Development and application of an LC–MS/MS method for measuring the effect of (partial) agonists on cAMP accumulation in vitro

Goutier, Wouter; Spaans, P.A.; Neut, Martina A.W. van der; McCreary, Andrew C.; Reinders, Jan Hendrik

doi:10.1016/j.jneumeth.2010.01.027

Cited by 23 publications

(8 citation statements)

References 13 publications

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“…These methods were modified from [ 31 ]. Confluent cultures of hBASMCs in 48-well plates were serum-deprived (6–24 h) and the medium was then replaced with HBSS (300 μL) containing HEPES (5 mM) and BSA (0.1%, w/v).…”

Section: Methodsmentioning

confidence: 99%

Selective inhibition of histamine-evoked Ca2+ signals by compartmentalized cAMP in human bronchial airway smooth muscle cells

Dale

Head

Dowling³

et al. 2018

Cell Calcium

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Section: Methodsmentioning

confidence: 99%

Selective inhibition of histamine-evoked Ca2+ signals by compartmentalized cAMP in human bronchial airway smooth muscle cells

Dale

Head

Dowling³

et al. 2018

Cell Calcium

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“…The supernatant was collected, transferred to new tubes, and stored at Ϫ80°C. AMP, ADP, ATP, and PCr were measured using the modified approach described by Goutier et al (11). In brief, samples were diluted with mobile phase A (10 mmol/l ammonium bicarbonate buffer adjusted to pH 9.4 with ammonium hydroxide in 20% acetonitrile in HPLC-grade water), filtered through a 3,000-molecular weight cutoff device, and directly injected into a liquid chromatography (LC)-electrospray ionization (ES)/multistage mass spectrometry (MS/MS) system.…”

Section: Methodsmentioning

confidence: 99%

Exposing cardiomyocytes to subclinical concentrations of doxorubicin rapidly reduces their creatine transport

Darrabie

Arciniegas

Mantilla

et al. 2012

American Journal of Physiology-Heart and Circulatory Physiology

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Doxorubicin is commonly used to treat leukemia, lymphomas, and solid tumors, such as soft tissue sarcomas or breast cancer. A major side effect of doxorubicin therapy is dose-dependent cardiotoxicity. Doxorubicin's effects on cardiac energy metabolism are emerging as key elements mediating its toxicity. We evaluated the effect of doxorubicin on [(14)C]creatine uptake in rat neonatal cardiac myocytes and HL-1 murine cardiac cells expressing the human creatine transporter protein. A significant and irreversible decrease in creatine transport was detected after an incubation with 50-100 nmol/l doxorubicin. These concentrations are well below peak plasma levels (5 μmol/l) and within the ranges (25-250 nmol/l) for steady-state plasma concentrations reported after the administration of 15-90 mg/m(2) doxorubicin for chemotherapy. The decrease in creatine transport was not solely because of increased cell death due to doxorubicin's cytotoxic effects. Kinetic analysis showed that doxorubicin decreased V(max), K(m), and creatine transporter protein content. Cell surface biotinylation experiments confirmed that the amount of creatine transporter protein present at the cell surface was reduced. Cardiomyocytes rely on uptake by a dedicated creatine transporter to meet their intracellular creatine needs. Our findings show that the cardiomyocellular transport capacity for creatine is substantially decreased by doxorubicin administration and suggest that this effect may be an important early event in the pathogenesis of doxorubicin-mediated cardiotoxicity.

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“…The total run time using pre-sampling was less than 6 min. The method was applied to Chinese hamster ovarian cells cloned and expressing the human dopamine D2L receptor [34].…”

Section: Hydrophilic Interaction Liquid Chromatographymentioning

confidence: 99%

“…The pre-analytical phase, including sampling, transport of material and sample preparation, plays an important role in the whole analytical process. The treatment of the sample is based on the material used for analysis, such as whole blood [12], erythrocytes [13][14][15], mononuclear cells [14,[16][17][18][19][20][21][22][23][24], cultured cells [25][26][27][28][29][30][31][32][33][34][35], plasma [12,19,20,36,37], urine [38], dry blood spots [15] and others [36,[39][40][41][42][43][44][45][46][47]. The enzymes involved in purine and pyrimidine metabolism can fundamentally change the nucleotide pool during the pre-analytical process and this should be stopped as soon as possible.…”

Section: Introductionmentioning

confidence: 99%

Mass Spectrometry for the Sensitive Analysis of Intracellular Nucleotides and Analogues

Mičová¹,

Friedecký²,

Adam³

2017

Mass Spectrometry

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Nowadays, mass spectrometry is very important and widely applied tool in nucleotide analysis. As a result of technological advances in sample preparation, separation and mass spectrometry detection, the developed methods allow sensitive and selective measurement of polar compounds occurring in low levels in various biological matrices. This enables more potential uses in clinical field. Direct methods require no special sample pre-treatment before analysis in contrast to indirect methods, where fractionation, dephosphorylation and purification are needed. The use of ion-pairing agent, ion exchange chromatography with pH gradient, porous graphitic carbon columns and HILIC in liquid chromatography represents the most common methods of nucleotide analysis. High separation efficiency is also achieved with the use of CE with MS detection. Analysis of nucleotides was also described by the means of MALDI-TOF, but poor reproducibility and lack of applications make a limitation for this approach. The chapter summarizes different techniques and approaches for determination of endogenous nucleotides and its analogues in various clinical applications. or deoxyribose) through a glycosidic bond. Nucleotides are formed by phosphorylation of nucleoside molecule. They represent a fundamental building block of the structure of nucleic acids formed by the polymerase-mediated synthesis of DNA and RNA from deoxynucleotides and nucleotides, respectively. All deoxynucleotides are synthesized from the corresponding nucleotides. Nucleotides play another important role in enzyme activation and metabolism. They are incorporated into important cofactors of enzymatic reactions such as transfer of various groups by coenzyme A (e.g. acetyl-CoA and succinyl-CoA) in many metabolic processes, NAD, NADP and FAD as coenzymes of oxidoreductases catalyses redox reactions in the cell. ATP and other nucleoside triphosphates are engaged as source energy providers. In the form of coenzymes, they transfer phosphate residues or nucleotide constituent and participate in the activation (e.g. uridine diphosphate glucose for pasting glucose units to polymeric saccharides or cytidine diphosphate-choline for phospholipid synthesis). Cyclic nucleotides act as secondary messengers in many signal transduction pathways. The most important nucleotide signal molecule is cyclic adenosine monophosphate (cAMP), which is an activator of protein kinases and participates in many metabolic pathways by transferring the effect of hormones into cells. Analogical molecule-cyclic guanosine monophosphate (cGMP)-also acts as a secondary messenger in the phototransduction process.Nucleotide metabolism is one of the main therapeutic cellular targets. Synthetic analogues of these compounds have been widely applied in anti-cancer, anti-viral and immunosuppressive therapies [1][2][3][4][5][6]. Nucleoside analogues (e.g. cytarabine, gemcitabine and zidovudine) are modified at the base or sugar moiety. This 'prodrug' form requires intracellular activation by phosphorylation to mono, di or...

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Development and application of an LC–MS/MS method for measuring the effect of (partial) agonists on cAMP accumulation in vitro

Cited by 23 publications

References 13 publications

Selective inhibition of histamine-evoked Ca2+ signals by compartmentalized cAMP in human bronchial airway smooth muscle cells

Selective inhibition of histamine-evoked Ca2+ signals by compartmentalized cAMP in human bronchial airway smooth muscle cells

Exposing cardiomyocytes to subclinical concentrations of doxorubicin rapidly reduces their creatine transport

Mass Spectrometry for the Sensitive Analysis of Intracellular Nucleotides and Analogues

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