Photo-Reactivity of Natural Dissolved Organic Matter from Fresh to Marine Waters in the Florida Everglades, USA

Natural dissolved organic matter (DOM) is the major absorber of sunlight in most natural waters and a critical component of carbon cycling in aquatic systems. The combined effect of light absorbance properties and related photo-production of reactive species are essential in determining the reactivity of DOM. Optical properties and in particular excitation–emission matrix fluorescence spectroscopy combined with parallel factor analysis (EEM-PARAFAC) have been used increasingly to track sources and fate of DOM. Here we describe studies conducted in water from two estuarine systems in the Florida Everglades, with a salinity gradient of 2 to 37 and dissolved organic carbon concentrations from 19.3 to 5.74 mg C L−1, aimed at assessing how the quantity and quality of DOM is coupled to the formation rates and steady-state concentrations of reactive species including singlet oxygen, hydroxyl radical, and the triplet excited state of DOM. These species were related to optical properties and PARAFAC components of the DOM. The formation rate and steady-state concentration of the carbonate radical was calculated in all samples. The data suggests that formation rates, particularly for singlet oxygen and hydroxyl radicals, are strongly coupled to the abundance of terrestrial humic-like substances. A decrease in singlet oxygen, hydroxyl radical, and carbonate radical formation rates and steady-state concentration along the estuarine salinity gradient was observed as the relative concentration of terrestrial humic-like DOM decreased due to mixing with microbial humic-like and protein-like DOM components, while the formation rate of triplet excited-state DOM did not change. Fluorescent DOM was also found to be more tightly coupled to reactive species generation than chromophoric DOM.

Reactive Oxygen Species via Redox Signaling to PI3K/AKT Pathway Contribute to the Malignant Growth of 4-Hydroxy Estradiol-Transformed Mammary Epithelial Cells

The purpose of this study was to investigate the effects of 17-β-estradiol (E2)-induced reactive oxygen species (ROS) on the induction of mammary tumorigenesis. We found that ROS-induced by repeated exposures to 4-hydroxy-estradiol (4-OH-E2), a predominant catechol metabolite of E2, caused transformation of normal human mammary epithelial MCF-10A cells with malignant growth in nude mice. This was evident from inhibition of estrogen-induced breast tumor formation in the xenograft model by both overexpression of catalase as well as by co-treatment with Ebselen. To understand how 4-OH-E2 induces this malignant phenotype through ROS, we investigated the effects of 4-OH-E2 on redox-sensitive signal transduction pathways. During the malignant transformation process we observed that 4-OH-E2 treatment increased AKT phosphorylation through PI3K activation. The PI3K-mediated phosphorylation of AKT in 4-OH-E2-treated cells was inhibited by ROS modifiers as well as by silencing of AKT expression. RNA interference of AKT markedly inhibited 4-OH-E2-induced in vitro tumor formation. The expression of cell cycle genes, cdc2, PRC1 and PCNA and one of transcription factors that control the expression of these genes – nuclear respiratory factor-1 (NRF-1) was significantly up-regulated during the 4-OH-E2-mediated malignant transformation process. The increased expression of these genes was inhibited by ROS modifiers as well as by silencing of AKT expression. These results indicate that 4-OH-E2-induced cell transformation may be mediated, in part, through redox-sensitive AKT signal transduction pathways by up-regulating the expression of cell cycle genes cdc2, PRC1 and PCNA, and the transcription factor – NRF-1. In summary, our study has demonstrated that: (i) 4-OH-E2 is one of the main estrogen metabolites that induce mammary tumorigenesis and (ii) ROS-mediated signaling leading to the activation of PI3K/AKT pathway plays an important role in the generation of 4-OH-E2-induced malignant phenotype of breast epithelial cells. In conclusion, ROS are important signaling molecules in the development of estrogen-induced malignant breast lesions.