Microarray analysis reveals the inhibition of nuclear factor-kappa B signaling by aristolochic acid in normal human kidney (HK-2) cells.
OBJECTIVE
To study the molecular mechanism underlying the effect of aristolochic acid (AA), a major active component of plants from the Aristolochiaceae family using microarray analysis.


METHODS
Human kidney (HK-2) cells were treated with AA (0, 10, 30, and 90 micromol/L) for 24 h, and the cell viability was measured by a 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide assay. Complementary DNA microarrays were used to investigate the gene expression pattern of HK-2 cells exposed to AA in triplicate. A quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) assay was used to verify the microarray data for selected nuclear factor kappa B (NF-kappaB)-regulated genes. Furthermore, the subcellular localization of NF-kappaB p65 was visualized by immunofluorescence confocal microscopy in HK-2 cells. The NF-kappaB activity was examined by a luciferase reporter assay in HK-2/NF-kappaB transgenic cells.


RESULTS
AA exhibited a dose-dependent cytotoxic effect in HK-2 cells and induced alterations in the gene expression profiles related to the DNA damage response, DNA repair, macromolecule metabolic process, carbohydrate metabolic process, DNA metabolic process, apoptosis, cell cycle, and transcription. In addition, 9 biological pathways associated with immunomodulatory functions were down-regulated in AA-treated HK-2 cells. A network analysis revealed that NF-kappaB played a central role in the network topology. Among NF-kappaB-regulated genes, 8 differentially expressed genes were verified by qRT-PCR. The inhibition of NF-kappaB activity by AA was further confirmed by immunofluorescence confocal microscopy and by NF-kappaB luciferase reporter assay.


CONCLUSIONS
Our data revealed that AA could suppress NF-kappaB activity in normal human cells, perhaps partially accounting for the reported anti-inflammatory effects of some plants from the genus Aristolochia.
Chen
Ya-yin
YY
China Medical University, Taichung, Taiwan, China.
Chiang
Su-yin
SY
Wu
Hsiu-ching
HC
Kao
Shung-te
ST
Hsiang
Chien-yun
CY
Ho
Tin-yun
TY
Lin
Jaung-geng
JG
D034341:Aristolochic Acids / Q000494:pharmacology*; D001483:Base Sequence; D002461:Cell Line, Transformed; D017931:DNA Primers; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction
D034341:Aristolochic Acids; D017931:DNA Primers; D016328:NF-kappa B; C000228:aristolochic acid I