Dichlorodiphenyltrichloroethane specifically depletes dopaminergic neurons in primary cell culture

Toxicity of dichlorodiphenyltrichloroethane (DDT) to dopaminergic neurons in primary cell culture was investigated in the present study. Developing neurons from the substantia nigra of neonatal rats were cultured. After treatments with different concentrations of DDT (5-12.5 μM), specific cell death of tyrosine-hydroxylase-immunoreactive dopaminergic neurons was observed in the culture by flow cytometric analysis. More than 60% of dopaminergic neurons were depleted after treatments with 10 and 12.5 μM of DDT. In addition, significant reductions of intensity levels of tyrosine hydroxylase immunofluorescence were observed in dopaminergic neurons after DDT treatments even at low concentrations of DDT. The present findings indicate that dopaminergic neurons are more susceptible to DDT toxicity than other types of neurons in the primary cell culture. Moreover, it is shown that the synthesis of dopamine in dopaminergic neurons is also depressed. Previous studies have demonstrated that perinatal exposure of DDT causes neurons to be more susceptible to neurotoxic damages in later adult life. The present findings thus provide evidence that dopaminergic neurons that are undergoing growth and development are targets of DDT neurotoxic effects. Exposure to DDT from contaminated environments is therefore a potential risk of onset of Parkinson's disease. Copyright © 2003 S. Karger AG, Basel.published_or_final_versio

Molecular and cellular aspects of plasticity after neural injury

This review focuses on our effort to address plasticity of the nervous system after neural injury. We have used different animal models to examine cellular mechanisms of plasticity underlying the pathological and repair processes. After severance of sensory input from one inner ear, topographic representations of spacecentered coordinates in the brain undergo plastic changes. During vestibular compensation, tissue plasticity constitutes an important component for functional recovery of neuronal network. In Parkinsonian animals, modulation of signaling via glutamatergic synapses, neurotrophins and neurokinins contributes to the protection of basal ganglion neurons from degeneration, thereby delaying deterioration of motor functions. With the use of animal models of neural injury, we further overcome the molecular restriction at the glial scar to enhance neural regrowth and remyelination, pointing to the possibility of developing new therapeutic strategies to stimulate neural plasticity and repair in the adult nervous system.published_or_final_versio

Targeting GRB7/ERK/FOXM1 Signaling Pathway Impairs Aggressiveness of Ovarian Cancer Cells

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DLX1 acts as a crucial target of FOXM1 to promote ovarian cancer aggressiveness by enhancing TGF-β/SMAD4 signaling

Recent evidence from a comprehensive genome analysis and functional studies have revealed that FOXM1 is a crucial metastatic regulator that drives cancer progression. However, the regulatory mechanism by which FOXM1 exerts its metastatic functions in cancer cells remains obscure. Here, we report that DLX1 acts as a FOXM1 downstream target, exerting pro-metastatic function in ovarian cancers. Both FOXM1 isoforms (FOXM1B or FOXM1C) could transcriptionally upregulate DLX1 through two conserved binding sites, located at +61 to +69bp downstream (TFBS1) and -675 to -667bp upstream (TFBS2) of the DLX1 promoter, respectively. This regulation was further accentuated by the significant correlation between the nuclear expression of FOXM1 and DLX1 in high-grade serous ovarian cancers. Functionally, the ectopic expression of DLX1 promoted ovarian cancer cell growth, cell migration/invasion and intraperitoneal dissemination of ovarian cancer in mice, whereas small interfering RNA-mediated DLX1 knockdown in FOXM1-overexpressing ovarian cancer cells abrogated these oncogenic capacities. In contrast, depletion of FOXM1 by shRNAi only partially attenuated tumor growth and exerted almost no effect on cell migration/invasion and the intraperitoneal dissemination of DLX1-overexpressing ovarian cancer cells. Furthermore, the mechanistic studies showed that DLX1 positively modulates TGF- signaling by upregulating PAI-1 and JUNB through direct interaction with SMAD4 in the nucleus upon TGF-1 induction. Taken together, these data strongly suggest that DLX1 plays a pivotal role in FOXM1 signaling to promote cancer aggressiveness through intensifying TGF-/SMAD4 signaling in high-grade serous ovarian cancer cells.published_or_final_versio

GRO-α and IL-8 enhance ovarian cancer metastatic potential via the CXCR2-mediated TAK1/NFκB signaling cascade

Intraperitoneal metastasis is a common occurrence and is usually involved in the poor prognosis of ovarian cancer. Its specific metastatic pattern implies that certain indispensable microenvironmental factors secreted in the peritoneal cavity can direct metastatic ovarian cancer cells to permissive niches for secondary lesion formation. However, the underlying molecular mechanisms are ill defined. Herein, we report that GRO-α and IL-8 are predominately upregulated in culture media derived from either normal or cancerous omenta and are associated with increased ovarian cancer aggressiveness. Methods: OCM was established from culture medium of fresh human omental tissues. Primary and metastatic ovarian cancer cell lines were generated from human tumor tissues and verified by specific antibodies. The functional roles of GRO-α, IL-8, and their specific receptor CXCR2 were examined by neutralizing antibodies, shRNA gene knockdown, CRISPR/Cas9 gene knockout and pharmaceutical CXCR2 inhibitor SB225002. The oncogenic properties of ovarian cancer cells were examined by in vitro and in vivo mouse models. Results: Both GRO-α and IL-8 can activate TAK1/NFκB signaling via the CXCR2 receptor. Intriguingly, TAK1/NFκB signaling activity was higher in metastatic ovarian cancer cells; this higher activity makes them more susceptible to OCM-induced tumor aggressiveness. Treatment of ovarian cancer cells with GRO-α and IL-8 neutralizing antibodies or ablation of CXCR2 by shRNA gene knockdown, CRISPR/Cas9 gene knockout, or CXCR2 inhibitor SB225002 treatment significantly attenuated TAK1/NFκB signaling and decreased in vitro and in vivo oncogenic and metastatic potential, suggesting CXCR2 plays a key role in the GRO-α and IL-8-governed metastatic spreading of ovarian cancer cells in the intraperitoneal cavity. Conclusion: This study highlights the significance of GRO-α and IL-8 as the key chemokines in the peritoneal tumor microenvironment and suggests the utility of targeting their receptor CXCR2 as a potential target-based therapy for peritoneal metastases of ovarian cancer.published_or_final_versio

Methylation-associated silencing of miR-193a-3p promotes ovarian cancer aggressiveness by targeting GRB7 and MAPK/ERK pathways

Human growth factor receptor-bound protein-7 (GRB7) is a pivotal mediator involved in receptor tyrosine kinase signaling and governing diverse cellular processes. Aberrant upregulation of GRB7 is frequently associated with the progression of human cancers. However, the molecular mechanisms leading to the upregulation of GRB7 remain largely unknown. Here, we propose that the epigenetic modification of GRB7 at the post-transcriptional level may be a crucial factor leading to GRB7 upregulation in ovarian cancers. Methods: The upstream miRNA regulators were predicted by in silico analysis. Expression of GRB7 was examined by qPCR, immunoblotting and immunohistochemical analyses, while miR-193a-3p levels were evaluated by qPCR and in situ hybridization in ovarian cancer cell lines and clinical tissue arrays. MS-PCR and pyrosequencing analyses were used to assess the methylation status of miR-193a-3p. Stable overexpression or gene knockdown and Tet-on inducible approaches, in combination with in vitro and in vivo tumorigenic assays, were employed to investigate the functions of GRB7 and miR-193a-3p in ovarian cancer cells. Results: Both miR-193a-3p and its isoform, miR-193b-3p, directly targeted the 3' UTR of GRB7. However, only miR-193a-3p showed a significantly inverse correlation with GRB7-upregulated ovarian cancers. Epigenetic studies revealed that methylation-mediated silencing of miR-193a-3p led to a stepwise decrease in miR-193a-3p expression from low to high-grade ovarian cancers. Intriguingly, miR-193a-3p not only modulated GRB7 but also ERBB4, SOS2 and KRAS in the MAPK/ERK signaling pathway to enhance the oncogenic properties of ovarian cancer cells in vitro and in vivo. Conclusion: These findings suggest that epigenetic silencing of miR-193a-3p by DNA hypermethylation is a dynamic process in ovarian cancer progression, and miR-193a-3p may be explored as a promising miRNA replacement therapy in this disease.published_or_final_versio

5-HT excites globus pallidus neurons by multiple receptor mechanisms

Anatomical and neurochemical studies indicated that the globus pallidus receives serotonergic innervation from raphe nuclei but the membrane effects of 5-HT on globus pallidus neurons are not entirely clear. We address this question by applying whole-cell patch-clamp recordings on globus pallidus neurons in immature rat brain slices. Under current-clamp recording, 5-HT depolarized globus pallidus neurons and increased their firing rate, an action blocked by both 5-HT4 and 5-HT7 receptor antagonists and attributable to an increase in cation conductance(s). Further experiments indicated that 5-HT enhanced the hyperpolarization-activated inward conductance which is blocked by 5-HT7 receptor antagonist. To determine if 5-HT exerts any presynaptic effects on GABAergic and glutamatergic inputs, the actions of 5-HT on synaptic currents were studied. At 10 μM, 5-HT increased the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) but had no effect on both the frequency and amplitude of miniature inhibitory postsynaptic currents (mIPSCs). However, 5-HT at a higher concentration (50 μM) decreased the frequency but not the amplitude of the mIPSCs, indicating an inhibition of GABA release from the presynaptic terminals. This effect was sensitive to 5-HT1B receptor antagonist. In addition to the presynaptic effects on GABAergic neurotransmission, 5-HT at 50 μM had no consistent effects on glutamatergic neurotransmission, significantly increased the frequency of miniature excitatory postsynaptic currents (mEPSCs) in 4 of 11 neurons and decreased the frequency of mEPSCs in 3 of 11 neurons. In conclusion, we found that 5-HT could modulate the excitability of globus pallidus neurons by both pre- and post-synaptic mechanisms. In view of the extensive innervation by globus pallidus neurons on other basal ganglia nuclei, this action of 5-HT originated from the raphe may have a profound effect on the operation of the entire basal ganglia network. © 2008 IBRO.link_to_subscribed_fulltex

Neurokinin peptides and neurokinin receptors as potential therapeutic intervention targets of basal ganglia in the prevention and treatment of Parkinson's disease

Parkinson's disease (PD) is a serious motor disorder and it is the second most common brain degenerative disease in human. PD is known to be caused by degeneration of dopamine neurons in the substantia nigra but the cause of cell death is largely unknown. Mammalian neurokinins [NKs] are a group of neuropeptides that include substance P (SP; neurokinin-1, NK-1), substance K (SK; NK-2, neurokinin A), and neuromedin K (NK; NK-3; neurokinin B). Their biological effects as neurotransmitters, neuromodulators, or neurotrophic-like factors are mediated by three distinct neurokinin receptors, namely SP receptor (SPR: NK-1 receptor, NK-1R), SKR (NK-2R), and NKR (NK-3R). Several lines of evidence have indicated that neurokinins are implicated in the pathogenesis of PD. First, decreases of SP level and SP-immunoreactivity have been found in nigral and striatal tissues of animals with PD and postmortem PD patients. Second, NKs exert neuroprotective effects on neurons. In addition, NK receptors, namely NK-1 and NK-3 receptors, are abundantly localized in dopaminergic and cholinergic neurons of the basal ganglia, indicating that these neurons are under the physiological regulation of NKs. Moreover, modulation in motor activity occurred in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice, PD animal model, after systemic administration of NK receptor agonists. NKs and NK receptors, therefore, might be important molecules that are associated with functions and survival of neurons in the basal ganglia, in particular the dopamine neurons. Further studies should be devoted to elucidate the functional roles of NK systems in (a) the neuropathogenesis and neuroprotection during the course of PD, (b) the efficacy of NK receptor drugs towards PD, and (c) potential therapeutic intervention that targets at the prevention or treatment of PD. © 2004 Bentham Science Publishers Ltd.link_to_subscribed_fulltex

Modulation of the gene expression of N-methyl-D-aspartate receptor NR2B subunit in the rat neostriatum by a single dose of specific antisense oligodeoxynucleotide

N-methyl-D-aspartate receptors (NRs) are a group of ionotropic glutamate receptors in the brain and they are composed of heteromeric subunits (NR1, NR2A-D and NR3). In the neostriatum, a brain region that is associated with movement in animals, NMDA channels are known to involve in the motor control. Our previous report (Lai et al., 2000, Neuroscience 98, 493-500) has shown that a single dose of antisense oligodeoxynucleotides that are specific to NR1 subunit results in blockage of the gene expression of NR1 as well as NR2A subunits in the neostriatum. In the present study, antisense oligodeoxynucleotides that are specific to NR2B (ANR2B) were then employed as molecular tools to further investigate the molecular interactions of NMDA receptor subunits in the neostriatum. A single dose of ANR2B was injected unilaterally into the rat neostriatum. After one day of injection, no modification of motor behavior was found in the ANR2B-injected rats. The mRNA level of NR2B in the ANR2B-injected neostriatum was found to be decreased (-20.4%) by reverse transcriptase polymerase chain reaction (RT-PCR). However, the mRNA levels of NR1, NR2A, NR2C and NR2D in the ANR2B-treated neostriatum were found to be unchanged. After two days of injection, NR2B immunoreactivity was found to decrease in the ANR2B-treated neostriatum by immunofluorescence (-35.1%). At higher magnification, NR2B immunoreactivity was found to decrease in presumed spiny neurons of the neostriatum (-23.4%). No change in NR1 immunoreactivity was observed. These results indicate that a single dose of ANR2B can successfully block the gene expression of NR2B in neurons of the neostriatum and there is less effect on NR1 and other NR2 subunits. The blockage of the gene expression of NR2B is therefore specific and the present results may provide important implications in applications of antisense in research and in clinical therapy of neurological diseases. © 2001 Elsevier Science Ltd. All rights reserved.link_to_subscribed_fulltex