Study of cytochrome P4502E1 mRNA level of mononuclear cells in patients with alcoholic liver disease

Background: Cytochrome P-4502E1 (CYP2E1) is an important enzyme because of its unique ability to convert many substrates to cytotoxins. The increased production of reactive intermediates by elevated enzyme concentrations lead s to various pathological conditions. Therefore, it is important to detect induced CYP2E1 levels in alcoholic individuals to avoid xenobiotic-promoted liver injury. In the present investigation, we detected CYP2E1 mRNA levels of mononuclear cells obtained from 10 ml of blood by using competitive pol ymerase chain reaction (PCR) method. Methods: Mononuclear cells were obtained from healthy individuals who did a nd did not drink habitually and patients with alcoholic liver disease (ALD) . Complementary DNA synthesis was performed with RNA obtained from mononucl ear cells by reverse transcription-PCR. Competitive PCR of CYP2E1 was perfo rmed with the sense (5 ' -CTGCAACGTCATA-GCCGACA-3 ') and antisense (5 ' -TC CATTTCCACGAGCAGGCA-3 ') primer and competitor DNA. Competitive PCR of beta -actin also was performed. Electrophoresis was scanned. and each band was d igitized. The concentration of CYP2E1 and B-actin mRNA was calculated from the ratio of competitor DNA. Results: In healthy individuals who did and did not drink habitually, CYP2E 1 mRNA levels were 10(33) coppies/mul RNA and 10(1.7) copies/mul RNA, respe ctively. In actively drinking patients with ALD, CYP2E1 mRNA levels were 10 (3.5) copies/mul RNA, but those levels decreased to 10(1.7) copies/mul RNA after 4 days of abstinence. No significant difference was observed in CYP2E 1 mRNA levels between alcoholic fibrosis and cirrhosis. As control, we meas ured beta -actin mRNA levels in mononuclear cells in all samples. The mean value of beta -actin mRNA was 10(4.3) Copies/mul RNA in all cases, which in cluded patients with ALD. Conclusions: The results demonstrated that it is possible to measure the CY P2E1 mRNA levels of mononuclear cells in a 10 ml blood sample. The CYP2E1 m RNA level in mononuclear cells increases during drinking and decreases in a bstinence for a short period of 3 to 4 days. It is concluded that CYP2E1 mR NA level may be used as an effective marker for alcoholic intake.