Dopamine D1 receptor activation induces dehydroepiandrosterone sulfotransferase (SULT2A1) in HepG2 cells
Abstract
Jiao-jiao XU1, 3, Si-yuan WANG1, Ye CHEN1, Guang-ping CHEN2, Zai-quan LI4, Xue-yan SHAO1, Liang LI1, Wei LU1, Tian-yan ZHOU1, *
1Department of Pharmaceutics, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China; 2Department of Physiological Sciences, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA; 3Laboratory of Physicochemical Research, Department of Physicochemical & Toxicology, Zhejiang Provincial Centre for Disease Control and Prevention, Hangzhou 310051, China; 4Department of Pathology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
Aim: Dopamine receptors are present in the nervous system and also widely distributed in the periphery. The aim of this study was to investigate the role of D1 subtype dopamine receptors (DRD1) in the regulation of dehydroepiandrosterone sulfotransferase (SULT2A1) in HepG2 cells.
Methods: HepG2 cells were treated with DRD1 agonists with or without DRD1 antagonist for 9 d. DRD1 and SULT2A1 mRNA expression, protein expression, and SULT2A1 activity were detected using RT-PCR, Western blotting and HPLC, respectively. The level of cAMP was measured using a commercial kit.
Results: All the 5 DR subtypes (DRD1–DRD5) were found to be expressed in HepG2 cells. Treatment of HepG2 cells with the specific DRD1 agonists SKF82958 (2.5 μmol/L) or SKF38393 (5 and 50 μmol/L) significantly increased the mRNA and protein expression of both DRD1 and SULT2A1, and increased SULT2A1 activity and cAMP levels. These effects were partially blocked by co-treatment with the specific DRD1 antagonist SCH23390 (2.5 μmol/L). In addition, transfection of HepG2 cells with DRD1-specific siRNAs decreased DRD1 mRNA expression by 40%, which resulted in the reduction of SULT2A1 mRNA expression by 60%, protein expression by 40%, and enzyme activity by 20%.
Conclusion: DRD1 activation upregulates DRD1 and SULT2A1 expression and SULT2A1 activity in HepG2 cells, suggesting that the DRD1 subtype may be involved in the metabolism of drugs and xenobiotics through regulating SULT2A1.
Keywords: dopamine; D1 receptor; dehydroepiandrosterone sulfotransferase (SULT2A1); drug-metabolizing enzyme; SKF82958; SKF38393; SCH23390; siRNA; HepG2 cell
This work was supported by the National Natural Science Foundation of China (Grant 81072699) and by the Scientific Research Foundation for the Returned Overseas Chinese Scholars by the State Education Ministry (Grant [2011]508). The authors deeply appreciate the generous gift of human SULT2A1 antibody from Dr David RINGER of the American Cancer Society.
* To whom correspondence should be addressed.
E-mail tianyanzhou@bjmu.edu.cn
Received 2013-10-06 Accepted 2014-02-20
Keywords:
1Department of Pharmaceutics, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China; 2Department of Physiological Sciences, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA; 3Laboratory of Physicochemical Research, Department of Physicochemical & Toxicology, Zhejiang Provincial Centre for Disease Control and Prevention, Hangzhou 310051, China; 4Department of Pathology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
Aim: Dopamine receptors are present in the nervous system and also widely distributed in the periphery. The aim of this study was to investigate the role of D1 subtype dopamine receptors (DRD1) in the regulation of dehydroepiandrosterone sulfotransferase (SULT2A1) in HepG2 cells.
Methods: HepG2 cells were treated with DRD1 agonists with or without DRD1 antagonist for 9 d. DRD1 and SULT2A1 mRNA expression, protein expression, and SULT2A1 activity were detected using RT-PCR, Western blotting and HPLC, respectively. The level of cAMP was measured using a commercial kit.
Results: All the 5 DR subtypes (DRD1–DRD5) were found to be expressed in HepG2 cells. Treatment of HepG2 cells with the specific DRD1 agonists SKF82958 (2.5 μmol/L) or SKF38393 (5 and 50 μmol/L) significantly increased the mRNA and protein expression of both DRD1 and SULT2A1, and increased SULT2A1 activity and cAMP levels. These effects were partially blocked by co-treatment with the specific DRD1 antagonist SCH23390 (2.5 μmol/L). In addition, transfection of HepG2 cells with DRD1-specific siRNAs decreased DRD1 mRNA expression by 40%, which resulted in the reduction of SULT2A1 mRNA expression by 60%, protein expression by 40%, and enzyme activity by 20%.
Conclusion: DRD1 activation upregulates DRD1 and SULT2A1 expression and SULT2A1 activity in HepG2 cells, suggesting that the DRD1 subtype may be involved in the metabolism of drugs and xenobiotics through regulating SULT2A1.
Keywords: dopamine; D1 receptor; dehydroepiandrosterone sulfotransferase (SULT2A1); drug-metabolizing enzyme; SKF82958; SKF38393; SCH23390; siRNA; HepG2 cell
This work was supported by the National Natural Science Foundation of China (Grant 81072699) and by the Scientific Research Foundation for the Returned Overseas Chinese Scholars by the State Education Ministry (Grant [2011]508). The authors deeply appreciate the generous gift of human SULT2A1 antibody from Dr David RINGER of the American Cancer Society.
* To whom correspondence should be addressed.
E-mail tianyanzhou@bjmu.edu.cn
Received 2013-10-06 Accepted 2014-02-20