Humic acid induces apoptosis in human endothelial cells.
Humic acid (HA) has been implicated as an etiologic factor in the vasculopathy of Blackfoot disease. In this study, the ability of HA to induce apoptosis was studied in cultured human umbilical vein endothelial cells. Treatment of endothelial cells with a variety of concentrations of HA (50-200 microg/ml) resulted in dose- and time-dependent sequences of events marked by apoptosis as shown by loss of cell viability, chromatin condensation, and internucleosomal DNA fragmentation. Antioxidants (superoxide dismutase, vitamin C, and vitamin E) and Ca(2+) chelator (BAPTA) effectively suppressed HA-induced DNA fragmentation (apoptosis). Further studies have shown that HA induced dramatic Ca(2+)-dependent caspase activation (2, 3, 6, 8, and 9). In contrast, negligible caspase-1 activation was observed. The increase in HA-induced apoptosis correlated with a reduction in Bcl-2, a potent cell death inhibitor, and an increase in Bax protein levels, which heterodimerizes with and thereby inhibits Bcl-2. Both of the antioxidants vitamin C and vitamin E prevented the dysregulation of Bcl-2 and Bax in HA-treated endothelial cells. Furthermore, the increase in p53 protein levels correlated with an increase in HA-induced apoptosis. We concluded that both Ca(2+) and oxidative stress were mediators of apoptosis caused by HA and the induction of apoptotic cell death on endothelial cells may be important to the etiology of HA-induced vascular disorder of Blackfoot disease.
Hseu
You-Cheng
YC
Department of Medical Technology, Fooyin Institute of Technology, Kaohsiung, Taiwan.
Huang
Hurng-Wern
HW
Wang
Shi-Ying
SY
Chen
Hwei-Yan
HY
Lu
Fung-Jou
FJ
Gau
Rung-Jiun
RJ
Yang
Hsin-Ling
HL
D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism; D020170:Caspase 1 / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D019254:Genes, bcl-2 / Q000187:drug effects; D006801:Humans; D006812:Humic Substances / Q000633:toxicity*; D020287:In Situ Nick-End Labeling; D016491:Peripheral Vascular Diseases / Q000139:chemically induced* / Q000378:metabolism / Q000473:pathology; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2; D013482:Superoxide Dismutase / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D014801:Vitamin A / Q000494:pharmacology; D014810:Vitamin E / Q000494:pharmacology; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D006812:Humic Substances; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; D014801:Vitamin A; D014810:Vitamin E; D004533:Egtazic Acid; D013482:Superoxide Dismutase; D020170:Caspase 1; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium