Heat shock increases cytosolic free Ca2+ concentration via Na(+)-Ca2+ exchange in human epidermoid A 431 cells

1992 ◽  
Vol 263 (1) ◽  
pp. C30-C38 ◽  
Author(s):  
J. G. Kiang ◽  
M. L. Koenig ◽  
R. C. Smallridge
Keyword(s):  
Heat Treatment ◽  
Heat Shock ◽  
Time Dependent ◽  
Maximal Velocity ◽  
Dependent Manner ◽  
Michaelis Constant ◽  
Exchange System ◽  
Maximal Increase ◽  
Normal Cells ◽  
Dimethyl Amiloride

This study characterized cytosolic free Ca2+ concentration ([Ca2+]i) in normal and thermally injured human epidermoid A 431 cells. The resting [Ca2+]i in normal cells at 37 degrees C was 87 +/- 5 nM (n = 105). When cells were subjected to hyperthermia (40-50 degrees C), [Ca2+]i increased in a temperature- and time-dependent manner. The maximal increase in cells exposed to 45 degrees C was observed at 20 min; [Ca2+]i returned to normal within 1 h. The heat-induced [Ca2+]i increase depended on the presence of external Ca2+. La3+ and Cd2+ but not Co2+, verapamil, or nifedipine attenuated the heat-induced [Ca2+]i increase. TMB-8 partially blocked the increase in [Ca2+]i but pertussis toxin and cholera toxin pretreatment did not. The magnitude of the heat-induced [Ca2+]i increase or 45Ca2+ uptake depended on the presence of extracellular Na+. Heat treatment reduced the apparent Michaelis constant for external Ca2+ from 490 +/- 91 to 210 +/- 60 microM, whereas the maximal velocity remained the same. The intracellular Na+ concentration decreased 62.5% after heating. The heat-induced [Ca2+]i increase was completely blocked by amiloride (5 microM) and 5'-(N,N-dimethyl)-amiloride (1 microM). These results suggest heat activates the Na(+)-Ca2+ exchange system so as to increase [Ca2+]i and reduce [Na+]i.

Exponential function for calculating saturable enzyme kinetics.

1988 ◽  
Vol 34 (12) ◽  
pp. 2486-2489 ◽  
Author(s):  
F Keller ◽  
C Emde ◽  
A Schwarz
Keyword(s):  
Steady State ◽  
Enzyme Kinetics ◽  
Exponential Function ◽  
Association Constant ◽  
Time Dependent ◽  
Mathematical Representation ◽  
Hyperbolic Function ◽  
Maximal Velocity ◽  
Michaelis Constant ◽  
General Meaning

Abstract Enzyme kinetics are usually described by the Michaelis-Menten equation, where the time-dependent decrease of substrate (-dS/dt) is a hyperbolic function of maximal velocity (Vmax), Michaelis constant (Km), and amount of substrate (S). Because the Michaelis-Menten function in its most general meaning requires an assumption of steady-state, it is less curvilinear than true enzyme kinetics. A saturation-type exponential function is more curvilinear than the hyperbolic function and more closely approximates enzyme kinetics: -dS/dt = Vmax [1 - exp(-S/Km)]. The mathematical representation of enzyme kinetics can be further improved by introducing a deceleration term (Vdec), to make the assumption of a steady state unnecessary. For the action of chymotrypsin on N-acetyltyrosylethylester, the Michaelis-Menten equation yields the following: Vmax = 3.74 mumol/min and Km = 833 mumol. According to decelerated enzyme kinetics, the values Vmax = 4.80 mumol/min, Vdec = 0.0118 mumol/min, and the association constant (Ka) = 0.00111/mumol are more nearly accurate for this reaction (where 1/Ka = 901 mumol approximately Km).

Heat Shock Inhibits NF-kB Activation in a Dose- and Time-Dependent Manner

2005 ◽  
Vol 129 (1) ◽  
pp. 90-93 ◽  
Author(s):  
Michael T. Schell ◽  
Austin L. Spitzer ◽  
Jennifer A. Johnson ◽  
Diana Lee ◽  
Hobart W. Harris
Keyword(s):  
Heat Shock ◽  
Time Dependent ◽  
Dependent Manner

scholarly journals Urocortin increases the expression of heat shock protein 90 in rat cardiac myocytes in a MEK1/2-dependent manner

2002 ◽  
Vol 172 (2) ◽  
pp. 283-293 ◽  
Author(s):  
BK Brar ◽  
J Railson ◽  
A Stephanou ◽  
RA Knight ◽  
DS Latchman
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Cardiac Myocytes ◽  
Primary Cultures ◽  
Heat Shock Protein 90 ◽  
Mitogen Activated Protein Kinase ◽  
Dependent Manner ◽  
Maximal Increase ◽  
Mitogen Activated Protein ◽  
Hsp90 Protein

We have previously demonstrated that urocortin protects cultured cardiac myocytes from ischaemic and reoxygenation injury and decreases the infarct size in the rat heart exposed to regional ischaemia and reperfusion. Urocortin-mediated cardioprotection is via activation of the mitogen-activated protein kinase (MAP kinase, MEK1/2) pathway. In addition, it is well documented that heat shock protein (hsp) 70 and hsp90 are cardioprotective against lethal stress. In this study we show, for the first time, that urocortin induces the expression of hsp90 but not hsp70 in primary cultures of rat neonatal cardiac myocytes. Levels of hsp90 protein increase by 1.5-fold over untreated cells within 10 min of urocortin treatment and are sustained for 24 h with a maximal increase of 2.5-fold at 60 min (P<0.05 at all time points). The increase in hsp90 expression by urocortin was not inhibited by actinomycin D, and urocortin failed to increase hsp90 promoter activity. Urocortin induction of hsp90 was inhibited by the MEK1/2 inhibitor PD98059 (P<0.001) and by cycloheximide, and both inhibitors abrogate urocortin-mediated cardioprotection (P<0.05 for cycloheximide, P<0.001 for PD98059). Hence, MEK1/2 and protein synthesis are involved in the cardioprotective effect of urocortin against hypoxic-mediated cell death, possibly due to an increase in expression of hsp90 protein. This is the first report of heat shock protein induction by urocortin or any other member of the corticotrophin-releasing hormone family.

scholarly journals Ethanolic Neem (Azadirachta indica) Leaf Extract Prevents Growth of MCF-7 and HeLa Cells and Potentiates the Therapeutic Index of Cisplatin

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
Chhavi Sharma ◽  
Andrea J. Vas ◽  
Payal Goala ◽  
Taher M. Gheewala ◽  
Tahir A. Rizvi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cancer Cells ◽  
Time Dependent ◽  
Cytochrome P450 Monooxygenases ◽  
Dependent Manner ◽  
Morphological Examination ◽  
Cell Viability Assay ◽  
Normal Cells ◽  
Neem Leaves ◽  
Mcf 7

The present study was designed to gain insight into the antiproliferative activity of ethanolic neem leaves extract (ENLE) alone or in combination with cisplatin by cell viability assay on human breast (MCF-7) and cervical (HeLa) cancer cells. Nuclear morphological examination and cell cycle analysis were performed to determine the mode of cell death. Further, to identify its molecular targets, the expression of genes involved in apoptosis, cell cycle progression, and drug metabolism was analyzed by RT-PCR. Treatment of MCF-7, HeLa, and normal cells with ENLE differentially suppressed the growth of cancer cells in a dose- and time-dependent manner through apoptosis. Additionally, lower dose combinations of ENLE with cisplatin resulted in synergistic growth inhibition of these cells compared to the individual drugs (combination index <1). ENLE significantly modulated the expression of bax, cyclin D1, and cytochrome P450 monooxygenases (CYP 1A1 and CYP 1A2) in a time-dependent manner in these cells. Conclusively, these results emphasize the chemopreventive ability of neem alone or in combination with chemotherapeutic treatment to reduce the cytotoxic effects on normal cells, while potentiating their efficacy at lower doses. Thus, neem may be a prospective therapeutic agent to combat gynecological cancers.

scholarly journals The Role of Oxidative Stress in Koenimbine-Induced DNA Damage and Heat Shock Protein Modulation in HepG2 Cells

2016 ◽  
Vol 16 (4) ◽  
pp. 563-571 ◽  
Author(s):  
Yahya Hasan Hobani
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Ascorbic Acid ◽  
Heat Shock ◽  
Hepg2 Cells ◽  
Time Dependent ◽  
Dependent Manner ◽  
Hsp 70 ◽  
Hsp 90

Background. Murraya koenigii (L.) Spreng, is a significant herb of traditional Ayurvedic system of medicine. Koenimbine, a carbazole alkaloid isolated from this plant holds antiproliferative and apoptotic effects. The aim of this study was to assess koenimbine-induced DNA damage and to clarify the role of free radicals in cell death mechanisms, using HepG2 cells. Methods. The level of cytotoxicity was assayed by MTT assay. To elucidate the role of glutathione (GSH), the intracellular GSH level was analyzed. The effect of koenimbine in the cell mitochondria was evaluated using mitochondrial membrane potential (MMP) changes. Single cell gel electrophoresis assay was used to examine the level of DNA damage. Heat shock proteins, Hsp 70 and Hsp 90 expressions were checked at mRNA and protein level. Ascorbic acid and catalase were used as control antioxidants. Results. It was observed that koenimbine considerably increased DNA damage in HepG2 cells at subcytotoxic concentrations. Koenimbine induced the increased levels of reactive oxygen species (ROS) and reduction of GSH level in HepG2 cells, together with time-dependent loss of MMP. In addition, results clearly showed that koenimbine encouraged cells to express Hsp 70 and Hsp 90 in a concentration-dependent manner up to a concentration of 100 µM and a time-dependent manner at 24-hour incubation both at transcriptional and translational levels. The antioxidant capacity of ascorbic acid was found to be not as prominent as to catalase throughout the study. Conclusion. Based on these data it can be concluded that koenimbine causes DNA strand breaks in HepG2 cells, probably through oxidative stress. Moreover, the oxidative stress induced was closely associated with MMP reduction and GSH depletion associated with HSP modulation at subcytotoxic concentration.

scholarly journals Mild Hyperthermia-Induced Myogenic Differentiation in Skeletal Muscle Cells: Implications for Local Hyperthermic Therapy for Skeletal Muscle Injury

2018 ◽  
Vol 2018 ◽  
pp. 1-9
Author(s):  
Hojun Lee ◽  
Seung-Jun Choi
Keyword(s):  
Heat Treatment ◽  
Skeletal Muscle ◽  
Treatment Time ◽  
Myogenic Differentiation ◽  
Mitochondrial Dynamics ◽  
Time Dependent ◽  
Dependent Manner ◽  
Myogenic Regulatory Factor ◽  
Mild Hyperthermia ◽  
Mild Heat

The percutaneous application of controlled temperature on damaged muscle is regarded as a prevalent remedy. However, specific mechanisms are not completely understood. Therefore, cellular behaviors of myoblasts were investigated under a physiological hyperthermic temperature. The myoblasts were cultured under no treatment (NT, 37°C, 24 h/day), intermittent heat treatment (IHT, 39°C, 2 h/day), and continuous heat treatment (CHT, 39°C, 24 h/day) during proliferation, migration, or myogenic differentiation. Although the effects of mild heat on migration were not observed, the proliferation was promoted by both IHT and CHT. The myogenic differentiation was also enhanced in a treatment time-dependent manner, as evidenced by an increase in myotube size and fusion index. The gene expressions of mitochondrial biogenesis (Pgc-1α, Nrf1, and Tfam), a subset of mitochondrial dynamics (Mfn1 and Drp1), and a myogenic regulatory factor (myogenin) were increased in a heat treatment time-dependent manner. Interestingly, the mild heat-induced myogenic differentiation and myogenin expression were retarded significantly in PGC-1α-targeted siRNA-transfected cells, suggesting that mild hyperthermia promotes myogenic differentiation via the modulation of PGC-1α. This study provides cellular evidence supporting that local hyperthermic treatment at 39°C is regarded as an effective therapeutic strategy to promote satellite cell activities in regenerating myofibers.

The Natural Flavonoid Naringenin Inhibits the Cell Growth of WilmsTumorinChildren by Suppressing TLR4/NF-κB Signaling

2020 ◽  
Vol 20 ◽  
Author(s):  
Hongtao Li ◽  
Peng Chen ◽  
Lei Chen ◽  
Xinning Wang
Keyword(s):  
Cell Growth ◽  
Western Blot ◽  
Wilms Tumor ◽  
Critical Role ◽  
Time Dependent ◽  
Luciferase Assay ◽  
Dependent Manner ◽  
Tlr4 Expression ◽  
Protein Levels

Background: Nuclear factor kappa B (NF-κB) is usually activated in Wilms tumor (WT) cells and plays a critical role in WT development. Objective: The study purpose was to screen a NF-κB inhibitor from natural product library and explore its effects on WT development. Methods: Luciferase assay was employed to assess the effects of natural chemical son NF-κB activity. CCK-8 assay was conducted to assess cell growth in response to naringenin. WT xenograft model was established to analyze the effect of naringenin in vivo. Quantitative real-time PCR and Western blot were performed to examine the mRNA and protein levels of relative genes, respectively. Results: Naringenin displayed significant inhibitory effect on NF-κB activation in SK-NEP-1 cells. In SK-NEP-1 and G-401 cells, naringenin inhibited p65 phosphorylation. Moreover, naringenin suppressed TNF-α-induced p65 phosphorylation in WT cells. Naringenin inhibited TLR4 expression at both mRNA and protein levels in WT cells. CCK-8 staining showed that naringenin inhibited cell growth of the two above WT cells in dose-and time-dependent manner, whereas Toll-like receptor 4 (TLR4) over expression partially reversed the above phenomena. Besides, naringenin suppressed WT tumor growth in dose-and time-dependent manner in vivo. Western blot found that naringenin inhibited TLR4 expression and p65 phosphorylation in WT xenograft tumors. Conclusion: Naringenin inhibits WT development viasuppressing TLR4/NF-κB signaling

Peroxidases from Tulip Bulbs (Tulipa fosteriana, L.) Oxidize Xenobiotics NNitrosodimethylamine and N-Nitroso-N-methylaniline in vitro

1997 ◽  
Vol 62 (11) ◽  
pp. 1804-1814 ◽  
Author(s):  
Marie Stiborová ◽  
Hana Hansíková
Keyword(s):  
Cytochromes P450 ◽  
Kinetic Studies ◽  
The Other ◽  
Maximal Velocity ◽  
Michaelis Constant ◽  
Other Hand ◽  
Plant Peroxidases ◽  
Tulip Bulbs

Tulip bulbs (Tulipa fosteriana, L.) contain peroxidases catalyzing the oxidation of the xenobiotics N-nitrosodimethylamine (NDMA) and N-nitroso-N-methylaniline (NMA). Three anionic (A1, A2, A3) and four cationic (B, C, D, E) peroxidases were purified from this tissue, partially characterized and used for kinetic studies. Demethylation of NDMA and NMA producing formaldehyde is catalyzed by one anionic (A1) and three cationic (C, D, E) peroxidases. The oxidation of NDMA by tulip peroxidases exhibits the Michaelis-Menten kinetics. The apparent Michaelis constant and the maximal velocity values for this substrate were determined. On the other hand, non-Michaelian kinetics for the NMA oxidation were observed with tulip peroxidases. The most abundant cationic peroxidase (peroxidase C) was used for detailed enzymatic studies. In addition to formation of formaldehyde, methylaniline, aniline, 4-aminophenol and phenol were found to be metabolites formed from NMA. Phenol was formed presumably by N-demethylation via a benzenediazonium ion, while methylaniline, aniline and 4-aminophenol were products of denitrosation of the substrate. The efficiencies of plant peroxidases to oxidize NDMA and NMA in vitro are compared with those of cytochromes P450 and discussed.

scholarly journals Synthesis and Evaluation of the Antitumor Activity of Novel 1-(4-Substituted phenyl)-2-ethyl Imidazole Apoptosis Inducers In Vitro

Molecules ◽  
2020 ◽  
Vol 25 (18) ◽  
pp. 4293
Author(s):  
Zhen-Wang Li ◽  
Chun-Yan Zhong ◽  
Xiao-Ran Wang ◽  
Shi-Nian Li ◽  
Chun-Yuan Pan ◽  
...  
Keyword(s):  
Antitumor Activity ◽  
Tumor Cells ◽  
Antiproliferative Activity ◽  
Antitumor Agent ◽  
Positive Control ◽  
Selectivity Index ◽  
Time Dependent ◽  
Potential Candidate ◽  
Dependent Manner

Novel imidazole derivatives were designed, prepared, and evaluated in vitro for antitumor activity. The majority of the tested derivatives showed improved antiproliferative activity compared to the positive control drugs 5-FU and MTX. Among them, compound 4f exhibited outstanding antiproliferative activity against three cancer cell lines and was considerably more potent than both 5-FU and MTX. In particular, the selectivity index indicated that the tolerance of normal L-02 cells to 4f was 23–46-fold higher than that of tumor cells. This selectivity was significantly higher than that exhibited by the positive control drugs. Furthermore, compound 4f induced cell apoptosis by increasing the protein expression levels of Bax and decreasing those of Bcl-2 in a time-dependent manner. Therefore, 4f could be a potential candidate for the development of a novel antitumor agent.

Sign in / Sign up

Export Citation Format

Share Document