Humic acid induces the endothelial nitric oxide synthase phosphorylation at Ser1177 and Thr495 Via Hsp90α and Hsp90ß upregulation in human umbilical vein endothelial cells.

Humic acid (HA) has been implicated as a contributory factor for blackfoot disease, which is an endemic peripheral vascular disease. We investigated the effect of HA on the regulation of endothelial nitric oxide (NO) synthase (eNOS) in human umbilical vein endothelial cells (HUVECs) to evaluate the involvement of eNOS and related factors in peripheral vascular impairment with HA exposure. Treatment of HUVECs with HA induced upregulation of eNOS. This result coincides with those of previous studies. Furthermore this is the first study to report that HA induces upregulation of heat shock protein (Hsp)90α, Hsp90ß, eNOS phosphorylation at Ser1177, and eNOS phosphorylation at Thr495, as compared to that in the control. In contrast, treatment with BAPTA, an intracellular Ca(2+) chelator, inhibited upregulation of these proteins induced by HA. This study demonstrates that HA treatment leads to increases in both Hsp90α and Hsp90ß proteins and indicates that Hsp90α leads to eNOS phosphorylation at Ser1177 and that Hsp90ß leads to eNOS phosphorylation at Thr495, respectively. Upregulation of eNOS, Hsp90α, and Hsp90ß in HUVECs is regulated by intracellular Ca(2+) accumulation induced by HA. These results suggest that upregulation of eNOS phosphorylation at Ser1177 and eNOS phosphorylation at Thr495 produce NO and superoxide anions, respectively, resulting in generation of peroxynitrite, which causes impairment of vascular endothelial cells.

Effects of peat fires on the characteristics of humic acid extracted from peat soil in Central Kalimantan, Indonesia.

When peat forest fires happen, it leads to burn soil and also humic acids as a dominant organic matter contained in peat soil as well as the forest. The structure and properties of humic acids vary depending on their origin and environment, therefore the transformation of humic acid is also diverse. The impacts of the peat fires on peat soil from Central Kalimantan, Indonesia were investigated through the characterization of humic acids, extracted from soil in burnt and unburnt sites. The characterization of humic acids was performed by elemental composition, functional groups, molecular weight by HPSEC, pyrolysate compounds by pyrolysis-GC/MS, fluorescence spectrum by 3DEEM spectrofluorometer, and thermogravimetry. The elemental composition of each humic substance indicated that the value of H/C and O/C of humic acids from burnt sites were lower than that from unburnt sites. The molecular weight of humic acids from burnt sites was also lower than that from unburnt sites. Pyrolysate compounds of humic acids from unburnt sites differed from those of humic acids from burnt soil. The heating experiment showed that burning process caused the significant change in the properties of humic acids such as increasing the aromaticity and decreasing the molecular weight.

Mechanism of the toxicity induced by natural humic acid on human vascular endothelial cells.

Humic acid (HA), a group of high-molecular weight organic compounds characterized by an ability to bind heavy metals, is normally found in natural water. Although the impairment of vascular endothelial cells in the presence of humic substances has been reported to be involved in some diseases, the mechanisms responsible for this involvement remain unclear. In this study, we examined the cytotoxicity of HA obtained from peatland in Central Kalimantan, Indonesia, to human vascular endothelial cells, as well as the mechanisms behind these effects. It was found that 50 mg/L HA showed cytotoxicity, which we considered to be mediated by apoptosis through the mitochondrial pathway because of an increase in the expression of caspases 6 and 9 in response to HA administration. In addition, this cytotoxicity was enhanced when cells in this experimental system were exposed to oxidative stress, while it was decreased by the addition of vitamin C. Thus, we conclude that the apoptosis induced by HA depends upon oxidative stress. Furthermore, an iron chelator, DFO, showed a tendency to decrease HA-induced cytotoxicity, suggesting that iron may potentially mediate HA-induced oxidative stress. In conclusion, long-term consumption of HA-rich water obtained from our study area may cause damage to endothelial cells and subsequent chronic health problems.