Establishment of cytochrome P450 3A4 and glutathione S-transferase A1-transfected human hepatoma cell line and functional analysis.
This study aimed to enhance the drug metabolism function of the human hepatoma cell line C3A and to explore the related significance for patients with severe liver disease. The important liver phase I and phase II drug metabolism enzymes, cytochrome P450 3A4 (CYP 3A4) and glutathione S-transferase A1 (GST A1), were constructed into a double expression vector and then transfected into C3A cells. Furthermore, in order to increase the expression of CYP 3A4 and GST A1, they were optimized according to human optimal codons. Another double-expression vector, pBudCE4.1-optimized CYP 3A4-optimized GST A1, was constructed and then transfected into C3A to establish a stable cell line. The drug metabolism function of C3A was evaluated. Sequence determination and analysis results showed that the recombinant plasmid pBudCE4.1-CYP 3A4-GST A1 met the application standard and its transfection was successful. The expression and activity of CYP 3A4 and GST A1 in unoptimized C3A cells were higher than those in blank C3A cells. Unoptimized C3A had a better drug metabolism function. Although some C3A cells transfected with pBudCE4.1-optimized CYP 3A4-optimized GST A1 survived, they grew slowly, and were therefore not applicable in clinical practice. Unoptimized C3A is superior to blank C3A in drug metabolism, and could be applied in the bioartificial liver support system as a new material.
Non-Infectious Liver Diseases Diagnosis and Treatment Center, 302 Military Hospital, Beijing, China.
B X
BX
Chang
Liver Failure Treatment and Research Center, 302 Military Hospital, Beijing, China.
S L
SL
You
Liver Failure Treatment and Research Center, 302 Military Hospital, Beijing, China.
H L
HL
Liu
Experimental Technologies and Research Center, 302 Military Hospital, Beijing, China.
P Y
PY
Mao
Liver Failure Treatment and Research Center, 302 Military Hospital, Beijing, China xinshaojiecn@163.com.
S J
SJ
Xin
D000779:Anesthetics, Local / Q000008:administration & dosage / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics; D051544:Cytochrome P-450 CYP3A / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008012:Lidocaine / Q000008:administration & dosage / Q000378:metabolism; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D004364:Pharmaceutical Preparations / Q000008:administration & dosage / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D000779:Anesthetics, Local; D007527:Isoenzymes; D004364:Pharmaceutical Preparations; D008012:Lidocaine; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D005982:Glutathione Transferase; C119995:glutathione S-transferase alpha