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Relationship between mRNA Levels Quantified by Reverse Transcription-Competitive PCR and Metabolic Activity of CYP3A4 and CYP2E1 in Human Liver
Reverse transcription-competitive polymerase chain reaction is a powerful and sensitive tool for quantifying the absolute amount of mRNA. Using this method with β-actin as the standard, we measured the mRNA level of CYP3A and CYP2E1 isoforms in human livers. We also determined the metabolic activiti...
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Published in: | Biochemical and biophysical research communications 1999-08, Vol.262 (2), p.499-503 |
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creator | Sumida, Akihiko Kinoshita, Kayoko Fukuda, Tsuyoshi Matsuda, Hideyasu Yamamoto, Isamu Inaba, Tadanobu Azuma, Junichi |
description | Reverse transcription-competitive polymerase chain reaction is a powerful and sensitive tool for quantifying the absolute amount of mRNA. Using this method with β-actin as the standard, we measured the mRNA level of CYP3A and CYP2E1 isoforms in human livers. We also determined the metabolic activities for both CYP isoforms. The absolute amounts of CYP3A4 and CYP2E1 mRNA in 15 liver tissues ranged from 15 to 5127 and 15163 to 69289 copies/104 copies of β-actin (341- & 3.6-fold), respectively. The testosterone 6β-hydroxylation for CYP3A4 and chlorzoxazone 6-hydroxylation activity for CYP2E1 ranged from 30 to 505 pmol/mg/min (16-fold) and from 0.59 to 2.73 nmol/mg/ml (3.6-fold), respectively. The correlation between the mRNA level and activity of CYP3A4 was significant (r = 0.94), while there was no significant correlation for CYP2E1 (r = 0.04). In conclusion, we observed a significant correlation between enzyme activity and mRNA expression for CYP3A4 but not for CYP2E1. This fact indicates that CYP2E1, in addition to being less variable between individuals than CYP3A4, differs in its regulation mechanism. |
doi_str_mv | 10.1006/bbrc.1999.1233 |
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Using this method with β-actin as the standard, we measured the mRNA level of CYP3A and CYP2E1 isoforms in human livers. We also determined the metabolic activities for both CYP isoforms. The absolute amounts of CYP3A4 and CYP2E1 mRNA in 15 liver tissues ranged from 15 to 5127 and 15163 to 69289 copies/104 copies of β-actin (341- & 3.6-fold), respectively. The testosterone 6β-hydroxylation for CYP3A4 and chlorzoxazone 6-hydroxylation activity for CYP2E1 ranged from 30 to 505 pmol/mg/min (16-fold) and from 0.59 to 2.73 nmol/mg/ml (3.6-fold), respectively. The correlation between the mRNA level and activity of CYP3A4 was significant (r = 0.94), while there was no significant correlation for CYP2E1 (r = 0.04). In conclusion, we observed a significant correlation between enzyme activity and mRNA expression for CYP3A4 but not for CYP2E1. This fact indicates that CYP2E1, in addition to being less variable between individuals than CYP3A4, differs in its regulation mechanism.</description><identifier>ISSN: 0006-291X</identifier><identifier>EISSN: 1090-2104</identifier><identifier>DOI: 10.1006/bbrc.1999.1233</identifier><identifier>PMID: 10462503</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Adolescent ; Adult ; CYP2E1 ; CYP3A4 ; Cytochrome P-450 CYP2E1 - analysis ; Cytochrome P-450 CYP2E1 - genetics ; Cytochrome P-450 CYP3A ; Cytochrome P-450 Enzyme System - analysis ; Cytochrome P-450 Enzyme System - genetics ; European Continental Ancestry Group ; Female ; human liver ; Humans ; Male ; metabolic activity ; Microsomes, Liver - enzymology ; Middle Aged ; Mixed Function Oxygenases - analysis ; Mixed Function Oxygenases - genetics ; mRNA ; Reverse Transcriptase Polymerase Chain Reaction ; reverse transcription-competitive PCR ; RNA, Messenger - analysis</subject><ispartof>Biochemical and biophysical research communications, 1999-08, Vol.262 (2), p.499-503</ispartof><rights>1999 Academic Press</rights><rights>Copyright 1999 Academic Press.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c437t-b07b43f6045677002c0bf8ce409d4be7c0eec25a3c6f11c9b2089501256d9d73</citedby><cites>FETCH-LOGICAL-c437t-b07b43f6045677002c0bf8ce409d4be7c0eec25a3c6f11c9b2089501256d9d73</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/10462503$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Sumida, Akihiko</creatorcontrib><creatorcontrib>Kinoshita, Kayoko</creatorcontrib><creatorcontrib>Fukuda, Tsuyoshi</creatorcontrib><creatorcontrib>Matsuda, Hideyasu</creatorcontrib><creatorcontrib>Yamamoto, Isamu</creatorcontrib><creatorcontrib>Inaba, Tadanobu</creatorcontrib><creatorcontrib>Azuma, Junichi</creatorcontrib><title>Relationship between mRNA Levels Quantified by Reverse Transcription-Competitive PCR and Metabolic Activity of CYP3A4 and CYP2E1 in Human Liver</title><title>Biochemical and biophysical research communications</title><addtitle>Biochem Biophys Res Commun</addtitle><description>Reverse transcription-competitive polymerase chain reaction is a powerful and sensitive tool for quantifying the absolute amount of mRNA. Using this method with β-actin as the standard, we measured the mRNA level of CYP3A and CYP2E1 isoforms in human livers. We also determined the metabolic activities for both CYP isoforms. The absolute amounts of CYP3A4 and CYP2E1 mRNA in 15 liver tissues ranged from 15 to 5127 and 15163 to 69289 copies/104 copies of β-actin (341- & 3.6-fold), respectively. The testosterone 6β-hydroxylation for CYP3A4 and chlorzoxazone 6-hydroxylation activity for CYP2E1 ranged from 30 to 505 pmol/mg/min (16-fold) and from 0.59 to 2.73 nmol/mg/ml (3.6-fold), respectively. The correlation between the mRNA level and activity of CYP3A4 was significant (r = 0.94), while there was no significant correlation for CYP2E1 (r = 0.04). In conclusion, we observed a significant correlation between enzyme activity and mRNA expression for CYP3A4 but not for CYP2E1. This fact indicates that CYP2E1, in addition to being less variable between individuals than CYP3A4, differs in its regulation mechanism.</description><subject>Adolescent</subject><subject>Adult</subject><subject>CYP2E1</subject><subject>CYP3A4</subject><subject>Cytochrome P-450 CYP2E1 - analysis</subject><subject>Cytochrome P-450 CYP2E1 - genetics</subject><subject>Cytochrome P-450 CYP3A</subject><subject>Cytochrome P-450 Enzyme System - analysis</subject><subject>Cytochrome P-450 Enzyme System - genetics</subject><subject>European Continental Ancestry Group</subject><subject>Female</subject><subject>human liver</subject><subject>Humans</subject><subject>Male</subject><subject>metabolic activity</subject><subject>Microsomes, Liver - enzymology</subject><subject>Middle Aged</subject><subject>Mixed Function Oxygenases - analysis</subject><subject>Mixed Function Oxygenases - genetics</subject><subject>mRNA</subject><subject>Reverse Transcriptase Polymerase Chain Reaction</subject><subject>reverse transcription-competitive PCR</subject><subject>RNA, Messenger - analysis</subject><issn>0006-291X</issn><issn>1090-2104</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1999</creationdate><recordtype>article</recordtype><recordid>eNqFkT2PEzEQhi0E4nIHLSVyRbdh7PV64zKKDg4pwBGlgMqyvbPCaL-wvUH5FfxlvJcraBDVWDPP-xZ-CHnFYM0A5Ftrg1szpdSa8bJ8QlYMFBScgXhKVpCJgiv29Ypcx_gDgDEh1XNylc-SV1CuyO8Ddib5cYjf_UQtpl-IA-0Pn7Z0jyfsIv0ymyH51mND7Zke8jJEpMdghuiCn5ZssRv7CZNP_oT0fnegZmjoR0zGjp13dOvywaczHVu6-3ZfbsUDkJ_8llE_0Lu5NwPd53R4QZ61pov48nHekOO72-Purth_fv9ht90XTpR1KizUVpStBFHJugbgDmy7cShANcJi7QDR8cqUTraMOWU5bFQFjFeyUU1d3pA3l9opjD9njEn3PjrsOjPgOEct84_WUqj_gqzmkotaZHB9AV0YYwzY6in43oSzZqAXVXpRpRdVelGVA68fm2fbY_MXfnGTgc0FyBbw5DHo6DwODhsf0CXdjP5f3X8AKISiRQ</recordid><startdate>19990827</startdate><enddate>19990827</enddate><creator>Sumida, Akihiko</creator><creator>Kinoshita, Kayoko</creator><creator>Fukuda, Tsuyoshi</creator><creator>Matsuda, Hideyasu</creator><creator>Yamamoto, Isamu</creator><creator>Inaba, Tadanobu</creator><creator>Azuma, Junichi</creator><general>Elsevier Inc</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7TM</scope><scope>7X8</scope></search><sort><creationdate>19990827</creationdate><title>Relationship between mRNA Levels Quantified by Reverse Transcription-Competitive PCR and Metabolic Activity of CYP3A4 and CYP2E1 in Human Liver</title><author>Sumida, Akihiko ; Kinoshita, Kayoko ; Fukuda, Tsuyoshi ; Matsuda, Hideyasu ; Yamamoto, Isamu ; Inaba, Tadanobu ; Azuma, Junichi</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c437t-b07b43f6045677002c0bf8ce409d4be7c0eec25a3c6f11c9b2089501256d9d73</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1999</creationdate><topic>Adolescent</topic><topic>Adult</topic><topic>CYP2E1</topic><topic>CYP3A4</topic><topic>Cytochrome P-450 CYP2E1 - analysis</topic><topic>Cytochrome P-450 CYP2E1 - genetics</topic><topic>Cytochrome P-450 CYP3A</topic><topic>Cytochrome P-450 Enzyme System - analysis</topic><topic>Cytochrome P-450 Enzyme System - genetics</topic><topic>European Continental Ancestry Group</topic><topic>Female</topic><topic>human liver</topic><topic>Humans</topic><topic>Male</topic><topic>metabolic activity</topic><topic>Microsomes, Liver - enzymology</topic><topic>Middle Aged</topic><topic>Mixed Function Oxygenases - analysis</topic><topic>Mixed Function Oxygenases - genetics</topic><topic>mRNA</topic><topic>Reverse Transcriptase Polymerase Chain Reaction</topic><topic>reverse transcription-competitive PCR</topic><topic>RNA, Messenger - analysis</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Sumida, Akihiko</creatorcontrib><creatorcontrib>Kinoshita, Kayoko</creatorcontrib><creatorcontrib>Fukuda, Tsuyoshi</creatorcontrib><creatorcontrib>Matsuda, Hideyasu</creatorcontrib><creatorcontrib>Yamamoto, Isamu</creatorcontrib><creatorcontrib>Inaba, Tadanobu</creatorcontrib><creatorcontrib>Azuma, Junichi</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Biochemical and biophysical research communications</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Sumida, Akihiko</au><au>Kinoshita, Kayoko</au><au>Fukuda, Tsuyoshi</au><au>Matsuda, Hideyasu</au><au>Yamamoto, Isamu</au><au>Inaba, Tadanobu</au><au>Azuma, Junichi</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Relationship between mRNA Levels Quantified by Reverse Transcription-Competitive PCR and Metabolic Activity of CYP3A4 and CYP2E1 in Human Liver</atitle><jtitle>Biochemical and biophysical research communications</jtitle><addtitle>Biochem Biophys Res Commun</addtitle><date>1999-08-27</date><risdate>1999</risdate><volume>262</volume><issue>2</issue><spage>499</spage><epage>503</epage><pages>499-503</pages><issn>0006-291X</issn><eissn>1090-2104</eissn><abstract>Reverse transcription-competitive polymerase chain reaction is a powerful and sensitive tool for quantifying the absolute amount of mRNA. Using this method with β-actin as the standard, we measured the mRNA level of CYP3A and CYP2E1 isoforms in human livers. We also determined the metabolic activities for both CYP isoforms. The absolute amounts of CYP3A4 and CYP2E1 mRNA in 15 liver tissues ranged from 15 to 5127 and 15163 to 69289 copies/104 copies of β-actin (341- & 3.6-fold), respectively. The testosterone 6β-hydroxylation for CYP3A4 and chlorzoxazone 6-hydroxylation activity for CYP2E1 ranged from 30 to 505 pmol/mg/min (16-fold) and from 0.59 to 2.73 nmol/mg/ml (3.6-fold), respectively. The correlation between the mRNA level and activity of CYP3A4 was significant (r = 0.94), while there was no significant correlation for CYP2E1 (r = 0.04). In conclusion, we observed a significant correlation between enzyme activity and mRNA expression for CYP3A4 but not for CYP2E1. This fact indicates that CYP2E1, in addition to being less variable between individuals than CYP3A4, differs in its regulation mechanism.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>10462503</pmid><doi>10.1006/bbrc.1999.1233</doi><tpages>5</tpages></addata></record> |
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subjects | Adolescent Adult CYP2E1 CYP3A4 Cytochrome P-450 CYP2E1 - analysis Cytochrome P-450 CYP2E1 - genetics Cytochrome P-450 CYP3A Cytochrome P-450 Enzyme System - analysis Cytochrome P-450 Enzyme System - genetics European Continental Ancestry Group Female human liver Humans Male metabolic activity Microsomes, Liver - enzymology Middle Aged Mixed Function Oxygenases - analysis Mixed Function Oxygenases - genetics mRNA Reverse Transcriptase Polymerase Chain Reaction reverse transcription-competitive PCR RNA, Messenger - analysis |
title | Relationship between mRNA Levels Quantified by Reverse Transcription-Competitive PCR and Metabolic Activity of CYP3A4 and CYP2E1 in Human Liver |
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