Induction of Erythroid Differentiation in Human Erythroleukemia Cells by Depletion of Malic Enzyme 2

Malic enzyme 2 (ME2) is a mitochondrial enzyme that catalyzes the conversion of malate to pyruvate and CO2 and uses NAD as a cofactor. Higher expression of this enzyme correlates with the degree of cell de-differentiation. We found that ME2 is expressed in K562 erythroleukemia cells, in which a number of agents have been found to induce differentiation either along the erythroid or the myeloid lineage. We found that knockdown of ME2 led to diminished proliferation of tumor cells and increased apoptosis in vitro. These findings were accompanied by differentiation of K562 cells along the erythroid lineage, as confirmed by staining for glycophorin A and hemoglobin production. ME2 knockdown also totally abolished growth of K562 cells in nude mice. Increased ROS levels, likely reflecting increased mitochondrial production, and a decreased NADPH/NADP+ ratio were noted but use of a free radical scavenger to decrease inhibition of ROS levels did not reverse the differentiation or apoptotic phenotype, suggesting that ROS production is not causally involved in the resultant phenotype. As might be expected, depletion of ME2 induced an increase in the NAD+/NADH ratio and ATP levels fell significantly. Inhibition of the malate-aspartate shuttle was insufficient to induce K562 differentiation. We also examined several intracellular signaling pathways and expression of transcription factors and intermediate filament proteins whose expression is known to be modulated during erythroid differentiation in K562 cells. We found that silencing of ME2 leads to phospho-ERK1/2 inhibition, phospho-AKT activation, increased GATA-1 expression and diminished vimentin expression. Metabolomic analysis, conducted to gain insight into intermediary metabolic pathways that ME2 knockdown might affect, showed that ME2 depletion resulted in high orotate levels, suggesting potential impairment of pyrimidine metabolism. Collectively our data point to ME2 as a potentially novel metabolic target for leukemia therapy

Behavior of chlorpyrifos and its major metabolite TCP (3,5,6-trichloro-2-pyridinol) in agricultural soils amended with drinking water treatment residuals

Purpose: Chlorpyrifos can be effectively adsorbed by drinking water treatment residuals (WTR), ubiquitous and non-hazardous by-products of potable water production. The major metabolite 3,5,6-trichloro-2-pyridinol (TCP) was found to be much more mobile and toxic than its parent chlorpyrifos. To assess the feasibility of WTR amendment for attenuation of chlorpyrifos and TCP pollution, the sorption/desorption and degradation behavior of chlorpyrifos and TCP in WTR-amended agricultural soils was examined in the present study. Materials and methods: Two representative agricultural soils were sampled from southern and northern China, respectively. The soils were amended with WTR at the rates of 0, 2, 5, and 10 % (w/w). Batch sorption/desorption test were applied to investigate the sorption/desorption characteristics of chlorpyrifos and TCP in WTR-amended soils. The influence of WTR amendment on chlorpyrifos degradation and TCP formation was evaluated using the incubation test, and its effect on the soil bacterial abundance was further studied through DNA extraction and PCR amplification. Results and discussion: Results showed that WTR amendment (0–10 %, w/w) significantly enhanced the retention capacity of chlorpyrifos and TCP in both soils examined (P &lt; 0.05). Fractionation analyses further demonstrated that the bioavailability of chlorpyrifos was considerably reduced by WTR amendment, resulting in a decreased chlorpyrifos degradation rate. The WTR amendment also significantly reduced the mobility of TCP formed in chlorpyrifos-contaminated soils (P &lt; 0.001). The chlorpyrifos toxicity to soil bacteria community was largely mitigated following WTR amendment, resulting in increased total bacterial abundance. Conclusions: Results obtained in the present study indicate a great deal of potential for the beneficial reuse of WTR as soil amendments for chlorpyrifos and TCP pollution control.</p