Shuangyue Li

Chlorpyrifos Induces MLL Translocations Through Caspase 3-Dependent Genomic Instability and Topoisomerase II Inhibition in Human Fetal Liver Hematopoietic Stem Cells

Household pesticide exposure during pregnancy has been associated with a more than 2-fold increased risk in infant leukemia, and chlorpyrifos (CPF) is among the most frequently applied insecticides. During early fetal development, liver is a hematopoietic organ with majority of cells being CD34(+) hematopoietic stem cells (CD34(+)HSC). The in utero injury to CD34(+)HSC has been known to underlie the pathogenesis of several blood disorders, often involving rearrangements of the mixed-lineage leukemia (MLL) gene on 11q23. In this study, we evaluated the leukemogenic potential of CPF in human fetal liver-derived CD34(+)HSC. Specifically, exposure to 10 μM CPF led to decrease in viability, inhibition in proliferation and induction of DNA double-strand breaks (DSBs) and occurrence of MLL(+) rearrangements. In particular, we observed CPF-mediated cell cycle disturbance as shown by G0/G1 arrest, in contrast to etoposide (VP-16), an anticancer drug used as a positive control and known to induce G2/M arrest. Further study on mechanisms underlying DNA DSBs and MLL(+) rearrangements revealed that CPF might act as topoisomerase II poison, a mechanism of action similar to VP-16. On the other hand, CPF was also shown to induce early apoptosis through active caspase-3 activation, a pathway known to underlie DNA DSBs and MLL(+) translocations. Our data indicate that in utero injury of CD34(+)HSC by CPF may contribute to the increased risk of infant leukemia. Future work will elucidate the mechanism and the type of CPF-induced MLL(+) translocations in HSC.

Bone marrow mesenchymal stem cells attenuate 2,5-hexanedione-induced neuronal apoptosis through a NGF/AKT-dependent pathway

Growing evidence suggests that the increased neuronal apoptosis is involved in n-hexane-induced neuropathy. We have recently reported that bone marrow-mesenchymal stem cells-derived conditioned medium (BMSC-CM) attenuated 2,5-hexanedione (HD, the active metabolite of n-hexane)-induced apoptosis in PC12 cells. Here, we explored the anti-apoptotic efficacy of BMSC in vivo. HD-treated rats received BMSC by tail vein injection 5 weeks after HD intoxication. We found that in grafted rats, BMSC significantly attenuated HD-induced neuronal apoptosis in the spinal cord, which was associated with elevation of nerve growth factor (NGF). Neutralization of NGF in BMSC-CM blocked the protection against HD-induced apoptosis in VSC4.1 cells, suggesting that NGF is essential for BMSC-afforded anti-apoptosis. Mechanistically, we found that the decreased activation of Akt induced by HD was significantly recovered in the spinal cord by BMSC and in VSC4.1 cells by BMSC-CM in a TrkA-dependent manner, leading to dissociation of Bad/Bcl-xL complex in mitochondria and release of anti-apoptotic Bcl-xL. The importance of Akt was further corroborated by showing the reduced anti-apoptotic potency of BMSC in HD-intoxicated VSC4.1 cells in the presence of Akt inhibitor, MK-2206. Thus, our findings show that BMSC attenuated HD-induced neuronal apoptosis in vivo through a NGF/Akt-dependent manner, providing a novel solution against n-hexane-induced neurotoxicity.

Evaluation of PFOS-mediated neurotoxicity in rat primary neurons and astrocytes cultured separately or in co-culture.

Perfluorooctane sulfonate (PFOS) is a potential neurotoxicant reported by epidemiological investigations and experimental studies, while the underlying mechanisms are still unclear. Astrocytes not only support for the construction of neurons, but also conduct neuronal functions through glutamate-glutamine cycle in astrocyte-neuron crosstalk. In the present study, the effect of PFOS exposure on rat primary hippocampal neurons or cortex astrocytes was evaluated. Then the role of the astrocytes in PFOS-induced toxic effect on neurons was explored with astrocyte-neuron co-culture system. Exposure of rat primary hippocampal neurons to PFOS has led to oxidation-antioxidation imbalance, increased apoptosis and abnormal autophagy. The adverse effect of PFOS on rat primary cortex astrocytes manifested in the form of altered extracellular glutamate and glutamine concentrations, decreased glutamine synthase activity, as well as decreased gene expression of glutamine synthase, glutamate transporters and glutamine transporters in the glutamate-glutamine cycle. Especially, the alleviation of PFOS-inhibited neurite outgrowth in neurons could be observed in astrocyte-neuron co-culture system, though the ability of astrocytes in fostering neurite outgrowth was affected by PFOS. These results indicated that both astrocytes and neurons might be the targets of PFOS-induced neurotoxicity, and astrocytes could protect against PFOS-inhibited neurite outgrowth in primary cultured neurons. Our research might render some information in explaining the mechanisms of PFOS-induced neurotoxicity.

2,5-hexanedione downregulates nerve growth factor and induces neuron apoptosis in the spinal cord of rats via inhibition of the PI3K/Akt signaling pathway

2,5-hexanedione (2,5-HD) is the main active metabolite of n-hexane and induces apoptosis in nerve tissue, however, the mechanism of which remains unclear. In the present study, neuropathic animal models were successfully constructed in rats by injecting 100, 200 and 400 mg/kg 2,5-HD intraperitoneally for 5 weeks. Rats exposed to 2,5-HD exhibited progressive gait abnormalities and slower motor neural response in a dose-dependent manner. TUNEL analysis and immunofluorescence dual labeling revealed that the spinal cord of the 2,5-HD treated rats underwent significantly more apoptosis in the cells of spinal cord than that of the control group. The neuron apoptosis index in spinal cord was 4.1%, 6.7%, 9.8% respectively in rats exposed to 100, 200 and 400 mg/kg 2,5-HD, compared with 1.1% in the control group (p < 0.05). Biochemical analysis showed that 2,5-HD exposure downregulated NGF expression in the spinal cord of the intoxicated rats; inhibited the phosphorylation of Akt and Bad, two key players in PI3K/Akt pathway downstream of NGF; increased the dimerization of Bad with Bcl-xL in the mitochondrial fraction, followed by the release of cytochrome c and activation of caspase-3 in the spinal cord of rats. In vitro study showed that the NGF expression decreased significantly in VSC4.1 cells dosed with 5.0, 10.0 mM 2,5-HD in comparison with the control group. It was also found that NGF supplement repressed the induced apoptosis, and increased p-Akt and p-Bad level in 2,5-HD treated VSC4.1 cells, which could be antagonized by PI3K kinase (the upstream member of Akt) inhibitor LY294002. Taken together, our experimental results indicate that 2,5-HD may induce apoptosis in the spinal cord of rats via downregulating NGF expression and subsequently repressing PI3K/Akt signaling pathway.

Taurine Ameliorates Arsenic-Induced Apoptosis in the Hippocampus of Mice Through Intrinsic Pathway

Our group previously reported that arsenic (As) exposure induced apoptosis in hippocampus neurons. The aim of the present study was to clarify the protective capacity of taurine (Tau) on As-induced neuronal apoptosis and the related mechanism in mouse hippocampus. Mice were divided into: control group, Tau control group, As exposure group and Tau protective group, randomly. The apoptotic rate of mouse hippocampus was determined by TUNEL staining. The levels of Bcl-2 and Bax gene and protein were analyzed by real time RT-PCR and WB, respectively. Furthermore, cytochrome c (Cyt C) release, and the activity of caspase-8 and caspase-3 were also determined. The results showed that Tau treatment induced the decrease of TUNEL-positive cells, prohibited the disturbance of Bcl-2 and Bax expression, and inhibited Cyt C release and caspase-8 and caspase-3 activation significantly. The results indicated that Tau supplement markedly ameliorates As-induced apoptosis by mitochondria-related pathway in mouse hippocampus.

Subchronic Exposure to Arsenic Represses the TH/TRβ1-CaMK IV Signaling Pathway in Mouse Cerebellum

We previously reported that arsenic (As) impaired learning and memory by down-regulating calmodulin-dependent protein kinase IV (CaMK IV) in mouse cerebellum. It has been documented that the thyroid hormone receptor (TR)/retinoid X receptor (RXR) heterodimer and thyroid hormone (TH) may be involved in the regulation of CaMK IV. To investigate whether As affects the TR/RXR heterodimer and TH, we determined As concentration in serum and cerebellum, 3,5,3’-triiodothyronine (T3) and thyroxin (T4) levels in serum, and expression of CaMK IV, TR and RXR in cerebellum of mice exposed to As. Cognition function was examined by the step-down passive avoidance task and Morris water maze (MWM) tests. Morphology of the cerebellum was observed by Hematoxylin-Eosin staining under light microscope. Our results showed that the concentrations of As in the serum and cerebellum of mice both increased with increasing As-exposure level. A significant positive correlation was found between the two processes. Adeficit in learning and memory was found in the exposed mice. Abnormal morphologic changes of Purkinje cells were observed in cerebellum of the exposed mice. Moreover, the cerebellar expressions of CaMK IV protein and the TRβ gene, and TRβ1 protein were significantly lower in As-exposed mice than those in controls. Subchronic exposure to As appears to increase its level in serum and cerebella of mice, impairing learning and memory and down-regulating expression of TRβ1 as well as down-stream CaMK IV. It is also suggested that the increased As may be responsible for down-regulation of TRβ1 and CaMK IV in cerebellum and that the down-regulated TRβ1 may be involved in As-induced impairment of learning and memory via inhibiting CaMK IV and its down-stream pathway.

Protection of Taurine Against Arsenic-Induced DNA Damage of Mice Kidneys

The purpose of this study was to explore the protective capacity of taurine on arsenic (As)-induced neurotoxicity. Thirty mice were used and ten rats in each group. We treated the As exposure group with 4 ppm As2O3 for 60 days by drinking water and the protective group with 4 ppm As2O3 and 150 mg/kg taurine. Drinking water was only given in the control group. Pathologic changes and DNA damage in the mice kidney were examined by HE staining, immunohistochemistry and comet assay. Abnormal morphological changes were found in the kidney of As exposed mouse. Moreover, 8-hydroxy-2-deoxyguanosine (8-OHdG) expression and comet number, tail moment, and tail length of comet were markedly elevated in the As intoxication mice. However, histopathological changes and low expression of 8-OHdG were shown in the protective group. Our results indicate that supplementation of taurine protects against the histopathologic changes and DNA damage of mouse kidneys in As exposure group.

Protection of Taurine Against PFOS-Induced Neurotoxicity in PC12 Cells

As a new member of persistent organic pollutants, the potent neurotoxicity of perfluorooctane sulfonates (PFOS) found in epidemiological studies and laboratory research has drawn increasing attention around the world. Previous studies showed that apoptosis driven by oxidative stress and autophagy were both observed in PFOS-induced toxicity. Taurine has been demonstrated to exert potent protections against oxidative stress as an efficient antioxidant. Whether taurine could protect against the PFOS neurotoxicity is not known. In the present study, PC12 cells were treated with several concentrations of PFOS (31.25, 250 μM) for 24 h. 3-(4,5-Dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay was applied to assess the cell viability. DCFH-DA detector was used to explore the production of ROS. Caspase 3 activity was used to reflect the possible apoptosis pathway. The lyso-tracker red dying was invited to evaluate the autophagy. Our data showed that taurine could significantly reverse the decreased viability and the increased ROS production in PC12 cells treated with PFOS. Moreover, the increased autophagy and apoptosis elicited by PFOS in PC12 cells could also be attenuated by taurine. Collectively, our results indicate that taurine may be an effective antioxidant in fighting against PFOS cytotoxicity and therefore could potentially serve as a preventative and therapeutic agent for environmental pollution-related toxicities.

Taurine Attenuates As 2 O 3 -Induced Autophagy in Cerebrum of Mouse Through Nrf2 Pathway

We previously reported that the impairment of cerebrum may relate with neurotoxicity induced by arsenic (As) exposure. In the present study, we investigated whether autophagy of the cerebrum neurons were responsible for As-induced neurotoxicity and the protective role of taurine (Tau). Forty mice were randomly divided into control group, Tau control group, As exposure group and Tau protection group. The results showed that LC3 II expression was elevated and P62 expression was lower after As exposure, whereas the effects were obviously attenuated by Tau treatment. More important, As induced increase of MDA level and decrease of Nrf2 expression were significantly inversed in protective group. In sum, autophagy inhibition might play a strong role in the neuroprotection of Tau in As-induced toxicity via Nrf2 pathway.

Taurine Normalizes the Levels of Se, Cu, Fe in Mouse Liver and Kidney Exposed to Arsenic Subchronically

To evaluate the benefits of taurine on the homeostasis of trace elements induced by toxic metals, we investigated the concentration of Selenium (Se), Copper (Cu), Iron (Fe) and Manganese (Mn) in mouse liver and kidney after arsenic exposure for 2 months. The experimental animals were divided into control group, arsenic exposure group (1, 2, 4 ppm) and taurine protective group randomly. Concentrations of serum, liver and kidney trace elements such as Se, Cu, Fe, Mn were measured by Inductively Coupled Plasma-Mass Spectrometry. Our results showed that the concentration of Cu was higher, however, the concentration of Se and Fe was lower in mice liver and kidney exposed to arsenic. The levels of Se, Cu, Fe were alleviated by co-administered with taurine. Furthermore, there was no difference in the concentration of Mn between the three groups. Our finding suggests that taurine may relieve the disturbed levels of Se, Cu and Fe in liver and kidney induced by arsenic.