletal muscle …”

Section: Cytochrome P450 2u1mentioning

confidence: 99%

Orphans in the Human Cytochrome P450 Superfamily: Approaches to Discovering Functions and Relevance in Pharmacology

Guengerich

Cheng

2011

Pharmacol Rev

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Effects of the differentiated keratinocyte phenotype on expression levels of CYP1–4 family genes in human skin cells

Cited by 49 publications

References 49 publications

Enzyme-Mediated Protein Haptenation of Dapsone and Sulfamethoxazole in Human Keratinocytes: I. Expression and Role of Cytochromes P450

Enzyme-Mediated Protein Haptenation of Dapsone and Sulfamethoxazole in Human Keratinocytes: I. Expression and Role of Cytochromes P450

Differentiation-Specific Factors Modulate Epidermal CYP1–4 Gene Expression in Human Skin in Response to Retinoic Acid and Classic Aryl Hydrocarbon Receptor Ligands

Orphans in the Human Cytochrome P450 Superfamily: Approaches to Discovering Functions and Relevance in Pharmacology

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