Resveratrol diminishes bisphenol A-induced oxidative stress through TRPM2 channel in the mouse kidney cortical collecting duct cells.

Bisphenol A (BisPH-A) is a latent danger that threatens our health, which we frequently exposure in our modern life (e.g. the widespread use of drinking water in plastic pet bottles). But the BisPH-A induced transient receptor potential melastatin 2 (TRPM2)-mediated oxidative stress and apoptosis in these cells has not been studied yet. Calcium (Ca2+) plays an important role in a versatile intracellular signal transduction that works over a wide range to regulate oxidative stress processes. TRPM2 is activated by oxidative stress and it has emerged as an important Ca2+ signaling mechanism in a variety of cells, contributing many cellular functions including cell death. Resveratrol (RESV), which belongs to the polyphenol group, acts as an antioxidant, eliminating cellular oxidative stress and increasing the body's resistance to diseases. The current study aimed to elucidate the effect of antioxidant resveratrol on TRPM2-mediated oxidative stress induced by BisPH-A exposure in the mouse kidney cortical collecting duct cells (mpkCCDcl4). The cells were divided into four groups as control, resveratrol (50 µM for 24 h), BisPH-A (100 µM for 24 h) and BisPH-A + RESV. Intracellular free Ca2+ concentrations and TRPM2 channel currents were high in BisPH-A treated cells, but decreased with resveratrol treatment. In addition, BisPH-A induced mitochondrial membrane depolarization, reactive oxygen species (ROS), caspase 3, caspase 9 and apoptosis values were decreased by the resveratrol treatment. In conclusion, resveratrol protected cells from BisPH-A induced oxidative damage. In this study, we showed that TRPM2 channel mediates this protective effect of resveratrol.

Inhibitions of anandamide transport and FAAH synthesis decrease apoptosis and oxidative stress through inhibition of TRPV1 channel in an in vitro seizure model.

The expression level of TRPV1 is high in hippocampus which is a main epileptic area in the brain. In addition to the actions of capsaicin (CAP) and reactive oxygen species (ROS), the TRPV1 channel is activated in neurons by endogenous cannabinoid, anandamide (AEA). In the current study, we investigated the role of inhibitors of TRPV1 (capsazepine, CPZ), AEA transport (AM404), and FAAH (URB597) on the modulation of Ca2+ entry, apoptosis, and oxidative stress in in vitro seizure-induced rat hippocampus and human glioblastoma (DBTRG) cell line. The seizure was induced in the hippocampal and DBTRG neurons using in vitro 4-aminopyridine (4-AP) to trigger a seizure-like activity model. CPZ and AM404 were fully effective in reversing 4-AP-induced intracellular free Ca2+ concentration of the hippocampus and TRPV1 current density of DBTRG. However, AEA and CAP did not activate TRPV1 in the URB597-treated neurons. Hence, we observed TRPV1 blocker effects of URB597 in the DBTRG neurons. In addition, the AM404 and CPZ treatments decreased intracellular ROS production, mitochondrial membrane depolarization, apoptosis, caspases 3 and 9 values in the hippocampus. In conclusion, the results indicate that inhibition of AEA transport, FAAH synthesis, and TRPV1 activity can result in remarkable neuroprotective effects in the epileptic neurons. Possible molecular pathways of involvement of capsazepine (CPZ) and AM4040 in anandamide and capsaicin (CAP)-induced apoptosis, oxidative stress, and Ca2+ accumulation through TRPV1 channel in the seizure-induced rat hippocampus and human glioblastoma neurons. The TRPV1 channel is activated by different stimuli including reactive oxygen species (ROS), anandamide (AEA), and CAP and it is blocked by capsazepine (CPZ). Cannabinoid receptor type 1 (CB1) is also activated by AEA. The AEA levels in cytosol are decreased by fatty acid amide hydrolase (FAAH) enzyme. Inhibition of FAAH through URB597 induces stimulation of CB1 receptor through accumulation AEA. URB597 acts antiepileptic effects through inhibition of TRPV1. Overloaded Ca2+ concentration of mitochondria can induce an apoptotic program by stimulating the release of apoptosis-promoting factors such as caspases 3 and caspase 9 by generating ROS due to respiratory chain damage. AM404 and CPZ reduce TRPV1 channel activation and Ca2+ entry in the in vitro 4-AP seizure model-induced hippocampal and glioblastoma neurons.

Calorie restriction protects against apoptosis, mitochondrial oxidative stress and increased calcium signaling through inhibition of TRPV1 channel in the hippocampus and dorsal root ganglion of rats.

The TRPV1 channel is activated in neurons by capsaicin, oxidative stress, acidic pH and heat factors, and these factors are attenuated by the antioxidant role of calorie restriction (CR). Hence, we investigated the hypothesis that the antioxidant roles of CR and food frequency (FF) may modulate TRPV1 activity and apoptosis through inhibition of mitochondrial oxidative stress in hippocampal (HIPPON) and dorsal root ganglion neurons (DRGN). We investigated the contribution of FF and CR to neuronal injury and apoptosis through inhibition of TRPV1 in rats. We assigned rats to control, FF and FF + CR groups. A fixed amount of food ad libitum was supplemented to the control and FF groups for 20 weeks, respectively. FF + CR group were fed the same amount of food as the control group but with 20% less calories during the same period. In major results, TRPV1 currents, intracellular Ca2+ levels, apoptosis, reactive oxygen species, mitochondrial depolarization, PARP-1 expression, caspase 3 and 9 activity and expression values were found to be increased in the HIPPON and DRGN following FF treatment, and these effects were decreased following FF + CR treatment. The FF-induced decrease in cell viability of HIPPO and DRGN, and vitamin E concentration of brain, glutathione peroxidase, vitamin A, and ß-carotene values of the HIPPO, DRGN, plasma, liver and kidney were increased by FF + DR treatment, although lipid peroxidation levels in the same samples were decreased. In conclusion, CR reduces FF-induced increase of oxidative stress, apoptosis and Ca2+ entry through TRPV1 in the HIPPON and DRGN. Our findings may be relevant to the etiology and treatment of obesity following CR treatment.

Selenium potentiates the anticancer effect of cisplatin against oxidative stress and calcium ion signaling-induced intracellular toxicity in MCF-7 breast cancer cells: involvement of the TRPV1 channel.

BACKGROUND: In breast cancers, calcium signaling is a main cause of proliferation and apoptosis of breast cancer cells. Although previous studies have implicated the transient receptor potential vanilloid 1 (TRPV1) cation channel, the synergistic inhibition effects of selenium (Se) and cisplatin in cancer and the suppression of ongoing apoptosis have not yet been investigated in MCF-7 breast cancer cells. This study investigates the anticancer properties of Se through TRPV1 channel activity in MCF-7 breast cancer cell line cultures when given alone or in combination with cisplatin. MATERIALS: The MCF-7 cells were divided into four groups: the control group, the Se-treated group (200 nM), the cisplatin-treated group (40 µM) and the Se + cisplatin-treated group. RESULTS: The intracellular free calcium ion concentration and current densities increased with TRPV1 channel activator capsaicin (0.01 mM), but they decreased with the TRPV1 blocker capsazepine (0.1 mM), Se, cisplatin, and Se + cisplatin incubations. However, mitochondrial membrane depolarization, apoptosis, and the caspase 3, and caspase 9 values increased in the Se-treated group and the cisplatin-treated group, although Western blot (procaspase 3 and 9) results and the cell viability levels decreased with the Se and Se + cisplatin treatments. Apoptosis and caspase-3 were further increased with the Se + cisplatin treatment. Intracellular reactive oxygen species production increased with the cisplatin treatment, but not with the Se treatment. CONCLUSION: This study's results report, for the first time, that at a cellular level, Se and cisplatin interact on the same intracellular toxic cascade, and the combination of these two drugs can result in a remarkable anticancer effect through modulation of the TRPV1.

Investigation of the effects of distance from sources on apoptosis, oxidative stress and cytosolic calcium accumulation via TRPV1 channels induced by mobile phones and Wi-Fi in breast cancer cells.

TRPV1 is a Ca2+ permeable channel and gated by noxious heat, oxidative stress and capsaicin (CAP). Some reports have indicated that non-ionized electromagnetic radiation (EMR)-induces heat and oxidative stress effects. We aimed to investigate the effects of distance from sources on calcium signaling, cytosolic ROS production, cell viability, apoptosis, plus caspase-3 and -9 values induced by mobile phones and Wi-Fi in breast cancer cells MCF-7 human breast cancer cell lines were divided into A, B, C and D groups as control, 900, 1800 and 2450 MHz groups, respectively. Cells in Group A were used as control and were kept in cell culture conditions without EMR exposure. Groups B, C and D were exposed to the EMR frequencies at different distances (0 cm, 1 cm, 5 cm, 10 cm, 20 cm and 25 cm) for 1h before CAP stimulation. The cytosolic ROS production, Ca2+ concentrations, apoptosis, caspase-3 and caspase-9 values were higher in groups B, C and D than in A group at 0 cm, 1 cm and 5 cm distances although cell viability (MTT) values were increased by the distances. There was no statistically significant difference in the values between control, 20 and 25 cm. Wi-Fi and mobile phone EMR placed within 10 cm of the cells induced excessive oxidative responses and apoptosis via TRPV1-induced cytosolic Ca2+ accumulation in the cancer cells. Using cell phones and Wi-Fi sources which are farther away than 10 cm may provide useful protection against oxidative stress, apoptosis and overload of intracellular Ca2+. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers.

Synergic Effects of Doxorubicin and Melatonin on Apoptosis and Mitochondrial Oxidative Stress in MCF-7 Breast Cancer Cells: Involvement of TRPV1 Channels.

Transient receptor transient receptor potential vanilloid 1 (TRPV1) is a Ca(2+)-permeable channel gated by oxidative stress and capsaicin (CAP) and modulated by melatonin (MEL) and capsazepine (CPZ). A combination of doxorubicin (DOX) and MEL may offer a potential therapy for breast cancer by exerting antitumor and anti-apoptotic effects and modulating Ca(2+) influx and TRPV1 activity. We aimed to investigate the effects of MEL and DOX on the oxidative toxicity of MCF-7 human breast cancer cells, in addition to the activity of the TRPV1 channel and apoptosis. The MCF-7 cells were divided into the following six treatment groups: control, incubated with MEL (0.3 mM), incubated with 0.5 µM DOX, incubated with 1 µM DOX, incubated with MEL + 0.5 µM DOX, or incubated with MEL + 1 µM DOX. The intracellular free Ca(2+) concentration was higher in the DOX groups than in the control, and the concentration was decreased by MEL. The intracellular free Ca(2+) concentration was further increased by treatment with the TRPV1 channel activator CAP (0.01 mM), and it was decreased by the CPZ (0.1 mM). The intracellular production of reactive oxygen species, mitochondrial membrane depolarization, apoptosis level, procaspase 9 and PARP activities, and caspase 3 and caspase 9 activities were higher in the DOX and MEL groups than in the control. Apoptosis and the activity of caspase 9 were further increased in the DOX plus MEL groups. Taken together, the findings indicate that MEL supported the effects of DOX by activation of TRPV1 and apoptosis, as well as by inducing MCF-7 cell death. As the apoptosis and caspase activity of cancer cells increase because of their elevated metabolism, MEL may be useful in supporting their apoptotic capacity.

Epilepsy but not mobile phone frequency (900 MHz) induces apoptosis and calcium entry in hippocampus of epileptic rat: involvement of TRPV1 channels.

Electromagnetic radiation (EMR) and epilepsy are reported to mediate the regulation of apoptosis and oxidative stress through Ca(2+) influx. Results of recent reports indicated that EMR can increase temperature and oxidative stress of body cells, and TRPV1 channel is activated by noxious heat, oxidative stress, and capsaicin (CAP). We investigated the effects of mobile phone (900 MHz) EMR exposure on Ca(2+) influx, apoptosis, oxidative stress, and TRPV1 channel activations in the hippocampus of pentylenetetrazol (PTZ)-induced epileptic rats. Freshly isolated hippocampal neurons of twenty-one rats were used in study within three groups namely control, PTZ, and PTZ + EMR. The neurons in the three groups were stimulated by CAP. Epilepsy was induced by PTZ administration. The neurons in PTZ + EMR group were exposed to the 900 MHz EMR for 1 h. The apoptosis, mitochondrial membrane depolarization, intracellular reactive oxygen species (ROS), and caspase-3 and caspase-9 values were higher in PTZ and PTZ + EMR groups than in control. However, EMR did not add additional increase effects on the values in the hippocampal neurons. Intracellular-free Ca(2+) concentrations in fura-2 analyses were also higher in PTZ + CAP group than in control although their concentrations were decreased by TRPV1 channel blocker, capsazepine. However, there were no statistical changes on the Ca(2+) concentrations between epilepsy and EMR groups. In conclusion, apoptosis, mitochondrial, ROS, and Ca(2+) influx via TRPV1 channel were increased in the hippocampal neurons by epilepsy induction although the mobile phone did not change the values. The results indicated that TRPV1 channels in hippocampus may possibly be a novel target for effective target of epilepsy.

The efficiency of Poly(ADP-Ribose) Polymerase (PARP) cleavage on detection of apoptosis in an experimental model of testicular torsion.

The aim of this study was to evaluate the histopathological and apoptotic changes occurring in the rat ipsilateral and contralateral testes, after experimental spermatic cord torsion, and to explore and the role of poly(ADP-ribose) polymerase (PARP) cleavage in testicular torsion-detorsion injury. A total of 37 Wistar albino rats were subjected to 720° unilateral spermatic cord torsion for 1, 2 and 4 h, followed by 4-h reperfusion, or else to a sham operation (control group). Histology of the testicle was evaluated using haematoxylin-eosin (H&E) staining and Johnsen's scoring system. Germ cell apoptosis was evaluated via active caspase-3 immunostaining, and PARP expression levels were evaluated via Western blotting. The mean Johnsen's tubular biopsy scores (JTBS) of the ipsilateral testicles were lower for all torsion groups than for the controls (P < 0.05), but the JTBS of the contralateral testicles were only lower in the 4-h torsion group (P < 0.05). The mean apoptosis score (AS) of the ipsilateral and contralateral testicles was significantly higher in the torsion groups than in the sham group. AS increased correlatively with torsion time, in both testicles. The effect of testicular torsion on PARP cleavage was time dependent, with the highest effect observed after 4 h of testicular torsion (P < 0.05). Testicular torsion caused time-dependent histological changes, apoptosis and increases in PARP cleavage. Our results suggest that testicular torsion-detorsion injury caused cell damage and germ cell apoptosis that apparently involved cleavage of PARP. Increased PARP cleavage could, in turn, lead to enhanced apoptosis.

Melatonin and selenium reduce plasma cytokine and brain oxidative stress levels in diabetic rats.

BACKGROUND: Hyperglycaemia-induced progression of brain and erythrocyte oxidative injuries might be modulated by melatonin and selenium as potent antioxidants. The present study was conducted to explore whether melatonin and selenium protect against diabetic brain and erythrocyte oxidative stress levels in streptozotocin (STZ)-induced diabetic rats. MATERIALS AND METHODS: Seventy rats were equally divided into seven groups. The first and second groups were used as untreated and placebo treated controls. The third group was treated with STZ to induce diabetes. The fourth and sixth groups received 10 mg kg(-1) melatonin. The fifth and seventh groups were treated with 1.5 mg kg(-1) selenium (sodium selenite). The sixth and seventh groups were treated with STZ administered with melatonin and selenium as described for the fourth and fifth groups. RESULTS: Brain and erythrocyte lipid peroxidation levels and plasma IL-1ß and IL-4 levels were high in the STZ group, although they were low in melatonin and selenium treatments. Decreased glutathione peroxidase, reduced glutathione, total antioxidant status, vitamins A and vitamin E values in brain and erythrocyte of STZ group were increased by melatonin and selenium treatments. DISCUSSION: Melatonin and selenium induced protective effects against diabetes-induced brain and erythrocyte oxidative injuries through regulation of the antioxidant level and cytokine production.

Hypericum perforatum modulates apoptosis and calcium mobilization through voltage-gated and TRPM2 calcium channels in neutrophil of patients with Behcet's disease.

Behcet's disease (BD) is a chronic, inflammatory, and multisystemic condition although its pathogenesis is uncertain. Main component of St. John's wort (Hypericum perforatum, HP) is hyperforin and induces antiinflammatory and antioxidant properties. We aimed to investigate effects of HP on oxidative stress, apoptosis, and cytosolic-free Ca²âº [Ca²âº](i) concentration in neutrophil of BD patients. Nine new-diagnosed active patients with BD and nine control subjects were included in the study. Disease activity was considered by clinical findings. Neutrophil samples were obtained from the patients and controls. The neutrophils from patients were divided into three subgroups and were incubated with HP, voltage-gated calcium channel (VGCC) blockers, (verapamil+dilitiazem) and non-specific TRPM2 channel blocker (2-aminoethyl diphenylborinate, 2-APB), respectively. The neutrophils were stimulated by fMLP as a Ca²âº-concentration agonist and oxidative stress former. Caspase-3, caspase-9, apoptosis, lipid peroxidation, and [Ca²âº](i) values were high in the patient groups, although cell viability, glutathione (GSH), and glutathione peroxidase (GSH-Px) values were low in patient group. However, the [Ca²âº](i), caspase-3, and caspase-9 values decreased markedly in patient+HP group although GSH and GSH-Px values increased in the group. The [Ca²âº](i) concentration was also decreased in the patient group by V+D, 2-APB, and HP incubations. In conclusion, we observed the importance of neutrophil Ca²âº entry, apoptosis, and oxidative stress through gating VGCC and TRPM2 channels in the neutrophils in the pathogenesis and activation of the patients with BD. HP induced protective effects on oxidative stress by modulating Ca²âº influx in BD patients.

Role of TRPM2 cation channels in dorsal root ganglion of rats after experimental spinal cord injury.

INTRODUCTION: We sought to determine the contribution of oxidative stress-dependent activation of TRPM2 and L-type voltage-gated Ca(2+) channels (VGCC) in dorsal root ganglion (DRG) neurons of rats after spinal cord injury (SCI). METHODS: The rats were divided into 4 groups: control; sham control; SCI; and SCI+nimodipine groups. The neurons of the SCI groups were also incubated with non-specific TRPM2 channel blockers, 2-aminoethoxydiphenylborate (2-APB) and N-(p-amylcinnamoyl)anthranilic acid (ACA), before H2 O2 stimulation. RESULTS: The [Ca(2+) ]i concentrations were higher in the SCI group than in the control groups, although their concentrations were decreased by nimodipine and 2-APB. The H2 O2 -induced TRPM2 current densities in patch-clamp experiments were decreased by ACA and 2-APB incubation. In the nimodipine group, the TRPM2 channels of neurons were not activated by H2 O2 or cumene hydroperoxide. CONCLUSIONS: Increased Ca(2+) influx and currents in DRG neurons after spinal injury indicated TRPM2 and voltage-gated Ca(2+) channel activation.

TRP Channels in Oxidative Stress Signalling.

It is well established that the accumulation of high levels of reactive oxygen species (ROS), due to excessive generation of ROS and/or impaired antioxidant capacity of cells, can result in oxidative stress and cause oxidative damage to cells and their functions [...].

Melatonin potentiates chemotherapy-induced cytotoxicity and apoptosis in rat pancreatic tumor cells.

Melatonin has antitumor activity via several mechanisms including its antiproliferative and proapoptotic effects in addition to its potent antioxidant action. Thus, melatonin has proven useful in the treatment of tumors in association with chemotherapeutic drugs. This study was performed to evaluate the effect of melatonin on the cytotoxicity and apoptosis induced by three different chemotherapeutic agents, namely 5-fluorouracil (5-FU), cisplatin, and doxorubicin in the rat pancreatic tumor cell line AR42J. We found that both melatonin and the three chemotherapeutic drugs induce a time-dependent decrease in AR42J cell viability, reaching the highest cytotoxic effect after 48 hr of incubation. Furthermore, melatonin significantly augmented the cytotoxicity of the chemotherapeutic agents. Consistently, cotreatment of AR42J cells with each of the chemotherapeutic agents in the presence of melatonin increased the population of apoptotic cells, elevated mitochondrial membrane depolarization, and augmented intracellular reactive oxygen species (ROS) production compared to treatment with each chemotherapeutic agent alone. These results provide evidence that in vitro melatonin enhances chemotherapy-induced cytotoxicity and apoptosis in rat pancreatic tumor AR42J cells and, therefore, melatonin may be potentially applied to pancreatic tumor treatment as a powerful synergistic agent in combination with chemotherapeutic drugs.

Aminoethoxydiphenyl borate and flufenamic acid inhibit Ca2+ influx through TRPM2 channels in rat dorsal root ganglion neurons activated by ADP-ribose and rotenone.

Exposure to oxidative stress causes health problems, including sensory neuron neuropathy and pain. Rotenone is a toxin used to generate intracellular oxidative stress in neurons. However, the mechanism of toxicity in dorsal root ganglion (DRG) neurons has not been characterized. Melastatin-like transient receptor potential 2 (TRPM2) channel activation and inhibition in response to oxidative stress, ADP-ribose (ADPR), flufenamic acid (FFA) and 2-aminoethoxydiphenyl borate (2-APB) in DRG neurons are also not clear. We tested the effects of FFA and 2-APB on ADPR and rotenone-induced TRPM2 cation channel activation in DRG neurons of rats. DRG neurons were freshly isolated from rats and studied with the conventional whole-cell patch-clamp technique. Rotenone, FFA and 2-APB were extracellularly added through the patch chamber, and ADPR was applied intracellularly through the patch pipette. TRPM2 cation currents were consistently induced by ADPR and rotenone. Current densities of the neurons were higher in the ADPR and rotenone groups than in control. The time courses (gating times) in the neurons were longer in the rotenone than in the ADPR group. ADPR and rotenone-induced TRPM2 currents were totally blocked by 2-APB and partially blocked by FFA. In conclusion, TRPM2 channels were constitutively activated by ADPR and rotenone, and 2-APB and FFA induced an inhibitory effect on TRPM2 cation channel currents in rat DRG neurons. Since oxidative stress is a common feature of neuropathic pain and diseases of sensory neurons, the present findings have broad application to the etiology of neuropathic pain and diseases of DRG neurons.

Glutathione modulates Ca(2+) influx and oxidative toxicity through TRPM2 channel in rat dorsal root ganglion neurons.

Glutathione (GSH) is the most abundant thiol antioxidant in mammalian cells and maintains thiol redox in the cells. GSH depletion has been implicated in the neurobiology of sensory neurons. Because the mechanisms that lead to melastatin-like transient receptor potential 2 (TRPM2) channel activation/inhibition in response to glutathione depletion and 2-aminoethyldiphenyl borinate (2-APB) administration are not understood, we tested the effects of 2-APB and GSH on oxidative stress and buthionine sulfoximine (BSO)-induced TRPM2 cation channel currents in dorsal root ganglion (DRG) neurons of rats. DRG neurons were freshly isolated from rats and the neurons were incubated for 24 h with BSO. In whole-cell patch clamp experiments, TRPM2 currents in the rat were consistently induced by H(2)O(2) or BSO. TRPM2 channels current densities and cytosolic free Ca(2+) content of the neurons were higher in BSO and H(2)O(2) groups than in control. However, the current densities and cytosolic Ca(2+) release were also higher in the BSO + H(2)O(2) group than in the H(2)O(2) alone. When intracellular GSH is introduced by pipette TRPM2 channel currents were not activated by BSO, H(2)O(2) or rotenone. BSO and H(2)O(2)-induced Ca(2+) gates were blocked by the 2-APB. Glutathione peroxidase activity, lipid peroxidation and GSH levels in the DRG neurons were also modulated by GSH and 2-APB inhibition. In conclusion, we observed the protective role of 2-APB and GSH on Ca(2+) influx through a TRPM2 channel in intracellular GSH depleted DRG neurons. Since cytosolic glutathione depletion is a common feature of neuropathic pain and diseases of sensory neuron, our findings are relevant to the etiology of neuropathology in DRG neurons.

Colchicine modulates oxidative stress in serum and neutrophil of patients with Behçet disease through regulation of Ca²âº release and antioxidant system.

Behçet disease (BD) is a chronic, inflammatory, and multisystemic condition with an uncertain pathogenesis. One of the major immunologic findings in BD pathogenesis is increase in activity of neutrophil. An increase in the cytosolic free Ca²âº[Ca²âº](i) concentration that induces Ca²âº signaling is an important step that participates in the neutrophil activation and reactive oxygen species production that leads to tissue damage in body cells. We aimed to investigate the effects of colchicine on oxidative stress and Ca²âº release in serum and neutrophil of BD patients with active and inactive periods. Twelve Behçet patients (6 active and 6 inactive) and 6 control subject were included in the study. Disease activity was considered by clinical findings. Serum and neutrophil samples were obtained from the patients and control subjects. Neutrophils from patients with active BD were divided into three subgroups and were incubated with colchicine, verapamil + diltiazem, and colchicine + verapamil + diltiazem, respectively. Erythrocyte sedimentation rate, leucocytes counts, serum C-reactive protein, neutrophil, and serum lipid peroxidation and intracellular Ca²âº release levels were higher in active and inactive groups than in the control group, although their levels were lower in active group than in inactive group. However, neutrophil Ca²âº release levels were decreased in colchicine, verapamil + diltiazem, and colchicine + verapamil + diltiazem groups group compared to active group. Serum glutathione, vitamin A, vitamin E, and ß-carotene concentrations were lower in active and inactive groups than in the control group, although serum vitamin E and ß-carotene concentrations were higher in the inactive group than in the active group. Neutrophil and serum glutathione peroxidase activity within the three groups did not change. In conclusion, we observed the importance of Ca²âº influx into the neutrophils and oxidative stress in the pathogenesis and activation of the patients with BD. Colchicine induced protective effects on oxidative stress by modulating Ca²âº influx in BD patients.

The protective role of selenium against dental amalgam-induced intracellular oxidative toxicity through the TRPV1 channel in DBTRG glioblastoma cells.

OBJECTIVE: The exposure to mercury (Hg) from dental amalgams is a suspected causative factor in neurological diseases. This study investigated the toxic effects of two different amalgam compositions related to Hg and the protective effects of selenium against the toxic effects of Hg through the TRPV1 channel in the human DBTRG glioblastoma cell line. METHODOLOGY: Six groups of the cells were organized. Analyses of cell viability, apoptosis, caspase 3 and caspase 9 activities, mitochondrial membrane depolarization, reactive oxygen species (ROS) production, and Western Blotting for protein expression levels were performed. RESULTS: Cell viability values were lower in amalgam with high copper (HCu) and low copper (LCu) groups independently of time but were increased by selenium and capsazepine (p<0.001 and p<0.05). Conversely, apoptosis rates, caspase 3 and caspase 9 expression, ROS formation, mitochondrial membrane depolarization, and protein expression levels were higher in the HCu and LCu groups but were decreased by selenium (p<0.001 and p<0.05). CONCLUSIONS: Selenium combined with an amalgam of either HCu or LCu decreases the toxic effects created by Hg in human DBTRG glioblastoma cells.

Noopept Attenuates Diabetes-Mediated Neuropathic Pain and Oxidative Hippocampal Neurotoxicity via Inhibition of TRPV1 Channel in Rats.

Neuropathic pain and oxidative neurotoxicity are two adverse main actions of diabetes mellitus (DM). The expression levels of calcium ion (Ca2+) permeable TRPV1 channels are high in the dorsal root ganglion (DRGs) and hippocampus (HIPPO). TRPV1 is activated by capsaicin and reactive free oxygen radicals (fROS) to mediate peripheral neuropathy and neurotoxicity. Noopept (NP) acted several protective antioxidant actions against oxidative neurotoxicity. As DM is known to increase the levels of fROS, the protective roles of antioxidant NP were evaluated on the DM-mediated neurotoxicity and neuropathic pain via the modulation of TRPV1 in rats. Thirty-six rats were equally divided into control, NP, DM (streptozotocin, STZ), and STZ + NP groups. A decrease on the STZ-mediated increase of neuropathic pain (via the analyses of Von Frey and hot plate) and blood glucose level was observed by the treatment of NP. A protective role of NP via downregulation of TRPV1 activity on the STZ-induced increase of apoptosis, mitochondrial fROS, lipid peroxidation, caspase -3 (CASP-3), caspase -9 (CASP-9), TRPV1 current density, glutathione (GSH), cytosolic free Zn2+, and Ca2+ concentrations in the DRGs and HIPPO was also observed. The STZ-mediated decrease of glutathione peroxidase, GSH, vitamin E, and ß-carotene concentrations in the brain cortex, erythrocyte, liver, kidney, and plasma was also attenuated by the treatment of NP. The STZ-mediated increase of TRPV1, CASP-3, and CASP-9 expressions was decreased in the DRGs and HIPPO by the treatment of NP. In conclusion, the treatment of NP induced protective effects against STZ-induced adverse peripheral pain and HIPPO oxidative neurotoxicity. These effects might attribute to the potent antioxidant property of NP.

Modulator role of infliximab and methotrexate through the transient receptor potential melastatin 2 (TRPM2) channel in neutrophils of patients with rheumatoid arthritis: a pilot study.

INTRODUCTION: Rheumatoid arthritis (RA) is a chronic, systemic, inflammatory disease causing symmetric polyarthritis. In this study, we aimed to investigate the effects of infliximab (INF) and methotrexate (MTX) on apoptosis, oxidative stress, and calcium signaling in the neutrophils of RA patients. MATERIAL AND METHODS: Neutrophils were isolated from 10 patients with newly diagnosed RA and 10 healthy controls. They were divided into four groups (control, RA, RA + MTX, RA + INF) and incubated with MTX and INF. In the cell viability (MTT) test, the ideal non-toxic dose and incubation time of MTX were found to be 0.1 mM and 1 h, respectively. The neutrophils were also incubated with the TRPM2 channel blocker N-(p-amylcinnamoyl) anthranilic acid (ACA). RESULTS: Intracellular free Ca2+ concentration, intracellular reactive oxygen species (ROS) production, mitochondrial depolarization, lipid peroxidation, apoptosis, and caspase 3 and caspase 9 activities were found to be significantly higher in the neutrophils of RA patients compared to controls. MTT, reduced glutathione (GSH) level, and glutathione peroxidase (GSHPx) activity were significantly lower in the neutrophils of RA patients. However, MTT, GSH and GSHPx values were detected to be significantly increased with INF and MTX therapies. The Ca2+ concentrations were further decreased by the ACA therapy. CONCLUSIONS: Our results suggest that INF and MTX are useful antagonists in apoptosis and mitochondrial oxidative stress in the neutrophils of RA patients. INF and MTX decreased the Ca2+ concentration through inhibition of the TRPM2 channel in the neutrophils of RA patients. It may be a new pathway in the mechanisms of anti-rheumatic drugs.

Albumin evokes Ca2+-induced cell oxidative stress and apoptosis through TRPM2 channel in renal collecting duct cells reduced by curcumin.

In proteinuric nephropathies of chronic kidney disease, the epithelial cells of the nephron including the collecting duct are exposed to high concentrations of luminal albumin. Albumin is taken up from collecting duct cells by endocytosis causing excessive reactive oxygen species (ROS) production and a proinflammatory response. Curcumin used in the traditional medicine possesses anti-inflammatory and antioxidant effects. ROS and ADP-ribose (ADPR) activate the cation channel TRPM2. We hypothesize, that albumin-induced cell stress and proinflammatory response are mediated by Ca2+ and can be reduced by curcumin. The cortical collecting duct (CCD) cells mpkCCDc14 exhibit spontaneous and inducible Ca2+ oscillations, which can be blocked by pre-treatment with curcumin. Curcumin accumulates in plasma membrane and intracellular vesicles, where it interferes with TRPM2 and decreases the influx of Ca2+. Albumin reduces cell viability and increases apoptosis, NF-κB activation, and mitochondrial membrane depolarization via Ca2+-dependent signaling, which results in increased ROS production. Albumin-induced cell stress is diminished by the inhibition of TRPM2 after administration of curcumin and ADPR (PARP1) inhibitors. Curcumin did not reduce the Ca2+ elevation induced by thapsigargin in Ca2+-free medium, but it reduced the function of store-operated Ca2+ channels and ATP-evoked Ca2+ response. In conclusion, albumin-induced oxidative stress is mediated by Ca2+-dependent signaling via TRPM2 and leads to cell damage and a proinflammatory response, strengthening the role of CCD cells in the progression of chronic kidney disease.