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Histone acetylation and arachidonic acid cytotoxicity in HepG2 cells overexpressing CYP2E1.

Holownia A, Mroz RM, Wielgat P, Jakubow P, Jablonski J, Sulek J, Braszko JJ - Naunyn Schmiedebergs Arch. Pharmacol. (2013)

Bottom Line: Arachidonic acid altered cell proliferation and was cytotoxic mostly to cells engineered to overexpress CYP2E1 but both effects were significantly lower in cells pretreated with ethanol or acetate.Cytotoxicity was also significantly decreased by 4-methylpyrazole--a CYP2E1 inhibitor and by trichostatin--an inhibitor of histone deacetylases.In cells pretreated with acetate or ethanol, the oxidative stress induced by arachidonic acid was also significantly lower.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Pharmacology, Medical University of Bialystok, Waszyngtona 15A, 15-274, Bialystok, Poland, Holow_sinai@hotmail.com.

ABSTRACT
The aim of this work was to assess the role of ethanol-derived acetate and acetate-mediated histone acetylation in arachidonic acid-induced stress in HepG2 cells and cells overexpressing CYP2E1. Cells were grown for 7 days with 1 mM sodium acetate or 100 mM ethanol; their acetylated histone proteins and histone deacetylase 2 expression was quantified using Western blot. Ethanol- or acetate-pretreated cells were also treated for 24 h with 60 μM arachidonic acid to induce oxidative stress. Cytotoxicity was estimated by lactate dehydrogenase release, 3-[4,5-dimethylthiazolyl-2] 2,5-diphenyltetrazolium bromide test, and by DNA damage, while oxidative stress was quantified using dichlorofluorescein diacetate. Cells grown with ethanol or acetate had increased acetylated histone H3 levels in both cell types and elevated acetylated histone H4 levels in cells overexpressing CYP2E1 but not in naïve cells. In cells overexpressing CYP2E1 grown with ethanol, expression of histone deacetylase 2 was reduced by about 40 %. Arachidonic acid altered cell proliferation and was cytotoxic mostly to cells engineered to overexpress CYP2E1 but both effects were significantly lower in cells pretreated with ethanol or acetate. Cytotoxicity was also significantly decreased by 4-methylpyrazole--a CYP2E1 inhibitor and by trichostatin--an inhibitor of histone deacetylases. In cells pretreated with acetate or ethanol, the oxidative stress induced by arachidonic acid was also significantly lower. Our data indicate that histone hyperacetylation may in some extent protect the cells against oxidative stress. It is possible that acetate may act as an antioxidant at histone level. This mechanism may be relevant to alcohol-induced liver injury.

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Related in: MedlinePlus

Acetylated histone H3 (AcH3) and histone H4 (AcH4) in HepG2 cells and HepG2 cells overexpressing CYP2E1 grown for 1 week in culture medium supplemented with 1 mM acetate or 100 mM ethanol. Figures represent mean results ± SD. Data are relative values (digitized relative density) expressed as percentages over untreated cells taken as 100 %. Representative Western blot pictures are included. Statistically significant differences from controls are indicated as: *P < 0.05 or **P < 0.01
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Fig1: Acetylated histone H3 (AcH3) and histone H4 (AcH4) in HepG2 cells and HepG2 cells overexpressing CYP2E1 grown for 1 week in culture medium supplemented with 1 mM acetate or 100 mM ethanol. Figures represent mean results ± SD. Data are relative values (digitized relative density) expressed as percentages over untreated cells taken as 100 %. Representative Western blot pictures are included. Statistically significant differences from controls are indicated as: *P < 0.05 or **P < 0.01

Mentions: Figure 1 shows acetylated histone H3 (AcH3) and acetylated histone H4 (AcH4) in HepG2 cells and HepG2 cells overexpressing CYP2E1 grown for 7 days in culture medium supplemented with 1 mM acetate or 100 mM ethanol. Both treatments increased AcH3 in transduced and nontransduced cells, while AcH4 levels were significantly increased in cells overexpressing CYP2E1 grown with acetate or ethanol, but not in naïve HepG2 cells. The highest expression of acetylated histones (increase by 73 %; P < 0.01) was detected in H3 protein in cells overexpressing CYP2E1 grown with ethanol and slightly lower AcH3 levels (increase by 53 %; P < 0.05) were found in the same cell type grown with acetate. Lower but statistically significant increases were observed in AcH4 levels, in cells engineered to overexpress CYP2E1 treated with ethanol or acetate (increase by about 40 %; P < 0.05 in both groups), while in nontransduced cells AcH4 levels were not affected.Fig. 1


Histone acetylation and arachidonic acid cytotoxicity in HepG2 cells overexpressing CYP2E1.

Holownia A, Mroz RM, Wielgat P, Jakubow P, Jablonski J, Sulek J, Braszko JJ - Naunyn Schmiedebergs Arch. Pharmacol. (2013)

Acetylated histone H3 (AcH3) and histone H4 (AcH4) in HepG2 cells and HepG2 cells overexpressing CYP2E1 grown for 1 week in culture medium supplemented with 1 mM acetate or 100 mM ethanol. Figures represent mean results ± SD. Data are relative values (digitized relative density) expressed as percentages over untreated cells taken as 100 %. Representative Western blot pictures are included. Statistically significant differences from controls are indicated as: *P < 0.05 or **P < 0.01
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3927057&req=5

Fig1: Acetylated histone H3 (AcH3) and histone H4 (AcH4) in HepG2 cells and HepG2 cells overexpressing CYP2E1 grown for 1 week in culture medium supplemented with 1 mM acetate or 100 mM ethanol. Figures represent mean results ± SD. Data are relative values (digitized relative density) expressed as percentages over untreated cells taken as 100 %. Representative Western blot pictures are included. Statistically significant differences from controls are indicated as: *P < 0.05 or **P < 0.01
Mentions: Figure 1 shows acetylated histone H3 (AcH3) and acetylated histone H4 (AcH4) in HepG2 cells and HepG2 cells overexpressing CYP2E1 grown for 7 days in culture medium supplemented with 1 mM acetate or 100 mM ethanol. Both treatments increased AcH3 in transduced and nontransduced cells, while AcH4 levels were significantly increased in cells overexpressing CYP2E1 grown with acetate or ethanol, but not in naïve HepG2 cells. The highest expression of acetylated histones (increase by 73 %; P < 0.01) was detected in H3 protein in cells overexpressing CYP2E1 grown with ethanol and slightly lower AcH3 levels (increase by 53 %; P < 0.05) were found in the same cell type grown with acetate. Lower but statistically significant increases were observed in AcH4 levels, in cells engineered to overexpress CYP2E1 treated with ethanol or acetate (increase by about 40 %; P < 0.05 in both groups), while in nontransduced cells AcH4 levels were not affected.Fig. 1

Bottom Line: Arachidonic acid altered cell proliferation and was cytotoxic mostly to cells engineered to overexpress CYP2E1 but both effects were significantly lower in cells pretreated with ethanol or acetate.Cytotoxicity was also significantly decreased by 4-methylpyrazole--a CYP2E1 inhibitor and by trichostatin--an inhibitor of histone deacetylases.In cells pretreated with acetate or ethanol, the oxidative stress induced by arachidonic acid was also significantly lower.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Pharmacology, Medical University of Bialystok, Waszyngtona 15A, 15-274, Bialystok, Poland, Holow_sinai@hotmail.com.

ABSTRACT
The aim of this work was to assess the role of ethanol-derived acetate and acetate-mediated histone acetylation in arachidonic acid-induced stress in HepG2 cells and cells overexpressing CYP2E1. Cells were grown for 7 days with 1 mM sodium acetate or 100 mM ethanol; their acetylated histone proteins and histone deacetylase 2 expression was quantified using Western blot. Ethanol- or acetate-pretreated cells were also treated for 24 h with 60 μM arachidonic acid to induce oxidative stress. Cytotoxicity was estimated by lactate dehydrogenase release, 3-[4,5-dimethylthiazolyl-2] 2,5-diphenyltetrazolium bromide test, and by DNA damage, while oxidative stress was quantified using dichlorofluorescein diacetate. Cells grown with ethanol or acetate had increased acetylated histone H3 levels in both cell types and elevated acetylated histone H4 levels in cells overexpressing CYP2E1 but not in naïve cells. In cells overexpressing CYP2E1 grown with ethanol, expression of histone deacetylase 2 was reduced by about 40 %. Arachidonic acid altered cell proliferation and was cytotoxic mostly to cells engineered to overexpress CYP2E1 but both effects were significantly lower in cells pretreated with ethanol or acetate. Cytotoxicity was also significantly decreased by 4-methylpyrazole--a CYP2E1 inhibitor and by trichostatin--an inhibitor of histone deacetylases. In cells pretreated with acetate or ethanol, the oxidative stress induced by arachidonic acid was also significantly lower. Our data indicate that histone hyperacetylation may in some extent protect the cells against oxidative stress. It is possible that acetate may act as an antioxidant at histone level. This mechanism may be relevant to alcohol-induced liver injury.

Show MeSH
Related in: MedlinePlus