Involvement of oxidative stress-related apoptosis in chlorpyrifos-induced cytotoxicity and the ameliorating potential of the antioxidant vitamin E in human glioblastoma cells.

Organophosphate pesticides (OPs), which are among the most widely used synthetic chemicals for the control of a wide variety of pests, are however associated with various adverse reactions in animals and humans. Chlorpyrifos, an OP, has been shown to cause various health complications due to ingestion, inhalation, or skin absorption. The mechanisms underlying the adverse effect of chlorpyrifos on neurotoxicity have not been elucidated. Therefore, we aimed to determine the mechanism of chlorpyrifos-induced cytotoxicity and to examine whether the antioxidant vitamin E (VE) ameliorated these cytotoxic effects using DBTRG-05MG, a human glioblastoma cell line. The DBTRG-05MG cells were treated with chlorpyrifos, VE, or chlorpyrifos plus VE and compared with the untreated control cells. Chlorpyrifos induced a significant decrease in cell viability and caused morphological changes in treated cultures. Furthermore, chlorpyrifos led to the increased production of reactive oxygen species (ROS) accompanied by a decrease in the level of reduced glutathione. Additionally, chlorpyrifos induced apoptosis by upregulating the protein levels of Bax and cleaved caspase-9/caspase-3 and by downregulating the protein levels of Bcl-2. Moreover, chlorpyrifos modulated the antioxidant response by increasing the protein levels of Nrf2, HO-1, and NQO1. However, VE reversed the cytotoxicity and oxidative stress induced by chlorpyrifos treatment in DBTRG-05MG cells. Overall, these findings suggest that chlorpyrifos causes cytotoxicity through oxidative stress, a process that may play an important role in the development of chlorpyrifos-associated glioblastoma.

High Thioredoxin Domain-Containing Protein 11 Expression Is Associated with Tumour Progression in Glioma.

Glioblastoma (GBM) is the most common primary brain malignancy in adults. Despite multimodal treatment that involves maximal safe resection, concurrent chemoradiotherapy, and tumour treatment for supratentorial lesions, the prognosis remains poor. The current median overall survival is only <2 years, and the 5-year survival is only 7.2%. Thioredoxin domain-containing protein 11 (TXNDC11), also known as EF-hand binding protein 1, was reported as an endoplasmic reticulum stress-induced protein. The present study aimed to elucidate the prognostic role of TXNDC11 in GBM. We evaluated the clinical parameters and TXNDC11 scores in gliomas from hospitals. Additionally, proliferation, invasion, migration assays, apoptosis, and temozolomide (TMZ)-sensitivity assays of GBM cells were conducted to evaluate the effects of short interfering RNA (siRNA) on these processes. In addition, these cells were subjected to Western blotting to detect the expression levels of N-cadherin, E-cadherin, and Cyclin D1. High levels of TXNDC11 protein expression were significantly associated with World Health Organization (WHO) high-grade tumour classification and poor prognosis. Multivariate analysis revealed that in addition to the WHO grade, TXNDC11 protein expression was also an independent prognostic factor of glioma. In addition, TXNDC11 silencing inhibited proliferation, migration, and invasion and led to apoptosis of GBM cells. However, over-expression of TXNDC11 enhanced proliferation, migration, and invasion. Further, TXNDC11 knockdown downregulated N-cadherin and cyclin D1 expression and upregulated E-cadherin expression in GBM cells. Knock-in TXNDC11 return these. Finally, in vivo, orthotopic xenotransplantation of TXNDC11-silenced GBM cells into nude rats promoted slower tumour growth and prolonged survival time. TXNDC11 is a potential oncogene in GBMs and may be an emerging therapeutic target.

Investigation of cytotoxic effect of the bufanolide steroid compound cinobufagin and its related underlying mechanism in brain cell models.

Cinobufagin, a bufadienolide of toad venom of Bufo bufo gargarizans, is used as a cardiotonic, central nervous system (CNS) respiratory agent, as well as an analgesic and anesthetic. However, several research showed that bufadienolide has a few side effects on the CNS, such as breathlessness or coma. Although cinobufagin was shown to display pharmacological effects in various models, the toxic effect of cinobufagin is elusive in brain cell models. The aim of this study was to explore whether cinobufagin affected viability, Ca2+ homeostasis, and reactive oxygen species (ROS) production in Gibco® Human Astrocyte (GHA) and HCN-2 neuronal cell line. In GHA cells but not in HCN-2 cells, cinobufagin (20-60 µM) induced [Ca2+ ]i rises. In terms of cell viability, chelation of cytosolic Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N'N'-tetraacetic acid reduced cinobufagin-induced cytotoxicity in GHA cells. In GHA cells, cinobufagin-induced Ca2+ entry was inhibited by 2-aminoethoxydiphenyl borate or SKF96365. In a Ca2+ -free medium, treatment with thapsigargin or U73122 abolished cinobufagin-evoked [Ca2+ ]i rises. Furthermore, treatment with N-acetylcysteine reversed ROS production and cytotoxicity in cinobufagin-treated GHA cells. Together, in GHA cells but not in HCN-2 cells cinobufagin caused cytotoxicity that was linked to preceding [Ca2+ ]i rises by Ca2+ influx via store-operated Ca2+ entry and phospholipase C-dependent Ca2+ release from the endoplasmic reticulum. Moreover, cinobufagin induced ROS-associated cytotoxicity.

Mechanism of action of a diterpene alkaloid hypaconitine on cytotoxicity and inhibitory effect of BAPTA-AM in HCN-2 neuronal cells.

Hypaconitine, a neuromuscular blocker, is a diterpene alkaloid found in the root of Aconitum carmichaelii. Although hypaconitine was shown to affect various physiological responses in neurological models, the effect of hypaconitine on cell viability and the mechanism of its action of Ca2+ handling is elusive in cortical neurons. This study examined whether hypaconitine altered viability and Ca2+ signalling in HCN-2 neuronal cell lines. Cell viability was measured by the cell proliferation reagent (WST-1). Cytosolic Ca2+ concentrations [Ca2+ ]i was measured by the Ca2+ -sensitive fluorescent dye fura-2. In HCN-2 cells, hypaconitine (10-50 µmol/L) induced cytotoxicity and [Ca2+ ]i rises in a concentration-dependent manner. Removal of extracellular Ca2+ partially reduced the hypaconitine's effect on [Ca2+ ]i rises. Furthermore, chelation of cytosolic Ca2+ with BAPTA-AM reduced hypaconitine's cytotoxicity. In Ca2+ -containing medium, hypaconitine-induced Ca2+ entry was inhibited by modulators (2-APB and SKF96365) of store-operated Ca2+ channels and a protein kinase C (PKC) inhibitor (GF109203X). Hypaconitine induced Mn2+ influx indirectly suggesting that hypaconitine evoked Ca2+ entry. In Ca2+ -free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished hypaconitine-induced [Ca2+ ]i rises. Conversely, treatment with hypaconitine inhibited thapsigargin-induced [Ca2+ ]i rises. However, inhibition of phospholipase C (PLC) with U73122 did not inhibit hypaconitine-induced [Ca2+ ]i rises. Together, hypaconitine caused cytotoxicity that was linked to preceding [Ca2+ ]i rises by Ca2+ influx via store-operated Ca2+ entry involved PKC regulation and evoking PLC-independent Ca2+ release from the endoplasmic reticulum. Because BAPTA-AM loading only partially reversed hypaconitine-induced cell death, it suggests that hypaconitine induced a second Ca2+ -independent cytotoxicity in HCN-2 cells.

Inhibition of the pesticide rotenone-induced Ca2+ signaling, cytotoxicity and oxidative stress in HCN-2 neuronal cells by the phenolic compound hydroxytyrosol.

Rotenone, a plant-derived pesticide belonging to genera Derris and Lonchorcarpus, is an inhibitor of NADH dehydrogenase complex. Studies have shown that rotenone was applied as a neurotoxic agent in various neuronal models. Hydroxytyrosol [2-(3,4-dihydroxyphenyl)-ethanol] is a natural phenolic compound found in the olive (Olea europaea L.). Studies of hydroxytyrosol have dramatically increased because this compound may contribute to the prevention of neurodegenerative diseases. Although hydroxytyrosol has received increasing attention due to its multiple pharmacological activities, it is not explored whether hydroxytyrosol inhibited rotenone-induced cytotoxicity in the neuronal cell model. The aim of this study was to explore whether hydroxytyrosol prevented rotenone-induced Ca2+ signaling, cytotoxicity and oxidative stress in HCN-2 neuronal cell line. In HCN-2 cells, rotenone (5-30 µM) concentration-dependently induced cytosolic Ca2+ concentrations ([Ca2+]i) rises and cytotoxicity. Treatment with hydroxytyrosol (30 µM) reversed rotenone (20 µM)-induced cytotoxic responses. In Ca2+-containing medium, rotenone-induced Ca2+ entry was inhibited by 2-APB (a store-operated Ca2+ channel modulator) or hydroxytyrosol. In Ca2+-free medium, treatment with thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) or hydroxytyrosol significantly inhibited rotenone-induced [Ca2+]i rises. Furthermore, treatment with hydroxytyrosol reversed ROS levels, cytotoxic responses, and antioxidant enzyme activities (SOD, GPX and CAT) in rotenone-treated cells. Together, in HCN-2 cells, rotenone induced Ca2+ influx via store-operated Ca2+ entry and Ca2+ release from the endoplasmic reticulum and caused oxidative stress. Moreover, hydroxytyrosol ameliorated Ca2+ or ROS-associated cytotoxicity. It suggests that hydroxytyrosol might have a protective effect on rotenone-induced neurotoxicity in human neuronal cells.

Mechanisms underlying protective effects of vitamin E against mycotoxin deoxynivalenol-induced oxidative stress and its related cytotoxicity in primary human brain endothelial cells.

Fusarium mycotoxins are one of the largest families of mycotoxins. Among these mycotoxins, deoxynivalenol is the most widespread pollutant of grains. However, the mechanism underlying the effect of deoxynivalenol on cytotoxicity in human brain endothelial cells was still unclear. This study examined whether deoxynivalenol induced oxidative stress-associated cytotoxicity in primary human brain endothelial cells (HBEC-5i), and explored whether Vitamin E (VE), a selective antioxidant, had protective effects on deoxynivalenol-treated cells. Deoxynivalenol (10-50 µM) concentration-dependently induced cytotoxicity in HBEC-5i cells. Deoxynivalenol (IC50 = 20 µM) activated mitochondrial apoptotic pathway by modulating antioxidant protein expressions (Nrf2, HO-1 and NQO1). More significantly, pre-treatment with VE (20 µM) attenuated the deoxynivalenol-induced cytotoxicity in this cell model. Together, VE significantly alleviated the apoptotic effects of deoxynivalenol in HBEC-5i cells suggesting that it protected the cells against deoxynivalenol-induced oxidative damage. Our findings provided new insight that VE had the potential to ameliorate neurotoxicity of deoxynivalenol.

Uncovering malathion (an organophosphate insecticide) action on Ca2+ signal transduction and investigating the effects of BAPTA-AM (a cell-permeant Ca2+ chelator) on protective responses in glial cells.

Malathion, one of commonly used organophosphate insecticides, has a wide range of toxic actions in different models. However, the effect of this compound on Ca2+ homeostasis and its related cytotoxicity in glial cells is elusive. This study examined whether malathion evoked intracellular Ca2+ concentration ([Ca2+]i) rises and established the relationship between Ca2+ signaling and cytotoxicity in normal human astrocytes, rat astrocytes and human glioblastoma cells. The data show that malathion induced concentration-dependent [Ca2+]i rises in Gibco® Human Astrocytes (GHA cells), but not in DI TNC1 normal rat astrocytes and DBTRG-05MG human glioblastoma cells. In GHA cells, this Ca2+ signal response was reduced by removing extracellular Ca2+. In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished malathion-induced [Ca2+]i rises. Conversely, incubation with malathion abolished thapsigargin-induced [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 also blocked malathion-induced [Ca2+]i rises. In Ca2+-containing medium, malathion-induced [Ca2+]i rises was inhibited by store-operated Ca2+ channel blockers (2-APB, econazole or SKF96365) and the protein kinase C (PKC) inhibitor GF109203X. Malathion (5-25 µM) concentration-dependently caused cytotoxicity in GHA, DI TNC1 and DBTRG-05MG cells. This cytotoxic effect was partially prevented by prechelating cytosolic Ca2+ with BAPTA-AM (a selective Ca2+ chelator) only in GHA cells. Together, in GHA but not in DI TNC1 and DBTRG-05MG cells, malathion induced [Ca2+]i rises by inducing PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ channels. Furthermore, malathion induced Ca2+-associated cytotoxicity, suggesting that Ca2+ chelating may have a protective effect on malathion-induced cytotoxicity in normal human astrocytes.

Effect of tetramethylpyrazine (TMP) on Ca2+ signal transduction and cell viability in a model of renal tubular cells.

Tetramethylpyrazine (TMP) is a compound purified from herb. Its effect on Ca2+ concentrations ([Ca2+ ]i ) in renal cells is unclear. This study examined whether TMP altered Ca2+ signaling in Madin-Darby canine kidney (MDCK) cells. TMP at 100-800 µM induced [Ca2+ ]i rises, which were reduced by Ca2+ removal. TMP induced Mn2+ influx implicating Ca2+ entry. TMP-induced Ca2+ entry was inhibited by 30% by modulators of protein kinase C (PKC) and store-operated Ca2+ channels. Treatment with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) inhibited 93% of TMP-evoked [Ca2+ ]i rises. Treatment with TMP abolished BHQ-evoked [Ca2+ ]i rises. Inhibition of phospholipase C (PLC) abolished TMP-induced responses. TMP at 200-1000 µM decreased viability, which was not reversed by pretreatment with the Ca2+  chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester. Together, in MDCK cells, TMP induced [Ca2+ ]i rises by evoking PLC-dependent Ca2+ release from endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ entry. TMP also caused Ca2+ -independent cell death.

Evaluation of cytotoxicity of propofol and its related mechanism in glioblastoma cells and astrocytes.

Propofol (2,6-diisopropylphenol), one of the extensively and commonly used anesthetic agents, has been shown to affect the biological behavior of various models. Previous researches have shown that propofol-induced cytotoxicity might cause anticancer effect in different cells. However, the mechanisms underlying the effect of propofol on cytotoxicity is still elusive in human glioblastoma cells. The aims of this study were to evaluate effects of propofol on cytotoxicity, cell cycle distribution and ROS production, and establish the relationship between oxidative stress and cytotoxicity in GBM 8401 human glioblastoma cells and DI TNC1 rat astrocytes. Propofol (20-30 µM) concentration-dependently induced cytotoxicity, cell cycle arrest, and increased ROS production in GBM 8401 cells but not in DI TNC1 cells. In GBM 8401 cells, propofol induced G2/M phase cell arrest, which affected the CDK1, cyclin B1, p53, and p21 protein expression levels. Furthermore, propofol induced oxygen stresses by increasing O2- and H2 O2 levels but treatment with the antioxidant N-acetylcysteine (NAC) partially reversed propofol-regulated antioxidative enzyme levels (superoxide dismutase, catalase, and glutathione peroxidase). Most significantly, propofol induced apoptotic effects by decreasing Bcl-2 but increasing Bax, cleaved caspase-9/caspase-3 levels, which were partially reversed by NAC. Moreover, the pancaspase inhibitor Z-VAD-FMK also partially prevented propofol-induced apoptosis. Together, in GBM 8401 cells but not in DI TNC1 cells, propofol activated ROS-associated apoptosis that involved cell cycle arrest and caspase activation. These findings indicate that propofol not only can be an anesthetic agent which reduces pain but also has the potential to be used for the treatment of human glioblastoma.

Effect of Methoxsalen on Ca²âº Homeostasis and Viability in Human Osteosarcoma Cells.

Methoxsalen is a natural compound found in many seed plants. The effect of methoxsalen on Ca²âº- related physiology in human osteosarcoma is unclear. This study investigated the effect of methoxsalen on cytosolic free Ca²âº concentrations ([Ca²âº]i) in MG63 human osteosarcoma cells. Methoxsalen induced [Ca²âº]i rises concentration-dependently. Methoxsalen-induced Ca²âº entry was confirmed by Mn²âº-induced quench of fura-2 fluorescence. This Ca²âº entry was suppressed by nifedipine, econazole, and SKF96365. In Ca²âº-free medium, incubation with the endoplasmic reticulum Ca²âº pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) inhibited methoxsalen-evoked [Ca²âº]i rises by 96%. In contrast, incubation with methoxsalen abolished BHQ-evoked [Ca²âº]i rises. Inhibition of phospholipase C (PLC) with U73122 abolished methoxsalen-induced [Ca²âº]i rises. Methoxsalen was cytotoxic at 300-700 µM in a concentration-dependent fashion. Chelating cytosolic Ca²âº with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxymethyl ester (BAPTA/AM) did not prevent methoxsalen-induced cytotoxicity. Collectively, our data suggest that in MG63 cells, methoxsalen induced [Ca²âº]i rises by evoking PLC-dependent Ca²âº release from the endoplasmic reticulum, and Ca²âº entry via store-operated Ca²âº entry. Methoxsalen also induced Ca²âº- disassociated cell death.

Discriminating pyogenic brain abscesses, necrotic glioblastomas, and necrotic metastatic brain tumors by means of susceptibility-weighted imaging.

OBJECTIVES: To investigate the feasibility of using susceptibility-weighted imaging (SWI) to discriminate abscesses and necrotic tumours. METHODS: Twenty-one patients with pyogenic abscesses, 21 patients with rim-enhancing glioblastomas and 23 patients with rim-enhancing metastases underwent SWI. Intralesional susceptibility signal (ILSS) was analyzed employing both qualitative (QL) and semi-quantitative (SQ) methods. Logistic regression models and receiver operating characteristic analysis were used to demonstrate the discriminating power. RESULTS: In QL analysis, ILSSs were seen in 12 of 21 abscesses, in 20 of 21 glioblastomas, and in 16 of 23 metastases. In SQ analysis, a low degree of ILSS (85.8 %) was in the majority of abscesses and a high degree of ILSS (76.2 %) was in the majority of glioblastomas. SQ model was significantly better than QL model in distinguishing abscesses from glioblastomas (P < .001). A derived ILSS cutoff grade of 1 or less was quantified as having a sensitivity of 85.7 %, specificity of 90.5 %, accuracy of 88.1 %, PPV of 90.0 %, and NPV of 86.4 % in distinguishing abscesses from glioblastomas. CONCLUSIONS: A high-grade ILSS may help distinguish glioblastomas from abscesses and necrotic metastatic brain tumours. The lack of ILSS or low-grade ILSS can be a more specific sign in the imaging diagnosis of abscesses. KEY POINTS: • ILSS of SWI can contribute to differential diagnosis of rim-enhanced mass. • Low-grade ILSS can be a more specific sign in abscesses. • High-grade ILSS may help distinguish necrotic glioblastomas from abscesses. • ILSS spreads across the four ILSS categories in metastases.

Rise of [Ca²âº]i and apoptosis induced by M-3M3FBS in SCM1 human gastric cancer cells.

M-3M3FBS (2,4,6-trimethyl-N-(meta-3-trifluoromethyl-phenyl)-benzenesulfonamide is a presumed phospholipase C activator which induced Ca²âº movement and apoptosis in different cell models. How- ever, the effect of m-3M3FBS on cytosolic free Ca²âº concentrations ([Ca²âº]i) and apoptosis in SCM1 human gastric cancer cells is unclear. This study explored whether m-3M3FBS elevated basal [Ca²âº]i levels in suspended cells by using fura-2 as a Ca²âº-sensitive fluorescent dye. M-3M3FBS at concentrations between 5-50 µM increased [Ca²âº]i in a concentration-dependent manner. The Ca²âº signal was reduced by half by removing extracellular Ca²âº. M-3M3FBS-induced Ca²âº influx was inhibited by nifedipine, econazole, SK&F96365, aristolochic acid, and GF109203X. In Ca²âº-free medium, 50 µM m-3M3FBS pretreatment inhibited the [Ca²âº]i rise induced by the endoplasmic reticulum Ca²âº pump inhibitor thapsigargin. Conversely, pretreatment with thapsigargin partly reduced m-3M3FBS-induced [Ca²âº]i rise. Suppression of inositol 1,4,5-trisphosphate production with U73122 did not change m-3M3FBS- induced [Ca²âº]i rise. At concentrations between 25 and 50 µM m-3M3FBS killed cells in a concentration- dependent manner. The cytotoxic effect of m-3M3FBS was not reversed by prechelating cytosolic Ca²âº with acetoxy-methyl ester of bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA/AM). Annexin V/propidium iodide staining data suggest that m-3M3FBS induced apoptosis at 25 and 50 µM. M-3M3FBS also increased levels of superoxide. Together, in human gastric cancer cells, m-3M3FBS induced a [Ca²âº]i rise by inducing phospholipase C-independent Ca²âº release from the endoplasmic reticulum and Ca²âº entry via protein kinase C-sensitive store-operated Ca²âº channels. M-3M3FBS induced cell death that might involve apoptosis via reactive oxygen species production.

M-3M3FBS-induced Ca² ⁺ movement and apoptosis in HA59T human hepatoma cells.

The effect of 2,4,6-trimethyl-N-(meta-3-trifluoromethyl-phenyl)-benzenesulfonamide (m-3M3FBS), a presumed phospholipase C activator, on cytosolic free Ca² ⁺ concentrations ([Ca² ⁺ ]i ) in HA59T human hepatoma cells is unclear. This study explored whether m-3M3FBS elevated basal [Ca² ⁺ ]i levels in suspended cells by using fura-2 as a Ca² ⁺ -sensitive fluorescent dye. M-3M3FBS at concentrations of 10- 50 µM increased [Ca² ⁺ ]i in a concentration-dependent fashion. The Ca² ⁺ signal was reduced partly by removing extracellular Ca² ⁺ . M-3M3FBS-induced Ca² ⁺ influx was inhibited by nifedipine, econazole, SK&F96365, aristolochic acid, and GF109203X. In Ca² ⁺ -free medium, 50 µM m-3M3FBS pretreatment inhibited the [Ca² ⁺ ]i rise induced by the endoplasmic reticulum Ca² ⁺ pump inhibitor thapsigargin. Conversely, pretreatment with thapsigargin partly reduced m-3M3FBS-induced [Ca² ⁺ ]i rise. Inhibition of inositol 1,4,5-trisphosphate formation with U73122 did not alter m-3M3FBS-induced [Ca² ⁺ ]i rise. At concentrations between 10 and 40 µM m-3M3FBS killed cells in a concentration-dependent manner. The cytotoxic effect of m-3M3FBS was not reversed by prechelating cytosolic Ca² ⁺ with 1,2-bis(2- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). Annexin V/propidium iodide staining data suggest that m-3M3FBS induced apoptosis in a concentration-dependent manner. M-3M3FBS also increased levels of reactive oxygen species. Together, in human hepatoma cells, m-3M3FBS induced a [Ca² ⁺ ]i rise by inducing phospholipase C-independent Ca² ⁺ release from the endoplasmic reticulum and Ca² ⁺ entry via protein kinase C-sensitive store-operated Ca² ⁺ channels. M-3M3FBS induced cell death that might involve apoptosis via mitochondrial pathways.

Evaluation of the mycotoxin patulin on cytotoxicity and oxidative stress in human glioblastoma cells and investigation of protective effect of the antioxidant N-acetylcysteine (NAC).

Mycotoxins are secondary metabolites produced by various kinds of fungi that can induce disease in humans. The fungal species Penicillium expansum produces patulin (C7H6O4), a polyketide lactone mycotoxin found in fruits. Patulin is classified as noncarcinogen; however, recently, it has been associated with harmful effects on the central nervous system. Patulin's toxic action has been established in various brain models; however, its effect on human glioblastoma remains elusive. This study explores whether patulin induces cytotoxicity through oxidative stress in DBTRG-05MG human glioblastoma cells. This study also evaluates whether the antioxidant N-acetylcysteine (NAC) protects against patulin-induced cytotoxicity. In DBTRG-05MG cells, patulin concentration (10-60 µM) dependently induced cytotoxicity. Concerning oxidative stress, patulin (10 and 20 µM) increased the production of intracellular reactive oxygen species (ROS) but depleted reduced glutathione (GSH) contents and regulated the expressions of antioxidant-related proteins (Nrf2 and HO-1). Furthermore, patulin induced cytotoxicity via modulation of apoptosis-related protein expressions (Bax, cleaved caspase-9, and cleaved caspase-3). These cytotoxic responses were partially reversed via pretreatment with NAC (10 µM). In summary, these data help us understand the toxicology of patulin in human glioblastoma and evaluate whether NAC could clinically reduce patulin-affected brain damage.

Effect of diindolylmethane on Ca2+ homeostasis and viability in PC3 human prostate cancer cells.

The effect of the natural product diindolylmethane on cytosolic Ca(2+) concentrations ([Ca(2+)](i)) and viability in PC3 human prostate cancer cells was explored. The Ca(2+)-sensitive fluorescent dye fura-2 was applied to measure [Ca(2+)](i). Diindolylmethane at concentrations of 20-50 µM induced [Ca(2+)](i) rise in a concentration-dependent manner. The response was reduced partly by removing Ca(2+). Diindolylmethane-evoked Ca(2+) entry was suppressed by nifedipine, econazole, SK&F96365, protein kinase C modulators and aristolochic acid. In the absence of extracellular Ca(2+), incubation with the endoplasmic reticulum Ca(2+) pump inhibitor thapsigargin or 2,5-di-tert-butylhydroquinone (BHQ) inhibited or abolished diindolylmethane-induced [Ca(2+)](i) rise. Incubation with diindolylmethane also inhibited thapsigargin or BHQ-induced [Ca(2+)](i) rise. Inhibition of phospholipase C with U73122 reduced diindolylmethane-induced [Ca(2+)](i) rise. At concentrations of 50-100 µM, diindolylmethane killed cells in a concentration-dependent manner. This cytotoxic effect was not altered by chelating cytosolic Ca(2+) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). Annexin V/PI staining data implicate that diindolylmethane (50 and 100 µM) induced apoptosis in a concentration-dependent manner. In conclusion, diindolylmethane induced a [Ca(2+)](i) rise in PC3 cells by evoking phospholipase C-dependent Ca(2+) release from the endoplasmic reticulum and Ca(2+) entry via phospholipase A(2)-sensitive store-operated Ca(2+) channels. Diindolylmethane caused cell death in which apoptosis may participate.

Tumorigenesis and prognostic role of hepatoma-derived growth factor in human gliomas.

Hepatoma-derived growth factor (HDGF) is a neurotrophic factor found in mouse spinal cord and hippocampal neurons. In various malignant tumors, the role of HDGF in tumor progression and its use as a diagnostic biomarker or therapeutic target have been extensively explored. However, the prognostic function and mitogenic role of HDGF in gliomagenesis are yet to be verified. In this study, we found a significant incidence of HDGF prevalence between the different pathological types and stages of glioma in 105 patients. We also found a prognostic significance in 41 glioblastoma multiforme (GBM) patients, with prevalence of nuclear HDGF predicting short survival of patients with GBM after surgery. To delineate the mitogenic role of HDGF in gliomagenesis, an adenoviral-expressed HDGF small interfering RNA (Ad-HDGF siRNA) was used to knock down expression of nuclear HDGF. After knocking down nuclear HDGF expression in human GBM cells, cell growth and cell invasion and induction on apoptosis by caspase-3 activation were significantly inhibited. We conclude that HDGF is a mitogenic growth factor in glioma progression and can be a useful prognostic marker for GBM and therapeutic target for clinical management of glioma in the future.

Effect of celecoxib on Ca(2+) handling and viability in human prostate cancer cells (PC3).

Celecoxib has been shown to have an antitumor effect in previous studies, but the mechanisms are unclear. Ca(2+) is a key second messenger in most cells. The effect of celecoxib on cytosolic free Ca(2+) concentrations ([Ca(2+)](i)) in human suspended PC3 prostate cancer cells was explored by using fura-2 as a fluorescent dye. Celecoxib at concentrations between 5 and 30 µM increased [Ca(2+)](i) in a concentration-dependent manner. The Ca(2+) signal was reduced partly by removing extracellular Ca(2+). Celecoxib-induced Ca(2+) influx was not blocked by L-type Ca(2+) entry inhibitors or protein kinase C/A modulators [phorbol 12-myristate 13-acetate (PMA), GF109203X, H-89], but was inhibited by the phospholipase A(2) inhibitor, aristolochic acid. In Ca(2+)-free medium, 30 µM of celecoxib failed to induce a [Ca(2+)](i) rise after pretreatment with thapsigargin (an endoplasmic reticulum [ER] Ca(2+) pump inhibitor). Conversely, pretreatment with celecoxib inhibited thapsigargin-induced Ca(2+) release. Inhibition of phospholipase C with U73122 did not change celecoxib-induced [Ca(2+)](i) rises. Celecoxib induced slight cell death in a concentration-dependent manner, which was enhanced by chelating cytosolic Ca(2+) with BAPTA. Collectively, in PC3 cells, celecoxib induced [Ca(2+)](i) rises by causing phospholipase C-independent Ca(2+) release from the ER and Ca(2+) influx via non-L-type, phospholipase A(2)-regulated Ca(2+) channels. These data may contribute to the understanding of the effect of celecoxib on prostate cancer cells.

Deltamethrin-Induced [Ca ²âº ]i Rise and Death in HGB Human Glioblastoma Cells.

The effect of the pesticide deltamethrin on cytosolic free Ca ²âº concentrations ([Ca²âº ]i) and viability in human glioblastoma DBTRG-05MG cells is explored. [Ca²âº ]i was measured in suspended cells using fura-2 as a Ca²âº -sensitive fluorescent dye. Deltamethrin at concentrations of 5-60 µM increased [Ca²âº]i in a concentration-dependent fashion. The Ca²âº signal was reduced partly by removing extracellular Ca ²âº . Deltamethrin induced Mn ²âº entry leading to quenching of fura-2 fluorescence. Deltamethrin-induced [Ca²âº ]i rise was not inhibited by nifedipine, econazole, SKF96365, and phorbol 12-myristate 13 acetate, but was inhibited by the protein kinase C (PKC) inhibitor GF109203X via blocking Ca²âº release. In Ca²âº -free medium, 50 µM deltamethrin pretreatment abolished the [Ca²âº]i rise induced by the endoplasmic reticulum Ca²âº pump inhibitor thapsigargin (TG) or 2,5-di-tertbutylhydroquinone (BHQ). Conversely, pretreatment with TG/BHQ abolished deltamethrin-induced [Ca²âº]i rise. Inhibition of inositol 1,4,5-trisphosphate formation with U73122 suppressed 50% of deltamethrin-induced [Ca²âº]i rise. At concentrations between 10 and 80 µM deltamethrin killed cells in a concentration-dependent manner. The cytotoxic effect of deltamethrin was not reversed by prechelating cytosolic Ca²âº with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). Annexin V/ propidium iodide staining data suggest that deltamethrin (10-40 µM) induced apoptosis in a concentrationdependent manner. Deltamethrin also increased levels of reactive oxygen species. Together, in human glioblastoma cells, deltamethrin induced a [Ca²âº]i rise by inducing phospholipase C- and PKC-dependent Ca²âº release from the endoplasmic reticulum and Ca²âº entry via non-store-operated Ca²âº channels. Deltamethrin induced cell death that might involve apoptosis via mitochondrial pathways.

Intra-tumoral susceptibility signal: a post-processing technique for objective grading of astrocytoma with susceptibility-weighted imaging.

BACKGROUND: Susceptibility-weighted imaging (SWI) is sensitive to the accumulation of paramagnetic substances, such as hemorrhage and increased venous vasculature, both being frequently found in high-grade tumors. The purpose of this retrospective study is to differentiate high-grade and low-grade astrocytoma by objectively measuring quantitative intra-tumoral susceptibility signals (qITSS) on SWI. METHODS: Precontrast SWI and 3D contrast-enhanced T1WI of 65 patients with astrocytoma were collected at 1.5 Tesla. All tumors were histologically confirmed and classified into two groups: high grade (WHO grade III and IV, n=50) and low grade (WHO grade II, n=15). After manual delineation of the tumor on T1WI, normalized contrast (NC) was calculated voxel by voxel within the tumor by using the concept of contrast to noise ratio. Thresholding on NC was applied to detect qITSS, and the volumetric percentage of qITSS can be obtained for each tumor. Two-sample t-test was applied to examine significant difference of qITSS percentage between high-grade and low-grade astrocytoma for different NC thresholds, ranging from 4 to 20. Receiver operating characteristic analysis was performed to evaluate the performance of differentiation. RESULTS: P value was less than 0.01 for a large range of NC thresholds [4-20], reflecting significant difference of qITSS percentage between high-grade and low-grade astrocytoma. The area under the receiver operating characteristic curve was larger than 0.9 at NC thresholds from 8 to 16 and peaks at 0.949 with a NC threshold of 14. It was shown that astrocytoma grading by qITSS percentage is successful for a wide range of NC threshold, demonstrating robustness on threshold selection. CONCLUSIONS: Without relying on the selection of slice position and at the same time providing objective identification of hypointense signal in SWI, the qITSS percentage can be used to distinguish high-grade and low-grade astrocytoma reliably.

Adult Posterior Fossa Anaplastic Ependymoma, Case Series and Literature Review.

OBJECTIVE: Ependymomas are rare central nervous system tumors. The current treatment strategy is gross total tumor removal. Whether adjuvant therapy will be beneficial is controversial. We retrospectively analyzed 3 cases of World Health Organization (WHO) grade III posterior fossa anaplastic ependymomas treated with different treatment modalities. We aimed to identify possible treatment options for infratentorial WHO grade III anaplastic ependymoma in adults. METHODS: We performed a retrospective analysis of 3 patients diagnosed with infratentorial anaplastic ependymomas in our institution from 2016 to 2020. The demographic data were documented. This case series of 3 patients does not meet the Department of Health and Human Services definition of research and does not need Institutional Review Board approval. All patients' informed consents have been obtained. RESULTS: One patient underwent subtotal tumor resection combined with adjuvant radiotherapy and Gamma Knife radiosurgery while the other 2 patients underwent gross total tumor removal combined with Gamma Knife radiosurgery or adjuvant radiotherapy. Tumors recurred in the first patient 20 months later, while the other 2 patents did not develop recurrence. The modified Rankin scale scores of these patients were 1, 0, and 0. All patients are followed up with regular magnetic resonance imaging at our facility. CONCLUSIONS: The strategy for treating WHO grade III anaplastic ependymomas is controversial, but gross total tumor resection remains the key element. Adjuvant stereotactic radiosurgery after tumor removal might be considered if radiotherapy is not an option. The role of chemotherapy is unclear, and the use of chemotherapy should be tailored to individual patients.