TRPM2 contributes to LPS/IFNγ-induced production of nitric oxide via the p38/JNK pathway in microglia

Miyake, Takahito; Shirakawa, Hisashi; Kusano, Ayaka; Sakimoto, Shinya; Konno, Masakazu; Nakagawa, Takayuki; Mori, Yasuo; Kaneko, Shuji

doi:10.1016/j.bbrc.2014.01.022

Cited by 61 publications

(49 citation statements)

References 30 publications

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“…Moreover, activated amoeboid microglia in the developing CNS can be additionally stimulated by pathological stimuli or nerve injury, inducing a greater activation degree (over-activation) that gives rise to the massive production of cytokines and ROS, resulting in alterations of normal development. In this respect, the present results show that LPS treatment of in vitro cultured E8 retina explants induces iNOS upregulation in amoeboid microglia, as demonstrated by anti-iNOS immunolabeling, western-blot, and RT-PCR, suggesting that their activation level increases in response to LPS, a classic stimulant of microglial activation and inductor of iNOS that is frequently used in in vitro [19], [50], [70], [84], [101], [102] and in vivo [52], [103]–[105] studies.…”

Section: Discussionsupporting

confidence: 64%

Expression of Inducible Nitric Oxide Synthase (iNOS) in Microglia of the Developing Quail Retina

et al. 2014

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Inducible nitric oxide synthase (iNOS), which produce large amounts of nitric oxide (NO), is induced in macrophages and microglia in response to inflammatory mediators such as LPS and cytokines. Although iNOS is mainly expressed by microglia that become activated in different pathological and experimental situations, it was recently reported that undifferentiated amoeboid microglia can also express iNOS during normal development. The aim of this study was to investigate the pattern of iNOS expression in microglial cells during normal development and after their activation with LPS by using the quail retina as model. iNOS expression was analyzed by iNOS immunolabeling, western-blot, and RT-PCR. NO production was determined by using DAR-4M AM, a reliable fluorescent indicator of subcellular NO production by iNOS. Embryonic, postnatal, and adult in situ quail retinas were used to analyze the pattern of iNOS expression in microglial cells during normal development. iNOS expression and NO production in LPS-treated microglial cells were investigated by an in vitro approach based on organotypic cultures of E8 retinas, in which microglial cell behavior is similar to that of the in situ retina, as previously demonstrated in our laboratory. We show here that amoeboid microglia in the quail retina express iNOS during normal development. This expression is stronger in microglial cells migrating tangentially in the vitreal part of the retina and is downregulated, albeit maintained, when microglia differentiate and become ramified. LPS treatment of retina explants also induces changes in the morphology of amoeboid microglia compatible with their activation, increasing their lysosomal compartment and upregulating iNOS expression with a concomitant production of NO. Taken together, our findings demonstrate that immature microglial cells express iNOS during normal development, suggesting a certain degree of activation. Furthermore, LPS treatment induces overactivation of amoeboid microglia, resulting in a significant iNOS upregulation.

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Section: Discussionsupporting

confidence: 64%

Expression of Inducible Nitric Oxide Synthase (iNOS) in Microglia of the Developing Quail Retina

et al. 2014

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“…We started with using immunofluorescent confocal microscopy to confirm the TRPM2 expression in microglial cells7172737475. Positive immunostaining was observed in cells labelled with an anti-TRPM2 antibody but not in control cells (Fig.…”

Section: Resultsmentioning

confidence: 99%

Signalling mechanisms mediating Zn2+-induced TRPM2 channel activation and cell death in microglial cells

Mortadza

Sim

Stacey

et al. 2017

Sci Rep

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Excessive Zn2+ causes brain damage via promoting ROS generation. Here we investigated the role of ROS-sensitive TRPM2 channel in H2O2/Zn2+-induced Ca2+ signalling and cell death in microglial cells. H2O2/Zn2+ induced concentration-dependent increases in cytosolic Ca2+ concentration ([Ca2+]c), which was inhibited by PJ34, a PARP inhibitor, and abolished by TRPM2 knockout (TRPM2-KO). Pathological concentrations of H2O2/Zn2+ induced substantial cell death that was inhibited by PJ34 and DPQ, PARP inhibitors, 2-APB, a TRPM2 channel inhibitor, and prevented by TRPM2-KO. Further analysis indicate that Zn2+ induced ROS production, PARP-1 stimulation, increase in the [Ca2+]c and cell death, all of which were suppressed by chelerythrine, a protein kinase C inhibitor, DPI, a NADPH-dependent oxidase (NOX) inhibitor, GKT137831, a NOX1/4 inhibitor, and Phox-I2, a NOX2 inhibitor. Furthermore, Zn2+-induced PARP-1 stimulation, increase in the [Ca2+]c and cell death were inhibited by PF431396, a Ca2+-sensitive PYK2 inhibitor, and U0126, a MEK/ERK inhibitor. Taken together, our study shows PKC/NOX-mediated ROS generation and PARP-1 activation as an important mechanism in Zn2+-induced TRPM2 channel activation and, TRPM2-mediated increase in the [Ca2+]c to trigger the PYK2/MEK/ERK signalling pathway as a positive feedback mechanism that amplifies the TRPM2 channel activation. Activation of these TRPM2-depenent signalling mechanisms ultimately drives Zn2+-induced Ca2+ overloading and cell death.

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“…Intracellular Ca 2+ influx induced by oxidative stress has a significant role in the etiology of SCI, and studies recently reported the important role of TRPM2 and TRPV1 channels in converting oxidant radical signaling to cytosolic calcium ion homeostasis in neuropathic pain and SCI [2,3,14]. Results of recent studies suggest that TRPM2 and TRPV1 activations are increased by NADPH oxidase pathways [2,15] and TRPM2 activity through modulation of NADPH oxidase pathways is reduced in DRG of rats and neutrophils of patients with multiple sclerosis by Hypericum perforatum (HP) treatment [16][17][18]. Therefore, HP may modulate oxidative stress-induced TRPM2 and TRPV1 channel activation in SCI-induced DRG neurons and this possibility is the main reason for undertaking the current study.…”

Section: Introductionmentioning

confidence: 97%

Hypericum perforatum Attenuates Spinal Cord Injury-Induced Oxidative Stress and Apoptosis in the Dorsal Root Ganglion of Rats: Involvement of TRPM2 and TRPV1 Channels

et al. 2015

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Oxidative stress and cytosolic Ca(2+) overload have important roles on apoptosis in dorsal root ganglion (DRG) neurons after spinal cord injury (SCI). Hypericum perforatum (HP) has an antioxidant property in the DRGs due to its ability to modulate NADPH oxidase and protein kinase C pathways. We aimed to investigate the protective property of HP on oxidative stress, apoptosis, and Ca(2+) entry through transient receptor potential melastatin 2 (TRPM2) and transient receptor potential vanilloid 1 (TRPV1) channels in SCI-induced DRG neurons of rats. Rats were divided into four groups as control, HP, SCI, and SCI + HP. The HP groups received 30 mg/kg HP for three concessive days after SCI induction. The SCI-induced TRPM2 and TRPV1 currents and cytosolic free Ca(2+) concentration were reduced by HP. The SCI-induced decrease in glutathione peroxidase and cell viability values were ameliorated by HP treatment, and the SCI-induced increase in apoptosis, caspase 3, caspase 9, cytosolic reactive oxygen species (ROS) production, and mitochondrial membrane depolarization values in DRG of SCI group were overcome by HP treatment. In conclusion, we observed a protective role of HP on SCI-induced oxidative stress, apoptosis, and Ca(2+) entry through TRPM2 and TRPV1 in the DRG neurons. Our findings may be relevant to the etiology and treatment of SCI by HP. Graphical Abstract Possible molecular pathways of involvement of Hypericum perforatum (HP) on apoptosis, oxidative stress, and calcium accumulation through TRPM2 and TRPV1 channels in DRG neurons of SCI-induced rats. The TRPM2 channel is activated by ADP-ribose and oxidative stress through activation of ADP-ribose pyrophosphate although it was inhibited by N-(p-amylcinnamoyl) anthranilic acid (ACA) and 2-aminoethyl diphenylborinate (2APB). The TRPV1 channel is activated by oxidative stress and capsaicin and it is blocked by capsazepine. Injury in the DRG can result in augmented ROS release, leading to Ca(2+) uptake through TRPM2 and TRPV1 channels. Mitochondria were reported to accumulate Ca(2+), provided intracellular Ca(2+) rises, thereby leading to depolarization of mitochondrial membranes and release of apoptosis-inducing factors such as caspase 3 and caspase 9. HP via regulation of NADPH oxidase and PKC inhibits TRPM2 and TRPV1 channels. The molecular pathway may be a cause of SCI-induced pain and neuronal death, and the subject should be urgently investigated.

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TRPM2 contributes to LPS/IFNγ-induced production of nitric oxide via the p38/JNK pathway in microglia

Cited by 61 publications

References 30 publications

Expression of Inducible Nitric Oxide Synthase (iNOS) in Microglia of the Developing Quail Retina

Expression of Inducible Nitric Oxide Synthase (iNOS) in Microglia of the Developing Quail Retina

Signalling mechanisms mediating Zn2+-induced TRPM2 channel activation and cell death in microglial cells

Hypericum perforatum Attenuates Spinal Cord Injury-Induced Oxidative Stress and Apoptosis in the Dorsal Root Ganglion of Rats: Involvement of TRPM2 and TRPV1 Channels

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