TY - JOUR
T1 - Dual-color bioluminescent assay using infected HepG2 cells sheds new light on Chlamydia pneumoniae and human cytomegalovirus effects on human cholesterol 7α-hydroxylase (CYP7A1) transcription
AU - Michelini, Elisa
AU - Donati, Manuela
AU - Aldini, Rita
AU - Cevenini, Luca
AU - Mezzanotte, Laura
AU - Nardini, Paola
AU - Foschi, Claudio
AU - Zvi, Ido Ben
AU - Cevenini, Monica
AU - Montagnani, Marco
AU - Marangoni, Antonella
AU - Roda, Aldo
AU - Cevenini, Roberto
N1 - Funding Information:
We are very grateful to Bruce Branchini (Connecticut College) for providing the complementary DNA (cDNA) encoding for PpyRE-TS. This work was supported by grants PRIN 2009MB4AYL and FIRB 2008 RBFR08SZTR from the Ministry for Education, Universities, and Research (MIUR, Italy).
PY - 2012/11/1
Y1 - 2012/11/1
N2 - Chlamydia pneumoniae and human cytomegalovirus (HCMV) are intracellular pathogens able to infect hepatocytes, causing an increase in serum triglycerides and cholesterol levels due to the production of inflammatory cytokines. We investigated whether these pathogens could interfere with cholesterol metabolism by affecting activity of hepatic cholesterol 7α-hydroxylase (CYP7A1) promoter. CYP7A1 is the rate-limiting enzyme responsible for conversion of cholesterol to bile acids, which represents the main route of cholesterol catabolism. A straightforward dual-reporter bioluminescent assay was developed to simultaneously monitor CYP7A1 transcriptional regulation and cell viability in infected human hepatoblastoma HepG2 cells. C. pneumoniae and HCMV infection significantly decreased CYP7A1 promoter activity in a dose-dependent manner, with maximal inhibitions of 33 ± 10% and 32 ± 4%, respectively, at a multiplicity of infection of 1. To support in vitro experiments, serum cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides and glucose levels were also measured in Balb/c mice infected with C. pneumoniae. Serum cholesterol and triglycerides also increased in infected mice compared with controls. Although further investigation is required, this work presents the first experimental evidence that C. pneumoniae and HCMV inhibit CYP7A1 gene transcription in the cultured human hepatoblastoma cell line.
AB - Chlamydia pneumoniae and human cytomegalovirus (HCMV) are intracellular pathogens able to infect hepatocytes, causing an increase in serum triglycerides and cholesterol levels due to the production of inflammatory cytokines. We investigated whether these pathogens could interfere with cholesterol metabolism by affecting activity of hepatic cholesterol 7α-hydroxylase (CYP7A1) promoter. CYP7A1 is the rate-limiting enzyme responsible for conversion of cholesterol to bile acids, which represents the main route of cholesterol catabolism. A straightforward dual-reporter bioluminescent assay was developed to simultaneously monitor CYP7A1 transcriptional regulation and cell viability in infected human hepatoblastoma HepG2 cells. C. pneumoniae and HCMV infection significantly decreased CYP7A1 promoter activity in a dose-dependent manner, with maximal inhibitions of 33 ± 10% and 32 ± 4%, respectively, at a multiplicity of infection of 1. To support in vitro experiments, serum cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides and glucose levels were also measured in Balb/c mice infected with C. pneumoniae. Serum cholesterol and triglycerides also increased in infected mice compared with controls. Although further investigation is required, this work presents the first experimental evidence that C. pneumoniae and HCMV inhibit CYP7A1 gene transcription in the cultured human hepatoblastoma cell line.
UR - http://www.scopus.com/inward/record.url?scp=84865987459&partnerID=8YFLogxK
U2 - 10.1016/j.ab.2012.08.003
DO - 10.1016/j.ab.2012.08.003
M3 - Article
C2 - 22889738
AN - SCOPUS:84865987459
SN - 0003-2697
VL - 430
SP - 92
EP - 96
JO - Analytical Biochemistry
JF - Analytical Biochemistry
IS - 1
ER -