Humic acid in drinking well water induces inflammation through reactive oxygen species generation and activation of nuclear factor-κB/activator protein-1 signaling pathways: a possible role in atherosclerosis.
Humic acid (HA) has been implicated as one of the etiological factors in the peripheral vasculopathy of blackfoot disease (BFD) in Taiwan. However, the underlying pathophysiological mechanisms of BFD are not well defined. In this study, we used an in vitro and in vivo model, in which HA (25-200μg/mL) activated macrophages to produce pro-inflammatory molecules by activating their transcriptional factors. HA exposure induced NO and PGE2 production followed by induction of iNOS and COX-2 through NF-κB/AP-1 transactivation in macrophages. In addition, the production of TNF-α and IL-1β was significantly increased by HA. Moreover, HA-induced iNOS and COX-2 expression were down-regulated by the NF-κB and AP-1 inhibitors pyrrolidine dithiocarbamate (PDTC) and Tanshinone, respectively. Furthermore, generations of ROS and nitrotyrosine, as well as activation of the AKT and MAPKs signaling cascades were observed after HA exposure. Specifically, HA-induced NF-κB activation was mediated by ROS and AKT, and that HA-induced AP-1 activation was mediated by JNK and ERK. Notably, HA-mediated AKT, JNK, and ERK activation was ROS-independent. The inflammatory potential of HA was correlated with increased expression of HO-1 and Nrf2. Furthermore, an in vivo study confirms that mice exposed to HA, the serum levels of TNF-α and IL-1β was significantly increased in a dose-dependent manner. This report marks the first confirmation that environmental exposure of HA induces inflammation in macrophages, which may be one of the main causes of early atherogenesis in blackfoot disease.
Department of Cosmeceutics, China Medical University, Taichung 40402, Taiwan; Department of Molecular and Cellular Oncology, University of Texas, MD Anderson Cancer Center, TX 77030, USA.
You-Cheng
YC
Hseu
Department of Cosmeceutics, China Medical University, Taichung 40402, Taiwan.
K J
KJ
Senthil Kumar
Institute of Nutrition, China Medical University, Taichung 40402, Taiwan.
Chih-Sheng
CS
Chen
Institute of Nutrition, China Medical University, Taichung 40402, Taiwan.
Hsin-Ju
HJ
Cho
Institute of Nutrition, China Medical University, Taichung 40402, Taiwan.
Shu-Wei
SW
Lin
Institute of Nutrition, China Medical University, Taichung 40402, Taiwan.
Pei-Chun
PC
Shen
Department of Medical Laboratory Science and Biotechnology, China Medical University, Taichung 40402, Taiwan.
Cheng-Wen
CW
Lin
Institute of Medicine, Chun Shan Medical University, Taichung 40201, Taiwan.
Fung-Jou
FJ
Lu
Institute of Nutrition, China Medical University, Taichung 40402, Taiwan; Department of Molecular and Cellular Oncology, University of Texas, MD Anderson Cancer Center, TX 77030, USA. Electronic address: hlyang@mail.cmu.edu.tw.
Hsin-Ling
HL
Yang
D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D050197:Atherosclerosis / Q000209:etiology / Q000473:pathology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism; D015232:Dinoprostone / Q000097:blood; D060766:Drinking Water / Q000737:chemistry*; D005260:Female; D005786:Gene Expression Regulation; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism; D006812:Humic Substances / Q000009:adverse effects* / Q000032:analysis; D007249:Inflammation / Q000139:chemically induced* / Q000473:pathology; D053583:Interleukin-1beta / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000097:blood; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction; D013624:Taiwan; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D000975:Antioxidants; D060766:Drinking Water; D006812:Humic Substances; D053583:Interleukin-1beta; D008565:Membrane Proteins; D051267:NF-E2-Related Factor 2; D016328:NF-kappa B; C495636:Nfe2l2 protein, mouse; D017382:Reactive Oxygen Species; D018808:Transcription Factor AP-1; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D051547:Heme Oxygenase-1; C496542:Hmox1 protein, mouse; C077623:Ptgs2 protein, mouse; D051546:Cyclooxygenase 2; D015232:Dinoprostone; D000111:Acetylcysteine
AP-1; Activating protein-1; BFD; Blackfoot disease; COX-2; ERK; HA; HO-1; Heme oxygenase-1; Humic acid; IL-1β; IκB; JNK; Keap-1; Kelch-like ECH-associated protein 1; MAPK; NF-E2-related factor-2; NF-κB; NO; Nrf2; PDTC; PGE(2); ROS; TNF-α; c-JUN N-terminal kinase; cyclooxygenase-2; extracellular signal-regulated kinase; humic acid; iNOS; inducible nitric oxide synthase; inhibitor kappa B; interleukin-1 beta; mitogen activated protein kinase; nitric oxide (NO); nuclear factor-kappa B; prostaglandin E(2); pyrrolidine dithiocarbamate; reactive oxygen species; tumor necrosis factor-alpha