Toxicity of Industrially Relevant Chlorinated Organic Solvents In Vitro

2013 ◽  
Vol 32 (2) ◽  
pp. 136-145 ◽  
Author(s):  
Catherine McDermott ◽  
James J.A. Heffron
Keyword(s):  
Cell Proliferation ◽  
Organic Solvents ◽  
Caspase 3 ◽  
Free Calcium ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Chlorinated Organic ◽  
Free Calcium Concentration ◽  
Ros Formation

The cytotoxic effects of 4 industrially important chlorinated organic solvents, dichloromethane (DCM), 1,2-dichloroethane (DCE), trichloroethylene (TCE), and tetrachloroethylene (PERC) in vitro, were investigated. Jurkat T cells were exposed to the solvents individually for 72 hours and changes in reactive oxygen species (ROS) formation, cell proliferation, intracellular free calcium concentration ([Ca2+]), and caspase-3 activity were measured. There was a concentration-dependent increase in the ROS formation and intracellular free [Ca2+] following exposure to each of the solvents. This was accompanied by a decrease in the cell proliferation. Solvent potency decreased in the following order: PERC > TCE > DCM > DCE. Caspase-3 activity was increased in a concentration-dependent manner by TCE and PERC but was not significantly altered by DCM or DCE. n-Acetyl-l-cysteine pretreatment showed that changes in the intracellular free [Ca2+] and caspase-3 activity were independent of ROS formation. However, increased ROS formation did play a causal role in the decreased cell proliferation observed.

Evaluation of the effects of hydrophilic organic solvents on CYP3A-mediated drug-drug interaction in vitro

2006 ◽  
Vol 25 (12) ◽  
pp. 715-721 ◽  
Author(s):  
M Iwase ◽  
N Kurata ◽  
R Ehana ◽  
Y Nishimura ◽  
T Masamoto ◽  
...  
Keyword(s):  
Organic Solvent ◽  
Organic Solvents ◽  
Human Liver ◽  
Liver Microsomes ◽  
Dependent Manner ◽  
Cyp3a4 Activity ◽  
Modulation Effect ◽  
Concentration Dependent Manner ◽  
Hydroxylation Activity

This study evaluated the effects of the commonly used hydrophilic organic solvents, acetonitrile, methanol, ethanol, 1-propanol, dimethyl sulfoxide (DMSO), N,N-dimethylformamide, polyethylene glycol and propylene glycol, on CYP3A in pooled human liver microsomes, using testosterone and midazolam as substrates. Furthermore, we examined the modulation effect of organic solvents on CYP3A inhibition by ketoconazole. Testosterone 6b-hydroxylation activity was potently inhibited in the presence of DMSO and 1-propanol in a concentration-dependent manner. Midazolam 1'-hydroxylation activity, however, was weakly inhibited only by 1% of DMSO, the highest concentration used in this study. Moreover, the potency of ketoconazole to inhibit CYP3A activities was variable, depending on the organic solvent used as a dissolving solvent for ketoconazole. Our data indicate that each organic solvent had an effect on CYP3A4 activity, evaluated by both substrates with different magnitudes. Furthermore, it was shown that the effects of organic solvents on CYP3A activity are substrate-dependent. The present study also shows that methanol had little effect on either substrate.

scholarly journals Poly-L-Arginine Induces Apoptosis of NCI-H292 Cells via ERK1/2 Signaling Pathway

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Ya-Ni Wang ◽  
Ling-Ling Zhang ◽  
Xiao-Yun Fan ◽  
Sha-Sha Wu ◽  
Sheng-Quan Zhang
Keyword(s):  
Epithelial Cells ◽  
Caspase 3 ◽  
Underlying Mechanism ◽  
Mitochondrial Pathway ◽  
Airway Epithelial Cells ◽  
Dependent Manner ◽  
Airway Epithelial ◽  
Cationic Protein ◽  
Concentration Dependent Manner

Cationic protein is a cytotoxic protein secreted by eosinophils and takes part in the damage of airway epithelium in asthma. Poly-L-arginine (PLA), a synthetic cationic protein, is widely used to mimic the biological function of the natural cationic protein in vitro. Previous studies demonstrated the damage of the airway epithelial cells by cationic protein, but the molecular mechanism is unclear. The purpose of this study aimed at exploring whether PLA could induce apoptosis of human airway epithelial cells (NCI-H292) and the underlying mechanism. Methods. The morphology of apoptotic cells was observed by transmission electron microscopy. The rate of apoptosis was analyzed by flow cytometry (FCM). The expressions of the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2), Bcl-2/Bax, and cleaved caspase-3 were assessed by western blot. Results. PLA can induce apoptosis in NCI-H292 cells in a concentration-dependent manner. Moreover, the phosphorylation of the ERK1/2 and the unbalance of Bcl2/Bax, as well as the activation of caspase-3, were involved in the PLA-induced apoptosis. Conclusions. PLA can induce the apoptosis in NCI-H292 cells, and this process at least involved the ERK1/2 and mitochondrial pathway. The results could have some indications in revealing the apoptotic damage of the airway epithelial cells. Besides, inhibition of cationic protein-induced apoptotic death in airway epithelial cells could be considered as a potential target of anti-injury or antiremodeling in asthmatics.

scholarly journals Toxicity Effects of Methylene Blue on Rat Intervertebral Disc Annulus Fibrosus Cells

2019 ◽  
Vol 2 (22.2) ◽  
pp. 155-164
Author(s):  
Liang Zhang
Keyword(s):  
Methylene Blue ◽  
Cell Apoptosis ◽  
Annulus Fibrosus ◽  
Caspase 3 ◽  
In Vitro Study ◽  
Collagen Type I ◽  
Type I ◽  
Dependent Manner ◽  
Concentration Dependent Manner

Background: There is an increasing local application of methylene blue (MB) in the treatment of discogenic low back pain (LBP) and percutaneous transforaminal endoscopic discectomy (PTED) procedures. MB could generate DNA damage and induce apoptosis in different cell types; however, the effects of MB on intervertebral disc (IVD) annulus fibrosus (AF) cells are not clearly understood. Objective: The objective of this study was to investigate the effects of different concentrations of MB on rat AF cells in vitro. Study Design: This study used an experimental design. Setting: This research was conducted at the Orthopaedic Institute of the Clinical Medical College of Yangzhou University. Methods: AF cells were isolated and cultured with different concentrations of MB (0, 2, 20, and 200 μg/mL) and assessed to determine the possible cytotoxic effects of MB. The cell proliferation was detected by Cell Counting Kit-8 (CCK-8) assay. The inverted phase-contrast microscopy was used to perform morphological observation of apoptotic cells, and flow cytometry was used to measure the incidence of cell apoptosis. The mRNA and protein expression levels of apoptosis-associated genes (caspase-3, Bcl-2, and Bax) and other related genes (collagen type I, transforming growth factor β1 [TGF-β1], fibroblast growth factor [bFGF], and tissue inhibitor of metalloproteinase-1 [TIMP-1]) were analyzed by quantitative real-time PCR (RT-PCR) and Western blotting. Results: Our results indicated that MB reduced cell viability in a concentration- and timedependent manner. MB also induced marked AF cell apoptosis in a concentration-dependent manner observed by inverted phase-contrast microscopy, flow cytometry, and indicated by the increased expression of caspase-3. Both RT-PCR and Western blotting revealed significant upregulation of Bax and caspase-3 expression levels accompanied by decreased expression of Bcl2 in a concentration-dependent manner. Moreover, collagen type I, TGF-β1, bFGF, and TIMP-1 mRNA and protein levels were also found to be decreased by MB in a concentration-dependent manner. Limitations: Limitations of this study were the in vitro study design and lack of in vivo validation of the observed effects of MB on human IVD cells. Conclusions: Our results indicate that a high concentration of MB can not only inhibit proliferation and paracrine function of AF cells, but can also induce cell apoptosis in a concentration-dependent manner, suggesting that it is necessary to choose low concentrations of MB in practical application and limit the use of MB in the treatment of discogenic LBP to research protocols. Key words: Methylene blue, annulus fibrosus cell, proliferation, apoptosis, paracrine

scholarly journals Maitotoxin activates cation channels distinct from the receptor-activated non-selective cation channels of HL-60 cells

1994 ◽  
Vol 301 (2) ◽  
pp. 437-441 ◽  
Author(s):  
I F Musgrave ◽  
R Seifert ◽  
G Schultz
Keyword(s):  
Calcium Concentration ◽  
Cation Channels ◽  
Free Calcium ◽  
Dependent Manner ◽  
Dibutyryl Cyclic Amp ◽  
Myristate Acetate ◽  
Atpase Inhibitor ◽  
Concentration Dependent Manner ◽  
Free Calcium Concentration ◽  
Stimulation Of

We investigated whether maitotoxin activates non-selective cation channels, as was recently proposed [Soergel, Yasumoto, Daly and Gusovsky (1992) Mol. Pharmacol. 41, 487-493]. Stimulation of dibutyryl cyclic AMP-differentiated HL-60 cells with the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP; 0.1 microM), the Ca(2+)-ATPase inhibitor thapsigargin (0.1 microM) or maitotoxin (25 ng/ml) resulted in an increase in cytoplasmic free calcium concentration ([Ca2+]i). Unlike fMLP and thapsigargin, maitotoxin produced no increase in [Ca2+]i in the absence of extracellular Ca2+. The increase in [Ca2+]i induced by fMLP was blocked by pretreatment with pertussis toxin (100 ng/ml for 24 h) but not that induced by maitotoxin. Similarly, the increase in [Ca2+]i produced by fMLP but not that produced by maitotoxin was inhibited by pretreatment with phorbol myristate acetate (100 ng/ml). Both fMLP- and maitotoxin-induced increases in [Ca2+]i were blocked by 1-(beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenylethyl)-1H-imid azole hydrochloride (SKF 96365) in a concentration-dependent manner. However, the maitotoxin-induced increase in [Ca2+]i was more sensitive to inhibition by SKF 96365 than the fMLP-induced increase. fMLP-induced increases in [Ca2+]i were blocked by cations with Gd3+ being more effective than Cd2+, whereas for maitotoxin Cd2+ was more effective than Gd3+. Both fMLP and thapsigargin stimulated quenching of Fura-2 fluorescence in the presence of extracellular Mn2+, whereas maitotoxin produced no Mn2+ quenching. Taken together these results suggest that maitotoxin does not stimulate the nonselective cation channel activated by fMLP, but instead activates Ca2+ influx by a different mechanism.

Cytotoxic and Proapoptotic Activity of Diterpenoids from in vitro Cultivated Salvia sclarea Roots. Studies on the Leukemia Cell Lines

2006 ◽  
Vol 61 (7-8) ◽  
pp. 483-488 ◽  
Author(s):  
Marek Różalski ◽  
Łukasz Kuźma ◽  
Halina Wysokińska ◽  
Urszula Krajewska
Keyword(s):  
Caspase 3 ◽  
Leukemia Cell ◽  
Multidrug Resistant ◽  
Dependent Manner ◽  
Transformed Roots ◽  
Concentration Dependent Manner ◽  
Salvia Sclarea ◽  
Leukemia Cell Lines ◽  
High Cytotoxicity

Four diterpenoids, ferruginol, salvipisone, aethiopinone and 1-oxoaethiopinone, were isolated from transformed roots of Salvia sclarea. Salvipisone and aethiopinone showed relatively high cytotoxicity against HL-60 and NALM-6 leukemia cells (IC50 range 0.6-7.7 μg/ mL which is equal to 2.0-24.7 μᴍ), whereas 1-oxoaethiopinone and ferruginol were less active in this regard. Moreover, we have found that all four diterpenoids of S. sclarea had equal cytotoxic activity against parental HL-60 and multidrug-resistant HL-60 ADR cells, what indicates that they are poor substrates for transport by multidrug resistance-associated protein (MRP1). Caspase-3 activity determinations showed that salvipisone and aethiopinone were able to induce apoptosis in a time- and concentration-dependent manner. The results obtained in this study show that S. sclarea diterpenoids aethiopinone and salvipisone may be useful in the treatment of human cancers, especially in the case of drug resistance.

scholarly journals Low-Level Laser Irradiation Modulated Viability of Normal and Tumor Human Lymphocytes In Vitro

2020 ◽  
Vol 11 (2) ◽  
pp. 174-180
Author(s):  
Hesam Saghaei Bagheri ◽  
Seyed Hossein Rasta ◽  
Seyedeh Momeneh Mohammadi ◽  
Ali Akbar Rahim Rahimi ◽  
AliAkbar Movassaghpour ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Line ◽  
Laser Irradiation ◽  
Caspase 3 ◽  
Mononuclear Cells ◽  
Human Lymphocytes ◽  
Dependent Manner ◽  
Ki 67 ◽  
Protein Levels

Introduction: Laser radiation is a promising strategy against various malignancies. Recent studies have shown that the application of low-power laser therapy (LPLT) at different doses and exposure times could modulate the growth dynamic of tumor cells. Based on the type of laser, LPLT could potentially trigger cell proliferation, differentiation, and apoptosis in different cell lines. Methods: In this study, MTT assay was used to monitor the effect of low and high laser intensities on the viability of normal and cancer lymphocytes. The protein levels of Ki-67 (a proliferation marker) and Caspase-3 (an apoptosis factor) were measured in human peripheral mononuclear cells (PBMCs) and the B-lymphoblastic cell line (Nalm-6) using flow cytometry after being-exposed to 630-nm LPLT at low (2, 4, 6, and 10 J/cm2 ) and high (15, 30, 60, and 120 J/cm2 ) energy densities in a continuous mode for 48 and 72 hours. Results: By using higher energy densities, 60 and 120 J/cm2 , a significant decrease was shown in the viability of Nalm-6 cells, which reached 6.6 and 10.1% after 48 hours compared to the control cells (P<0.05). Notably, Cell exposure to doses 30, 60, and 120 J/cm2 yielded 7.5, 12.9, and 21.6 cell viability reduction after 72 hours. The collected data showed that the high-intensity parameters of LPLT (15 to 120 J/cm2 ) promoted significant apoptotic changes in the exposed cells coincided with the activation of Caspase-3 compared to the none-treated control cells (P<0.05). The data further showed the stimulation of the Ki-67 factor both in primary PBMCs and the lymphoblastic cell line treated with LPLT at energy densities of 4 and 6 J/cm2 (P<0.05), indicating enhanced cell proliferation. Similar to Nalm-6 cells, primary PBMCs showed apoptosis after 48 hours of being exposed to doses 60, and 120 J/cm2 , indicated by increased Caspase-3 levels (P<0.05). As expected, the Nalm-6 cells were resistant to cytotoxic effects of laser irradiation in the first 48 hours (P>0.05) compared to normal PBMCs. The exposure of Nalm-6 cells to low-intensity laser intensities increased a proliferation rate compared to the PBMCs treated with the same doses. Conclusion: We showed the potency of LPLT in the induction of apoptosis and proliferation in human primary PBMCs and Nalm-6 cells in a dose and time-dependent manner after 72 hours.

scholarly journals In Vitro Evidence for Effects of Magnesium Supplementation on Quinolone-treated Horse and Dog Chondrocytes

2001 ◽  
Vol 38 (2) ◽  
pp. 143-148 ◽  
Author(s):  
M. Egerbacher ◽  
B. Wolfesberger ◽  
C. Gabler
Keyword(s):  
Cell Proliferation ◽  
Magnesium Deficiency ◽  
Great Part ◽  
Control Group ◽  
Dependent Manner ◽  
Free Medium ◽  
Culture Dish ◽  
Magnesium Supplementation ◽  
Concentration Dependent Manner

Quinolones and magnesium deficiency cause similar lesions in joint cartilage of young animals. Chondrocytes cultivated in the presence of quinolones and in Mg-free medium show severe alterations in cytoskeleton and decreased ability to adhere to the culture dish. We investigated whether Mg2+ supplementation can prevent quinolone-mediated effects on chondrocytes in vitro. Chondrocytes cultivated in Dulbecco's modified Eagle's medium/HAM's F-12 medium were treated with ciprofloxacin (80 and 160 μg/ml) and enrofloxacin (100 and 150 μg/ml). Mg2+ was added at a concentration of 0.0612 mg/ml (MgCl) and 0.0488 mg/ml (MgSO4) or a triple dose. In addition, cells were cultivated in Mg-free medium and accordingly treated with Mg2+ supplementation. After 5 days in culture, the number of adherent cells per milliliter was determined. The number of chondrocytes in quinolone-treated groups decreased to 12-36% that of the control group within the culture period. With Mg2+ supplementation, the number of attached cells increased to 40-70% that of control cells. The threefold dose of Mg2+ led to better results than did the single dose. Cell proliferation tested by immunohistochemical staining with Ki67 (clone MIB5) decreased from 70% in control groups to 55%, 48%, and 30% in enrofloxacintreated groups in a concentration dependent manner (50, 100, and 150 μg/ml). Addition of Mg2+ did not increase the rate of cell proliferation. These results suggest that a great part of quinolone-induced damage is due to magnesium complex formation, as Mg2+ supplementation is able to reduce the effects in vitro. However, quinolone effects on cell proliferation seem to be an independent process that is not influenced by magnesium supplementation.

scholarly journals Glycerol 3-phosphate-induced ATP production in intact mitochondria from pancreatic B-cells

1995 ◽  
Vol 312 (1) ◽  
pp. 287-292 ◽  
Author(s):  
L Å Idahl ◽  
N Lembert
Keyword(s):  
Hill Coefficient ◽  
Mitochondrial Activity ◽  
Free Calcium ◽  
Dependent Manner ◽  
Glycerol Phosphate ◽  
Concentration Dependent Manner ◽  
Atp Production ◽  
Functional Defect ◽  
Order Of Magnitude ◽  
Free Calcium Concentration

A bioluminescent method is presented that allows monitoring of ATP production from mitochondria corresponding to one islet of Langerhans per sample. In mitochondria from ob/ob mice Ca2+ stimulates the ATP production in the presence of L-glycerol 3-phosphate (GP) by reducing the Km for GP by one order of magnitude to about 3 mM. Maximal ATP production in the presence of Ca2+ (200 nM) is obtained at 10 mM GP. The free calcium concentration required to reach half-maximal stimulation (K0.5Ca2+) depends on the GP concentration, thus half-maximal effects are observed at about 80 nM at low GP (1 mM) and 10 nM at high GP (10 mM). Sodium can replace Ca2+ as a stimulator of GP-induced ATP production. It activates ADP phosphorylation by B-cell mitochondria in a sigmoidal concentration-dependent manner in the absence of Cs2+ (Hill coefficient 2.3 +/- 0.2) but does not change K0.5ca2+ nor the maximal mitochondrial activity. Ca2+ concentrations higher than 300 nM are inhibitory at all tested substrate concentrations. Mitochondria from ob/ob mice showed no functional defect when compared with normal controls. It is concluded that activation of the glycerol phosphate shuttle may not be the main coupling site for glucose-induced insulin release at maximal cytoplasmic Ca2+ levels.

scholarly journals Effects of mTOR inhibitor, everolimus, on proliferation, autophagy and temozolomide sensitivity of glioma cells

2020 ◽  
Vol 19 (1) ◽  
pp. 77-82
Author(s):  
Wei Dong ◽  
Jialiang Wang ◽  
Haipeng Liu ◽  
Shuo Sun ◽  
Yanbin Wang
Keyword(s):  
Cell Proliferation ◽  
Western Blot ◽  
Glial Cells ◽  
Glioma Cell ◽  
Mtor Inhibitor ◽  
Glioma Cells ◽  
Expression Level ◽  
Dependent Manner ◽  
Concentration Dependent Manner

Purpose: To study the effect of the mTOR inhibitor, everolimus, on glioma cell proliferation, autophagy, and drug sensitivity to temozolomide (TMZ).Methods: Human glioma cell lines were cultured in vitro, and the effects of different concentrations of everolimus on the proliferation of brain glial cells were determined using CCK-8 method. The effect of different concentrations of everolimus on brain glial cell levels of autophagy protein were assayed by western blot method.Results: The results of CCK-8 analysis showed that everolimus inhibited the proliferation of glial cells in a time- and concentration-dependent manner. Western blot results showed that the expression levels of autophagy proteins, LC3-II and LC3-II/I, were gradually and concentration-dependently up-regulated, while p62 protein level was gradually decreased concentration-dependently, when compared with blank control (p < 0.05). Treatment with different concentrations of TMZ alone, and in combination with everolimus for 48 h inhibited the proliferation of brain glial cells in a concentration-dependent manner, but the inhibition due to TMZ-everolimus combination was significantly higher than that of TMZ singletreatment (p < 0.05). After 48 h, the expression level of Beclin-1 increased with the ratio of LC3-II/LC-I in TMZ-everolimus group, while the expression level of p62 decreased, when compared with TMZ alone, or control (p < 0.05).Conclusion: Everolimus significantly inhibits the proliferation of glioma cells and promotes the occurrence of autophagy. Combined use of TMZ and everolimus significantly enhances the sensitivity of TMZ to glioma cells, inhibits cell proliferation, and promotes autophagy better than TMZ alone. Keywords: mTOR inhibitor, Everolimus, Glioma cells, Proliferation, Autophagy

scholarly journals Effects of Inhibin A on Apoptosis and Proliferation of Bovine Granulosa Cells

Animals ◽  
2020 ◽  
Vol 10 (2) ◽  
pp. 367 ◽  
Author(s):  
Huitao Xu ◽  
Adnan Khan ◽  
Shanjiang Zhao ◽  
Huan Wang ◽  
Huiying Zou ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Granulosa Cells ◽  
Caspase 3 ◽  
Nuclear Antigen ◽  
Molecular Response ◽  
Dependent Manner ◽  
Inhibin A ◽  
The Impact

Inhibin A is well known for its inhibitory properties against follicle-stimulating hormone (FSH), released through a pituitary–gonadal negative feedback loop to regulate follicular development. Ovarian folliculogenesis, hormonal biosynthesis, and gametogenesis are dependent on inhibins, playing vital roles in promoting or inhibiting cell proliferation. The present study explored the physiological and molecular response of bovine granulosa cells (GCs) to different concentrations of inhibin A in vitro. We treated the primary GCs isolated from ovarian follicles (3–6 mm) with different levels of inhibin A (20, 50, and 100 ng/mL) along with the control (0 ng/mL) for 24 h. To evaluate the impact of inhibin A on GCs, several in vitro cellular parameters, including cell apoptosis, viability, cell cycle, and mitochondrial membrane potential (MMP) were detected. Besides, the transcriptional regulation of pro-apoptotic (BAX, Caspase-3) and cell proliferation (PCNA, CyclinB1) genes were also quantified. The results indicated a significant (p < 0.05) increase in the cell viability in a dose-dependent manner of inhibin A. Likewise, MMP was significantly (p < 0.05) enhanced when GCs were treated with high doses (50, 100 ng/mL) of inhibin A. Furthermore, inhibin A dose (100 ng/mL) markedly improved the progression of the G1 phase of the cell cycle and increased the cell number in the S phase, which was supported by the up-regulation of the proliferating cell nuclear antigen PCNA (20, 50, and 100ng/mL) and CyclinB (100 ng/mL) genes. In addition, higher doses of inhibin A (50 and 100 ng/mL) significantly (p < 0.05) decreased the apoptotic rate in GCs, which was manifested by down regulating BAX and Caspase-3 genes. Conclusively, our study presented a worthy strategy for the first time to characterize the cellular adaptation of bovine GCs under different concentrations of inhibin A. Our results conclude that inhibin A is a broad regulatory marker in GCs by regulating apoptosis and cellular progression.

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