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.
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.
Department of Chemistry "G. Ciamician", University of Bologna, Bologna, Italy. elisa.michelini8@unibo.it
Elisa
E
Michelini
Manuela
M
Donati
Rita
R
Aldini
Luca
L
Cevenini
Laura
L
Mezzanotte
Paola
P
Nardini
Claudio
C
Foschi
Ido Ben
IB
Zvi
Monica
M
Cevenini
Marco
M
Montagnani
Antonella
A
Marangoni
Aldo
A
Roda
Roberto
R
Cevenini
D000818:Animals; D001786:Blood Glucose / Q000378:metabolism; D023521:Chlamydophila Infections / Q000097:blood / Q000201:enzymology / Q000235:genetics; D016993:Chlamydophila pneumoniae / Q000472:pathogenicity / Q000502:physiology*; D002790:Cholesterol 7-alpha-Hydroxylase / Q000235:genetics*; D008076:Cholesterol, HDL / Q000097:blood; D003116:Color; D003587:Cytomegalovirus / Q000472:pathogenicity / Q000502:physiology*; D056945:Hep G2 Cells; D006801:Humans; D008163:Luminescent Measurements / Q000379:methods*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D014158:Transcription, Genetic; D014280:Triglycerides / Q000097:blood
D001786:Blood Glucose; D008076:Cholesterol, HDL; D014280:Triglycerides; C553219:CYP7A1 protein, human; D002790:Cholesterol 7-alpha-Hydroxylase