OBJECTIVETo investigate the effect of deltamethrin (DM) on production of reactive oxygen species (ROS) of rat pheochromocytoma (PC12) cells and its mechanism.

METHODSPC12 cells were treated with various dose of DM (0, 10 or 100 micromol/L) for 1, 6 or 12 h respectively. Furthermore, PC12 cells were treated with various dose of DM (0, 10 or 100 micromol/L) for 24 or 48 h, respectively. PC12 cells were pre-incubated with 10 mmol/L N-acetyl-L-cysteine (NAC) for 2 h, or with 500 micromol/L DL-Buthionine-[S, R]-Sulfoximine (BSO) for 16 h, or with 40 micromol/L tertiary butylhydroquinone (tBHQ) for 16 h, prior to exposure to DM and then with 10 micromol/L DM for 6 h. After treatment, ROS production in PC12 cells were measured by a molecular probe, 2', 7'-dichlorofluorescein diacetate (DCFH-DA).

RESULTSDM induced a dose-time dependent increase in ROS production (indicated by DCF fluorescence intensity). 10 micromol/L DM treatment for 6 h enhanced DCF fluorescence intensity that reached approximately 2.24 times of values of control group. Furthermore, a pretreatment with NAC, BSO or tBHQ significantly reduced the DM-enhanced DCF fluorescence intensity that reached approximately 22%, 62% or 38% of values of DM treatment, respectively (P < 0.05), indicating that all these pretreatments attenuate ROS production.

CONCLUSIONThe in vitro studies demonstrate that DM could enhance ROS production, and may be the influential factor for the decreased mercapto level and antioxidative function.

Animals ; Nitriles ; pharmacology ; PC12 Cells ; Pyrethrins ; pharmacology ; Rats ; Reactive Oxygen Species ; metabolism

OBJECTIVETo establish a human breast cancer MCF-7 cell model stably overexpressing the aromatase gene (MCF-7-aromatase) and aromatase inhibitor letrozole-resistant MCF-7 cell model (MCF-7-LR).

METHODSWe utilized the lentivirus-mediated gene transfer approach to establish MCF-7-aromatase cell and MCF-7 cell model stably overexpressing green fluorescent protein (GFP) (MCF-7-GFP). The expression of aromatase in the MCF-7-aromatase and MCF-7-GFP cells was determined by reverse transcription polymerase chain reaction (RT-PCR), real time quantitative PCR (RT-qPCR), Western blot and immunoprecipitation (IP) assay. The proliferative ability in vitro of MCF-7-aromatase and MCF-7-GFP cells treated with testostorone and β-estradiol (E2) was determined by WST-1 cell proliferation assay. The proliferative ability of MCF-7-aromatase cells treated with letrozole was determined by WST-1 assay. The half maximal inhibitory concentration (IC50 value) for letrozole was calculated from the nonlinear regression line of the plot of cell viability (percentage of control) versus letrozole concentration using Graphpad Prism software. MCF-7-aromatase cells were continuously cultured in the presence of testosterone and letrozole, thus letrozole-resistant MCF-7-LR cells were obtained. WST-1 assay was performed to determine their chemoresistance to letrozole.

RESULTSRT-PCR and RT-qPCR results revealed that the mRNA expression of aromatase was significantly increased in the MCF-7-aromatase cells compared with that in the MCF-7-GFP cells. Both Western blot and IP assays showed that the expression of aromatase protein was drastically increased in the MCF-7-aromatase cells, compared with that in the MCF-7-GFP cells. WST-1 assay showed that the cell proliferation rate of MCF-7-aromatase cells treated with 1 and 10 nmol/L testosterone was 1.43- and 1.53-fold higher than that of the control cells, respectively. The proliferation rate of MCF-7-aromatase cells treated with 1 and 10 nmol/L E2 was 1.41- and 1.55-fold higher than that of the control cells, respectively. In contrast, the proliferation rate of MCF-7-GFP cells treated with 10 nmol/L testosterone was 1.12-fold higher than that of the control cells, and the proliferation rate of MCF-7-GFP cells treated with 1 and 10 nmol/L E2 was 1.41- and 1.51-fold higher than that of the control cells, respectively. Letrozole treatment significantly inhibited the testosterone-induced proliferation ability of MCF-7-aromatase cells in a dose-dependent manner and the IC50 value was 5.3 nmol/L. In contrast, letrozole treatment showed no inhibitory effect on the proliferative ability of MCF-7-LR cells and the IC50 value was >1000 nmol/L.

CONCLUSIONSMCF-7-aromatase and MCF-7-LR cells exhibit different response to letrozole treatment, which provides an important basis for further investigating the mechanism of letrozole resistance.

Antineoplastic Agents ; pharmacology ; Aromatase ; metabolism ; Aromatase Inhibitors ; pharmacology ; Breast Neoplasms ; Cell Proliferation ; Drug Resistance, Neoplasm ; Humans ; MCF-7 Cells ; Models, Biological ; Nitriles ; pharmacology ; Triazoles ; pharmacology

OBJECTIVETo assess the joint toxicity of phoxim (Pho) and fenvalerate (Fen) on the spermatogenesis of male rats and to clarify its mechanism.

METHODSBased on the three administrative levels of factorial analysis (3 x 3) of Pho and Fen, i. e. their half lethal dose (LD50) 1/250 LD50 (5.9, 2.4 mg/kg) and 1/50 LD50 (29.4, 12.0 mg/kg) and the control, 135 adult male SD rats were randomly assigned to9 groups, the control (0.0 mg/kg), Pho (5.9, 29.4 mg/kg), Fen (2.4, 12.0 mg/kg), Pho + Fen (5.9 + 2.4, 5.9 + 12.0, 29.4 + 2.4, 29.4 + 12.0 mg/kg), and treated intragastrically with different doses of Pho, Fen and Pho + Fen for 15 and 30 days. The levels of serum follicle stimulating hormone (FSH), luteinizing hormone (LH), testosterone (T) and testis homogenate T were determined by radioimmunoassay (RIA), the activity of testicular marker enzymes such as acid phosphatases (ACP) and gamma-glutamyl transpeptidase (gamma-GT) examined, and the sperm head count measured for the changes of daily sperm production (Spr).

RESULTSAt 15 days, obvious interaction was observed between Pho and Fen in both serum LH and FSH (P < 0.05), as well as between their reproductive toxicities. With the increase in doses, the joint action was synergistic in LH (P < 0.05) and antagonistic in FSH (P < 0.01) at a high dose ( 29.4 + 12.0 mg/kg). At 30 days, marked interaction between Pho and Fen was noted in the content of homogenate T (P < 0.05) , with a joint synergistic effect. At 15 and 30 days, with the increase of doses, both Pho and Fen reduced Spr and the activity of ACP and gamma-GT, but Pho + Fen showed no obvious interaction in them (P > 0.05) , and their joint action was an additive effect.

CONCLUSIONPho and Fen jointly impaired spermatogenesis in a dose- and time-dependent manner. Their joint action exhibited mainly as a synergistic effect, an additive effect and increased toxicity.

Animals ; Drug Interactions ; Male ; Nitriles ; pharmacology ; Organothiophosphorus Compounds ; pharmacology ; Pyrethrins ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Sperm Count ; Sperm Motility ; Spermatogenesis ; drug effects

OBJECTIVETo study the effect of deltamethrin on the apoptotic rate and the expression of caspase-3 in rat neural cells.

METHODSMale Wistar rats were randomly divided into 5 groups: control, 5 h, 24 h, 48 h and 5 d exposed groups. Apoptotic rate and the expression of caspase-3 were measured by FACS420 Flow Cytometer; Ac-DEVD-pNa was used as a substrate to detect the activity of caspase-3.

RESULTSApoptotic rates in 24 h, 48 h and 5 d exposed groups in hippocampus and cerebral cortex [hippocampus: (8.45 +/- 1.02)%, (9.44 +/- 1.14)%, (7.58 +/- 0.75)%; cerebral cortex: (7.90 +/- 0.49)%, (8.01 +/- 0.87)%, (7.97 +/- 0.41)% respectively] were higher than those in the control [hippocampus: (2.97 +/- 0.36)%; cerebral cortex: (3.50 +/- 0.48)%] (P < 0.01); the activity of caspase-3 in 5 h, 24 h and 48 h exposed groups (A(405) nm in hippocampus: 0.389 +/- 0.038, 0.472 +/- 0.041, 0.295 +/- 0.049; A(405) nm in cerebral cortex: 0.321 +/- 0.068, 0.429 +/- 0.077, 0.344 +/- 0.047) and 5 d group of hippocampus (0.246 +/- 0.065) were all higher than those of the control (hippocampus: 0.184 +/- 0.054; cerebral cortex: 0.198 +/- 0.049) (P < 0.05, P < 0.01); the expression of caspase-3 in 5 h, 24 h and 48 h exposed groups increased apparently while 5 d group did not.

CONCLUSIONExposure to high dose of deltamethrin would affect the apoptosis, the activity and expression of caspase-3 in rat neural cells. The increase in caspase-3 activity and expression occurred before the rising of neuronal apoptotic rate may be the upstream event of apoptosis.

Animals ; Apoptosis ; drug effects ; Caspase 3 ; Caspases ; metabolism ; Cerebral Cortex ; enzymology ; pathology ; Hippocampus ; enzymology ; pathology ; Insecticides ; pharmacology ; Male ; Nitriles ; pharmacology ; Pyrethrins ; pharmacology ; Random Allocation ; Rats ; Rats, Wistar

OBJECTIVETo explore the effects of λ-cyhalothrin on hippocampus by interfering with estrogen.

METHODSThe healthy female ICR mice of postnatal 28 days were random divided into 12 groups, 4 of those were sham-operation include control, λ-cyhalothrin (LCT, 3.0 µg/g), Letrozole (Let, 1.0 µg/g), and LCT (3.0 µg/g)+Let (1.0 µg/g); and the last 8 were ovariectomized include OVX, Estradiol (E2, 10.0 µg/g), LCT, Let, E2+LCT, E2+Let, LCT+Let, E2+LCT+Let. 10 mice in every group received drugs by intraperitoneal injection for 2 days. Then half of every group initiate the ethological test (open field test and Morris water maze) 24 h later. The last half animals were sacrificed to made frozen section for immunofluorescent assay of postsynaptic density protein 95 (PSD95).

RESULTSIn ethological test, campared with Sham, OVX can lengthen incubation period in the first grid and to get on the platform (P < 0.05); campared with OVX, OVX+E2 can increase the total numbers of through grid and shorten the incubation period to get on the platform (P < 0.05); campared with OVX+E2, OVX+E2+LCT can reduce the number of grid and standing, lengthen incubation period to the platform (P < 0.05); campared with Sham, Sham+LCT can lengthen incubation period to the platform of Sham mice (P < 0.05), but campared with OVX, OVX+LCT can shoten incubation period in the first grid and to get on the platform in OVX mice (P < 0.05); campared with Sham+Let, Sham+LCT+Let can lengthen incubation period in the first grid, reduce the the number of grid and standing (P < 0.05). In the Immunohistochemical fluorescence experiment we find that, campared with Sham, OVX can reduce the expression of PSD95 in CA1,CA3 and DG (P < 0.05); however campared with OVX, E2 or LCT can both inhibit the effect of OVX (P < 0.05); campared with Sham, Sham+LCT can reduce the expression of PSD95, the same result when OVX+E2+LCT campared with OVX+E2 (P < 0.05); campared with OVX+E2+Let, OVX+E2+LCT+Let can reduce the expression of PSD95 in CA3 (P < 0.05); campared with OVX+Let, OVX+LCT+Let can increase the expression of PSD95 in DG (P < 0.05).

CONCLUSIONSWhen few estrogen exist in the body, LCT can show estrogen-like action to promote hippocampal synaptic development; but when circulating estrogen exist, LCT can inhibit synaptic development by interfering estrogen.

Animals ; Estradiol ; Estrogens ; pharmacology ; Female ; Hippocampus ; drug effects ; Humans ; Mice ; Mice, Inbred ICR ; Nitriles ; pharmacology ; Ovariectomy ; Pyrethrins ; pharmacology ; Random Allocation ; Synapses ; drug effects ; Triazoles

BACKGROUNDIt is believed that estrogen plays pivotal roles in the regulation of follicle/oocyte maturation and oocyte fertilizability. It is also involved in the functional preparation of the fallopian tubes for subsequent gamete interaction, in early embryonic development occurring in the tubal microenvironment, and in the preparation of the uterus for implantation. This study was designed to determine whether estrogen is required for follicular and embryonic development.

METHODSThe biosynthesis of estrogen was blocked by a daily injection of the aromatase inhibitor, Arimidex, at a dose of 100 micro g/d, using 3 - 4 week old C57B6 F1 female mice. Injections were continued for 3 days in experiment 1 (n = 10) and for 5 days in experiment 2 (n = 23). Mice in the control group (n = 27) were given the same amount of saline. Exogenous gonadotrophin [7.5 IU pregnant mare serum gonadotrophin (PMSG)] was administered to induce follicular growth and development on the second day. In experiment 1, we tested estrogen and progesterone levels and examined ovary morphology two days later. In experiment 2, 47 hours after PMSG injection, 5 IU human chorionic gonadotropin (hCG) was given and two female mice were then caged with a male mouse overnight. Two days later, we measured estrogen and progesterone levels. We then removed the embryos, cultured them, and examined embryonic development every 24 hours for 3 days.

RESULTSBefore hCG injection, estrogen levels in mice from the Arimidex group were suppressed by 94%, and progesterone levels were suppressed by 75%. There was no difference between the two groups in mean number of total follicles found per animal (30.4 follicles/animal in the control group and 27 follicles/animal in the Arimidex group). Two days after hCG injection, estrogen levels in the Arimidex group were significantly lower than that in the control group (P < 0.01), while progesterone levels were not significantly lower (P > 0.05). The rate of development of embryos, morulae, blastocysts, and hatching blastocysts was not significantly different between the two groups (P = 0.20, 0.10, 0.44, and 0.38, respectively).

CONCLUSIONSIn the present study, by depriving mice of normal estrogen support, we have been able to rule out the absolute need for rising levels of estrogen for the completion of the follicular maturation process and the development of embryos in vitro.

Animals ; Chorionic Gonadotropin ; pharmacology ; Embryonic and Fetal Development ; Estrogens ; physiology ; Female ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Nitriles ; pharmacology ; Oocytes ; physiology ; Ovarian Follicle ; physiology ; Pregnancy ; Triazoles ; pharmacology

Antiviral Agents/pharmacology* ; COVID-19 ; Coronavirus 3C Proteases ; Humans ; Lactams ; Leucine ; Molecular Docking Simulation ; Molecular Dynamics Simulation ; Nitriles ; Proline ; Protease Inhibitors/pharmacology* ; SARS-CoV-2

OBJECTIVETo determine the role of extracellular signal-regulated kinase (ERK)1/2 during focal cerebral ischemia.

METHODSLeft middle cerebral artery occlusion (MCAO) was undergone after the introduction of a nylon suture to the left internal carotid artery in 70 male adult CD-1 mice. ERK 1/2 phosphorylation was detected using Western blot analysis, and the morphological feature was determined by immunohistochemistry. An ERK pathway inhibitor, 1,4-diamino-2,3-dicyano-1,4-bis[2-amino-phenylthio] butadiene (U0126), was administered intravenously 20 minutes before MCAO, and the neurological deficit levels and the infarct volumes were measured 24 hours after MCAO.

RESULTSPhosphorylated ERK 1/2 (pERK 1/2) activity increased after 30 minutes of MCAO and peaked at 2 hours. The immunohistochemical study displayed a large number of pERK 1/2 positive cells in the ischemic basal ganglion and surrounding cortex. Double-labeled fluorescent staining identified the pERK1/2 positive cells as neurons or astrocytes. In U0126 treated mice which had undergone 24 hours of MCAO, the neurological deficit levels and the infarct volumes were 44.6% and 45.8% respectively, less than those of the control mice.

CONCLUSIONSERK plays an important role in focal cerebral ischemia and inhibition of the ERK pathway can help protect against ischemic brain injury, which may provide a therapeutic approach for cerebral ischemia.

Animals ; Basal Ganglia ; pathology ; Brain Ischemia ; metabolism ; pathology ; physiopathology ; Butadienes ; pharmacology ; Cerebral Cortex ; pathology ; Immunohistochemistry ; Male ; Mice ; Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; physiology ; Nitriles ; pharmacology ; Phosphorylation

OBJECTIVETo investigate the inhibition mechanisms of BAY11-7082 (IkappaB-alpha phosphorylation inhibitor) and Lactacystein (proteosome inhibitor) in CD154-induced NF-kappaB activation.

METHODSWe used recombinant CD154 to stimulate EBV/LMP1 negative Ramos B cell and observed the effects of BAY11-7082 and Lactacystein in CD154-induced NF-kappaB luciferase activation, phosphorylation and degradation of IkappaB-alpha, phosphorylation of p65, and nuclear translocation of NF-kappaB subunits upon CD154 stimulation.

RESULTSBoth BAY11-7082 and Lactacystein abrogated CD154-induced NF-kappaB luciferase activation in Ramos cells. While CD154-induced phosphorylation of p65, phosphorylation and degradation of IkappaB-alpha, and nuclear translocation of p50, p65, and c-Rel were all blocked by BAY11-7082; Lactacystein only inhibited degradation of IkappaB-alpha and p65 nuclear translocation.

CONCLUSIONBAY11-7082 and Lactacystein inhibit CD154-induced NF-kappaB activation through different mechanisms.

Acetylcysteine ; analogs & derivatives ; pharmacology ; Apoptosis ; drug effects ; Burkitt Lymphoma ; pathology ; CD40 Ligand ; pharmacology ; Cysteine Proteinase Inhibitors ; pharmacology ; Enzyme Activation ; drug effects ; Humans ; NF-kappa B ; antagonists & inhibitors ; metabolism ; Nitriles ; pharmacology ; Sulfones ; pharmacology ; Tumor Cells, Cultured

OBJECTIVETo study the inhibitory effect of the dual usage of BEZ235 and U0126, the inhibitor of phosphatidyl inositol-3-kinase/protein kinase B pathway and extracellular regulated proteinkinase/mitogen-activated protein kinase pathway, respectively, on cell proliferation.

METHODSPhosphatase and tensin homolog knockout mouse embryonic fibroblast (PTEN-/-MEF) cell lines were used as the cellular model for malignant tumors. BEZ235, the dual inhibitor of phosphatidyl inositol-3-kinase and mammalian target of rapamycin, and U0126, the inhibitor of mitogen-activated protein kinase were used to treat the cells individually and in a combination manner. The inhibitory effects to cell proliferation were monitored by MTT.

RESULTSBoth BEZ235 and U0126 suppressed PTEN knockout cell proliferation, and their half inhibitory concentrations were 6.257 nmol/L and 22.85 μmol/L, respectively. However, the combination treatment of the two drugs showed antagonistic rather than synergistic effect on cell proliferation.

CONCLUSIONBEZ235 and U0126 are not suitable for a combined target therapy regimen.

Animals ; Butadienes ; pharmacology ; Cell Line ; Cell Proliferation ; drug effects ; Drug Antagonism ; Fibroblasts ; drug effects ; Imidazoles ; pharmacology ; Mice ; Mice, Knockout ; Nitriles ; pharmacology ; Phosphatidylinositol 3-Kinase ; antagonists & inhibitors ; pharmacology ; Quinolines ; pharmacology