{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,21]],"date-time":"2026-08-21T13:15:05Z","timestamp":1787318105526,"version":"build-2736575974"},"reference-count":37,"publisher":"MDPI AG","issue":"20","license":[{"start":{"date-parts":[[2025,10,15]],"date-time":"2025-10-15T00:00:00Z","timestamp":1760486400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100012046","name":"Vietnam Academy of Science and Technology","doi-asserted-by":"crossref","award":["T\u00d0DIOX.04\/22-24"],"award-info":[{"award-number":["T\u00d0DIOX.04\/22-24"]}],"id":[{"id":"10.13039\/100012046","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>This study reports the construction and validation of a CAFLUX (Chemically Activated Fluorescent Expression) HepG2 reporter cell line engineered to express a histone H2B\u2013green fluorescent protein (H2B\u2013GFP) fusion protein under the control of a dioxin-responsive cytochrome P450 1A1 (CYP1A1) promoter. A lentiviral construct containing a synthetic promoter with multiple dioxin-responsive elements (DREs) upstream of the H2B\u2013EGFP coding sequence was cloned into the pFUGW vector, packaged in human embryonic kidney (HEK) 293FT cells, and used to transduce HepG2 hepatocellular carcinoma cells. Stable clones obtained by limiting dilution were screened for GFP expression in response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The resulting CAFLUX HepG2 cells exhibited dose-dependent nuclear GFP fluorescence when exposed to aryl hydrocarbon receptor (AhR) agonists, with limits of detection of approximately 0.01 pM for TCDD and 0.1 pM for benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon (PAH). This reporter activity correlated with endogenous CYP1A1 mRNA expression as determined by quantitative polymerase chain reaction (qPCR), confirming that GFP signals reflected native transcriptional responses. In functional assays, curcumin suppressed GFP expression in a concentration-dependent manner and induced apoptotic morphology at higher doses, while extracellular vesicles (EVs) derived from adipose-derived stem cells (ADSCs) significantly reduced both GFP fluorescence and CYP1A1 mRNA levels, suggesting an inhibitory effect on AhR-driven transcription. The CAFLUX HepG2 reporter system therefore provides a sensitive and reproducible platform for real-time, nuclear-localized monitoring of AhR-mediated gene expression. Its responsiveness to both agonists and antagonists underscores its potential utility in toxicological evaluation, drug discovery, and the investigation of EV-mediated signaling in liver cancer models.<\/jats:p>","DOI":"10.3390\/ijms262010029","type":"journal-article","created":{"date-parts":[[2025,10,15]],"date-time":"2025-10-15T11:57:42Z","timestamp":1760529462000},"page":"10029","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Development and Application of a CAFLUX HepG2 Reporter Cell Line for Real-Time Monitoring of AhR-Mediated CYP1A1 Gene Expression in Response to Environmental Toxicants and Bioactive Modulators"],"prefix":"10.3390","volume":"26","author":[{"given":"Huyen Thi","family":"La","sequence":"first","affiliation":[{"name":"Institute of Biology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"},{"name":"Faculty of Biology, Graduate University of Science and Technology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hanh Hong","family":"Hoang","sequence":"additional","affiliation":[{"name":"Institute of Biology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7078-7345","authenticated-orcid":false,"given":"Phuc Minh Thi","family":"Le","sequence":"additional","affiliation":[{"name":"Institute of Biology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Linh Thuy","family":"Nguyen","sequence":"additional","affiliation":[{"name":"Institute of Biology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Da Thi","family":"Nguyen","sequence":"additional","affiliation":[{"name":"Institute of Biology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Van Hanh","family":"Nguyen","sequence":"additional","affiliation":[{"name":"Faculty of Agricultural Technology, VNU University of Engineering and Technology, 144 Xuan Thuy, Cau Giay, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tam Minh Thi","family":"Ha","sequence":"additional","affiliation":[{"name":"Faculty of Early Childhood Education, HaNoi Pedagogical University 2, 32 Nguyen Van Linh, Xuan Hoa 290000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Long Hoang","family":"Nguyen","sequence":"additional","affiliation":[{"name":"Institute for Advanced Study in Technology, Ton Duc Thang University, Ho Chi Minh City 70000, Vietnam"},{"name":"Faculty of Pharmacy, Ton Duc Thang University, Ho Chi Minh City 70000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dat Tien","family":"Nguyen","sequence":"additional","affiliation":[{"name":"Center for High Technology Research and Development, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,10,15]]},"reference":[{"key":"ref_1","unstructured":"World Health Organization (2025, April 01). 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