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Methylmercury (MeHg), the causal substrate in Minamata disease, can lead to severe and chronic neurological disorders. The main symptom of Minamata disease is sensory impairment in the four extremities; however, the sensitivity of individual sensory modalities to MeHg has not been investigated extensively. In the present study, we performed stimulus-response behavioral experiments in MeHg-exposed rats to compare the sensitivities to pain, heat, cold, and mechanical sensations. MeHg (6.7 mg/kg/day) was orally administered to 9-week-old Wistar rats for 5 days and discontinued for 2 days, then administered daily for another 5 days. The four behavioral experiments were performed daily on each rat from the beginning of MeHg treatment for 68 days. The pain sensation decreased significantly from day 11 onwards, but recovered to control levels on day 48. Other sensory modalities were not affected by MeHg exposure. These findings suggest that the pain sensation is the sensory modality most susceptive to MeHg toxicity and that this sensitivity is reversible following discontinuation of the exposure. [ABSTRACT FROM AUTHOR]

Methylmercury (MeHg) exposure is known to induce neurodegeneration in both the central nervous system (CNS) and peripheral nervous system (PNS). Molecular mechanisms of MeHg-induced neurotoxicity have been well investigated in the CNS, however, it remains unclear in the PNS. In the present study, comprehensive gene expression analysis was performed by analyzing MeHg-exposed adult rat dorsal root ganglion (DRG) by DNA microarray. Methylmercuric chloride (6.7 mg/kg/day) was administered to nine-week-old male Wistar rats for five days, followed by two days without administration; this cycle was repeated once. Rats were anesthetized at 7 or 14 days after commencement of MeHg exposure, and their DRGs were removed and homogenized to make total RNA samples. DNA microarray data from Day 7 samples identified 100 out of 18,513 detected genes as annotated genes with more than two-fold upregulated or downregulated expression compared with controls. Database for Annotation, Visualization, and Integrated Discovery (DAVID) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses suggested strong involvement of immune activation and inflammation pathways in rat DRG exposed to MeHg, and some genes overlapped with previously reported genes affected by MeHg exposure in the cerebellum. The present results suggest that MeHg-induced neurotoxicity is associated with immune activation and inflammatory responses in rat DRG. [ABSTRACT FROM AUTHOR]

Exposure to organic mercury, especially methylmercury (MeHg), causes Minamata disease, a severe chronic neurological disorder. Minamata disease predominantly affects the central nervous system, and therefore, studies on the mechanisms of MeHg neurotoxicity have focused primarily on the brain. Although the peripheral nervous system is also affected by the organometallic compound and shows signs of neural degeneration, the mechanisms of peripheral MeHg neurotoxicity remain unclear. In the present study, we performed quantitative immunohistochemical analyses of the dorsal root ganglion (DRG) and associated sensory and motor fibers to clarify the mechanisms of MeHg-induced peripheral neurotoxicity in Wistar rats. Methylmercury chloride (6.7 mg/kg/day) was orally administrated for 5 days, followed by 2 days without administration, and this cycle was repeated once again. Seven and 14 days after the beginning of MeHg exposure, rats were anesthetized, and their DRGs and sensory and motor nerve fibers were removed and processed for immunohistochemical analyses. The frozen sections were immunostained for neuronal, Schwann cell, microglial and macrophage markers. DRG sensory neuron somata and axons showed significant degeneration on day 14. At the same time, an accumulation of microglia and the infiltration of macrophages were observed in the DRGs and sensory nerve fibers. In addition, MeHg caused significant Schwann cell proliferation in the sensory nerve fibers. In comparison, there was no noticeable change in the motor fibers. Our findings suggest that in the peripheral nervous system, MeHg toxicity is associated with neurodegenerative changes to DRG sensory neurons and the induction of a neuroprotective and/or enhancement of neurodegenerative host response. [ABSTRACT FROM AUTHOR]

Photodynamic therapy (PDT) is a Food and Drug Administration authorized method for cancer treatment, which uses photosensitizer and laser photo-irradiation to generate reactive oxygen species to induce cell death in tumors. Photosensitizers have been progressively developed, from first to third generation, with improvements in cell specificity, reduced side effects and toxicity, increased sensitivity for irradiation and reduced persistence of photosensitizer in healthy cells. These improvements have been achieved by basic comparative experiments between current and novel photosensitizers using cell lines; however, photosensitizers should be carefully evaluated because they may have cell type specificity. In the present study, we compared a third-generation photosensitizer, ß-mannose-conjugated chlorin (ß-Mchlorin), with the second generation, talaporfin sodium (NPe6), using seven different rat and human cell lines and a neuronal/glial primary culture prepared from rat embryos. NPe6 was more effective than ß-Mchlorin in human-derived cell lines, and ß-M-chlorin was more effective than NPe6 in rat primary cultures and rat-derived cell lines, except for the rat pheochromocytoma cell line, PC12. These differences of phototoxicity in different cell types are not because of differences in photosensitivity between the photosensitizers, but rather are associated with different distribution and accumulation rates in the different cell types. These data suggest that evaluation of photosensitizers for PDT should be carried out using as large a variety of cell types as possible because each photosensitizer may have cell type specificity [ABSTRACT FROM AUTHOR]

Methylmercury is an environmental polluting organometallic compound that exhibits neurotoxicity, as observed in Minamata disease patients. Methylmercury damages peripheral nerves in Minamata patients, causing more damage to sensory nerves than motor nerves. Peripheral nerves are composed of three cell types: dorsal root ganglion (DRG) cells, anterior horn cells (AHCs), and Schwann cells. In this study, we compared cultured these three cell types derived from the rat for susceptibility to methylmercury cytotoxicity, intracellular accumulation of mercury, expression of L-type amino acid transporter 1 (LAT1), which transports methylmercury into cells, and expression of multidrug resistanceassociated protein 2 (MRP2), which transports methylmercury-glutathione conjugates into the extracellular space. Of the cells examined, we found that DRG cells were the most susceptible to methylmercury with markedly higher intracellular accumulation of mercury. The constitutive level of LAT1 was higher and that of MRP2 lower in DRG cells compared with those in AHC and Schwann cells. Additionally, decreased cell viability caused by methylmercury was significantly reduced by either the LAT1 inhibitor, JPH203, or siRNA-mediated knockdown of LAT1. On the other hand, an MRP2 inhibitor, MK571, significantly intensified the decrease in the cell viability caused by methylmercury. Our results provide a cellular basis for sensory neve predominant injury in the peripheral nerves of Minamata disease patients. [ABSTRACT FROM AUTHOR]

Vascular endothelial cells serve as barriers between blood components and subendothelial tissue and regulate the blood coagulation-fibrinolytic system. Ionizing radiation is a common physical stimulant that induces a bystander effect whereby irradiated cells influence neighboring cells through signalings, including purinergic receptor signaling, activated by adenosine 5′-triphosphate (ATP), adenosine 5′-diphosphate (ADP), and adenosine as secondary soluble factors. Human vascular endothelial EA.hy926 cells were cultured and irradiated with γ-rays or treated with ATP, ADP, or adenosine under non-toxic conditions. RNA-seq, gene ontology, and hierarchical clustering analyses were performed. The transcriptome analysis of differentially expressed genes in vascular endothelial cells after γ-ray irradiations suggests that the change of gene expression by γ-irradiation is mediated by ATP and ADP. In addition, the expression and activity of the proteins related to blood coagulation and fibrinolysis systems appear to be secondarily regulated by ATP and ADP in vascular endothelial cells after exposure to γ-irradiation. Although it is unclear whether the changes of the gene expression related to blood coagulation and fibrinolysis systems by γ-irradiation affected the increased hemorrhagic tendency through the exposure to γ-irradiation or the negative feedback to the activated blood coagulation system, the present data indicate that toxicity associated with γ-irradiation involves the dysfunction of vascular endothelial cells related to the blood coagulation-fibrinolytic system, which is mediated by the signalings, including purinergic receptor signaling, activated by ATP and ADP. [ABSTRACT FROM AUTHOR]

Granule cell-selective toxicity of methylmercury in the cerebellum is one of the main unresolved issues in the pathogenesis of Minamata disease. Rats were orally administered methylmercury chloride (10 mg/kg/day) for 5 consecutive days, and their brains were harvested on days 1, 7, 14, 21, or 28 after the last administration for histological examination of the cerebellum. It was found that methylmercury caused a marked degenerative change to the granule cell layers but not to the Purkinje cell layers. The generative change of the granule cell layer was due to cell death, including apoptosis, which occurred at day 21 and beyond after the methylmercury administration. Meanwhile, cytotoxic T-lymphocytes and macrophages had infiltrated the granule cell layer. Additionally, granule cells are shown to be a cell type susceptible to TNF-α. Taken together, these results suggest that methylmercury causes small-scale damage to granule cells, triggering the infiltration of cytotoxic T-lymphocytes and macrophages into the granule cell layer, which secrete tumor necrosis factor-a (TNF-α) to induce apoptosis in granule cells. This chain is established based on the susceptibility of granule cells to methylmercury, the ability of cytotoxic T lymphocytes and macrophages to synthesize and secrete TNF-α, and the sensitivity of granule cells to TNF-α and methylmercury. We propose to call the pathology of methylmercury-induced cerebellar damage the "inflammation hypothesis". [ABSTRACT FROM AUTHOR]

Blood coagulation and the fibrinolytic system contribute to vascular lesions. Fibrinolysis in normal circulating blood strongly depends on the balance between tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) secreted from vascular endothelial cells; however, the mechanisms by which endothelial fibrinolysis is regulated remain to be fully understood. In the present study, human vascular endothelial EA.hy926 cells were transfected with small interfering RNA for nuclear factor erythroid 2-related factor 2 (NRF2) and the expression of t-PA and PAI-1 and fibrinolytic activity in the conditioned medium were examined. EA.hy926 cells were also treated with sulforaphane, an NRF2 activator, and fibrinolytic activity was examined to confirm the NRF2 signaling pathway's effect. Enhanced fibrinolytic activity in the conditioned medium was observed in association with increased expression and secretion levels of t-PA in NRF2 knockdown EA.hy926 cells. However, sulforaphane inhibited fibrinolytic activity and t-PA synthesis in EA.hy926 cells without any cell damage. The expression level of PAI-1 did not change in either NRF2 knockdown or sulforaphane treated cells. These results suggest that transcription factor NRF2 may play a role in down-regulating endothelial t-PA synthesis and fibrinolytic activity. [ABSTRACT FROM AUTHOR]

Photodynamic therapy (PDT) using talaporfin sodium (TS) is tumor cell-selective less invasive therapy for the treatment of malignant glioma. We previously demonstrated that PDT using TS (TS-PDT) treatment exhibits anti-tumor activity against not only glioblastoma cells but also malignant meningioma cells. In general, various stress response proteins have been reported to affect the sensitivity determination for anticancer agents against tumor cells. However, the relationship between the therapeutic effect of TS-PDT and stress response systems in tumor cells is not adequately investigated. In this study, we investigated the gene expression of stress response proteins, including Sod1, Cat1, Gstp1, Gpx1, Nqo1, and Hmox1, in rat malignant meningioma KMY-J cells after treatment of TS-PDT. TS-PDT treatment significantly decreased the cell viability when compared with the no laser irradiation group. In morphological observation, TS at 25.6 μM treatment exhibited a significant cytotoxic effect after 12 hr of laser irradiation to KMY-J cells. After 3 and 6 hr of TS-PDT treatment, mRNA expression of heme oxygenase-1 (HO-1, encoded by Hmox1) was significantly increased by TS-PDT treatment. We also demonstrated that zinc protoporphyrin IX (ZnPPIX), a HO-1 inhibitor, significantly augmented the cytotoxic effect of TS-PDT treatment. These data suggest that HO-1 induction may contribute to a protective response against TS-PDT treatment in the malignant meningioma cells and may attenuate the therapeutic effect for TS-PDT treatment. [ABSTRACT FROM AUTHOR]

Citrinin, a natural mycotoxin that is found in fermented foods, is known as a cytotoxin and nephrotoxin. Exposure to high doses of citrinin result in apoptosis; however, the effects of low doses are not fully understood. Glutamate excitotoxicity is responsible for neuronal death in acute neurological disorders including stroke, trauma and other neurodegenerative diseases. Here, we show the neuroprotective effect of low doses of citrinin against glutamate-induced excitotoxicity. We examined the effect of citrinin exposure on glutamate-induced cell death in cultured rat cortical neurons under two conditions: simultaneous treatment with citrinin 0.1 to 1,000 nM and glutamate (30 μM) for 1, 3 hr; the same simultaneous treatment for 3 hr after pretreatment with citrinin for 21 hr. Both the MTT and immunocytochemical assay showed significant neuroprotective effects at several doses and exposure times tested. All concentrations of citrinin tested showed no remarkable cell death following 14-day exposure, and no marked alterations to synapses. These data suggest that low doses of citrinin can be used as a neuroprotective agent against glutamate-induced excitotoxicity without additional harmful cellular alterations. [ABSTRACT FROM AUTHOR]