Limits...
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.

Show MeSH

Related in: MedlinePlus

The effect of arachidonic acid (AA; 60 μM; 24 h) on viability of HepG2 and HepG2 cells overexpressing CYP2E1. Cell viability was assessed with MTT test. The same treatment was done in cells grown for 1 week with medium supplemented with 1 mM acetate or 100 mM ethanol. Results from six independent experiments are expressed as mean percentages of viable cells ± SD and control was taken as 100 %. Statistically significant difference from the control is indicated as *P < 0.05 and ##P < 0.01—from ethanol-treated cells
© Copyright Policy - OpenAccess
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3927057&req=5

Fig4: The effect of arachidonic acid (AA; 60 μM; 24 h) on viability of HepG2 and HepG2 cells overexpressing CYP2E1. Cell viability was assessed with MTT test. The same treatment was done in cells grown for 1 week with medium supplemented with 1 mM acetate or 100 mM ethanol. Results from six independent experiments are expressed as mean percentages of viable cells ± SD and control was taken as 100 %. Statistically significant difference from the control is indicated as *P < 0.05 and ##P < 0.01—from ethanol-treated cells

Mentions: Cytotoxicity of AA applied for 24 h to the cells was also tested by MTT test (Fig. 4). In HepG2 cells overexpressing CYP2E1, AA significantly decreased cell viability (by about 37 %; P < 0.01). This effect was observed neither in naïve HepG2 cells nor in CYP2E1 overexpressing cells grown for 1 week with ethanol or acetate. Cytotoxicity of AA in transduced cells was abolished not only by CYP2E1 inhibitor-4MP, but also by cell pretreatment with acetate or HDAC inhibitor—TSA (Table 1). In cells grown with ethanol prior to AA treatment, cytotoxicity of AA was slightly lower than in naïve cells but still significant (P < 0.01) comparing to ethanol-only treated cells.Fig. 4


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)

The effect of arachidonic acid (AA; 60 μM; 24 h) on viability of HepG2 and HepG2 cells overexpressing CYP2E1. Cell viability was assessed with MTT test. The same treatment was done in cells grown for 1 week with medium supplemented with 1 mM acetate or 100 mM ethanol. Results from six independent experiments are expressed as mean percentages of viable cells ± SD and control was taken as 100 %. Statistically significant difference from the control is indicated as *P < 0.05 and ##P < 0.01—from ethanol-treated cells
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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

Fig4: The effect of arachidonic acid (AA; 60 μM; 24 h) on viability of HepG2 and HepG2 cells overexpressing CYP2E1. Cell viability was assessed with MTT test. The same treatment was done in cells grown for 1 week with medium supplemented with 1 mM acetate or 100 mM ethanol. Results from six independent experiments are expressed as mean percentages of viable cells ± SD and control was taken as 100 %. Statistically significant difference from the control is indicated as *P < 0.05 and ##P < 0.01—from ethanol-treated cells
Mentions: Cytotoxicity of AA applied for 24 h to the cells was also tested by MTT test (Fig. 4). In HepG2 cells overexpressing CYP2E1, AA significantly decreased cell viability (by about 37 %; P < 0.01). This effect was observed neither in naïve HepG2 cells nor in CYP2E1 overexpressing cells grown for 1 week with ethanol or acetate. Cytotoxicity of AA in transduced cells was abolished not only by CYP2E1 inhibitor-4MP, but also by cell pretreatment with acetate or HDAC inhibitor—TSA (Table 1). In cells grown with ethanol prior to AA treatment, cytotoxicity of AA was slightly lower than in naïve cells but still significant (P < 0.01) comparing to ethanol-only treated cells.Fig. 4

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