Determination of major metabolites of MAM-2201 and JWH-122 in in vitro and in vivo studies to distinguish their intake

ABSTRACT This investigation examined the effects of environmental alteration on the virulence of the oral treponemes Treponema denticolaand Treponema pectinovorum. The environmental effects were assessed by using a model of localized inflammatory abscesses in mice. In vitro growth of T. denticola and T. pectinovorum as a function of modification of the cysteine concentration significantly enhanced abscess formation and size. In contrast, growth of T. denticola or T. pectinovorum under iron-limiting conditions (e.g., dipyridyl chelation) had no effect on abscess induction in comparison to that when the strains were grown under normal iron conditions. In vivo modulation of the microenvironment at the focus of infection with Cytodex beads demonstrated that increasing the local inflammation had no effect on lesion induction or size. In vivo studies involved the determination of the effects of increased systemic iron availability (e.g., iron dextran or phenylhydrazine) on the induction, kinetics, and size of lesions. T. denticola induced significantly larger lesions in mice with iron pretreatment and demonstrated systemic manifestations of the infectious challenge and an accompanying spreading lesion with phenylhydrazine pretreatment (e.g., increases in circulating free hemoglobin). In contrast, T. pectinovorum virulence was minimally affected by this in vivo treatment to increase iron availability. T. denticolavirulence, as evaluated by lesion size, was increased additively by in vivo iron availability, and cysteine modified growth of the microorganism. Additionally, galactosamine sensitized mice to a lethal outcome following infection with both T. denticola andT. pectinovorum, suggesting an endotoxin-like activity in these treponemes. These findings demonstrated the ability to modify the virulence capacity of T. denticola andT. pectinovorum by environmental conditions which can be evaluated by using in vivo murine models.

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Determination of Adhesion and Colonisation Intensity of Staphylococcus epidermidis and Pseudomonas aeruginosa on the Surface of Originally Synthesised Biomaterials in vitro and in vivo Studies, and Their Impact on Inflammatory Cytokine Expression in Tissues. Summary of the Doctoral Thesis

10.25143/prom-rsu_2016-10_dts ◽

2016 ◽

Author(s):

◽

Reinis Aigars

Keyword(s):

Pseudomonas Aeruginosa ◽

Staphylococcus Epidermidis ◽

Inflammatory Cytokine ◽

Cytokine Expression ◽

In Vivo Studies ◽

Doctoral Thesis

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Mitochondrial Neurogastrointestinal Encephalomyopathy (MNGIE): Biochemical Features and Therapeutic Approaches

Bioscience Reports ◽

10.1007/s10540-007-9043-2 ◽

2007 ◽

Vol 27 (1-3) ◽

pp. 151-163 ◽

Cited By ~ 43

Author(s):

M. C. Lara ◽

M. L. Valentino ◽

J. Torres-Torronteras ◽

M. Hirano ◽

R. Martí

Keyword(s):

Molecular Genetic ◽

In Vivo Studies ◽

Gene Encoding ◽

Mitochondrial Neurogastrointestinal Encephalomyopathy ◽

Mtdna Replication ◽

Biochemical Features

Over the last 15 years, important research has expanded our knowledge of the clinical, molecular genetic, and biochemical features of mitochondrial neurogastrointestinal encephalomyopathy (MNGIE). The characterization of mitochondrial involvement in this disorder and the seminal determination of its genetic cause, have opened new possibilities for more detailed and deeper studies on the pathomechanisms in this progressive and fatal disease. It has been established that MNGIE is caused by mutations in the gene encoding thymidine phosphorylase (TP), which lead to absolute or nearly complete loss of its catalytic activity, producing systemic accumulations of its substrates, thymidine (dThd) and deoxyuridine (dUrd). Findings obtained from in vitro and in vivo studies indicate that the biochemical imbalances specifically impair mitochondrial DNA (mtDNA) replication, repair, or both leading to mitochondrial dysfunction. We have proposed that therapy for MNGIE should be aimed at reducing the concentrations of these toxic nucleosides to normal or nearly normal levels. The first treatment, allogeneic stem-cell transplantation (alloSCT) reported in 2006, produced a nearly full biochemical correction of the dThd and dUrd imbalances in blood. Clinical follow-up of this and other patients receiving alloSCT is necessary to determine whether this and other therapies based on a permanent restoration of TP will be effective treatment for MNGIE.

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Gouda Cheese with Modified Content of β-Casein as a Source of Peptides with ACE- and DPP-IV-Inhibiting Bioactivity: A Study Based on In Silico and In Vitro Protocol

International Journal of Molecular Sciences ◽

10.3390/ijms22062949 ◽

2021 ◽

Vol 22 (6) ◽

pp. 2949

Author(s):

Anna Iwaniak ◽

Damir Mogut ◽

Piotr Minkiewicz ◽

Justyna Żulewska ◽

Małgorzata Darewicz

Keyword(s):

In Silico ◽

Ace Inhibition ◽

Water Soluble ◽

In Vivo Studies ◽

Quantitative Parameters ◽

Dpp Iv ◽

Gouda Cheese

In silico and in vitro methods were used to analyze ACE- and DPP-IV-inhibiting potential of Gouda cheese with a modified content of β-casein. Firstly, the BIOPEP-UWM database was used to predict the presence of ACE and DPP-IV inhibitors in casein sequences. Then, the following Gouda cheeses were produced: with decreased, increased, and normative content of β-casein after 1 and 60 days of ripening each (six variants in total). Finally, determination of the ACE/DPP-IV-inhibitory activity and the identification of peptides in respective Gouda-derived water-soluble extracts were carried out. The identification analyses were supported with in silico calculations, i.e., heatmaps and quantitative parameters. All Gouda variants exhibited comparable ACE inhibition, whereas DPP-IV inhibition was more diversified among the samples. The samples derived from Gouda with the increased content of β-casein (both stages of ripening) had the highest DPP-IV-inhibiting potency compared to the same samples measured for ACE inhibition. Regardless of the results concerning ACE and DPP-IV inhibition among the cheese samples, the heatmap showed that the latter bioactivity was predominant in all Gouda variants, presumably because it was based on the qualitative approach (i.e., peptide presence in the sample). Our heatmap did not include the bioactivity of a single peptide as well as its quantity in the sample. In turn, the quantitative parameters showed that the best sources of ACE/DPP-IV inhibitors were all Gouda-derived extracts obtained after 60 days of the ripening. Although our protocol was efficient in showing some regularities among Gouda cheese variants, in vivo studies are recommended for more extensive investigations of this subject.

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Determination of potential solvents for novel N-substituted 5-(phenylamino)uracil derivatives and evaluation of their cytotoxic effects on Vero 76 Cells

Asia Pacific Journal of Molecular Biology and Biotechnology ◽

10.35118/apjmbb.2019.027.4.03 ◽

2019 ◽

pp. 19-29

Author(s):

Noor Fahitah Abu Hanipah ◽

Noor Farah Omar Ahmad ◽

Minaketan Tripathy ◽

Elena Gureeva ◽

Michail Novikov ◽

...

Keyword(s):

Cell Viability ◽

Sodium Benzoate ◽

Aqueous Media ◽

In Vivo Studies ◽

Cytotoxic Effects ◽

Uracil Derivatives ◽

Cytotoxicity Assays

N-substituted 5-(phenylamino)uracil derivatives have recently shown to possess potential antiviral properties. However, the high lipophilicity of these compounds has limited their ability to be dissolved in aqueous media for further in vitro and in vivo studies. This study aimed to determine the potential solvents for novel N-substituted 5-(phenylamino)uracil compounds and to evaluate the cytotoxic effects of these solvents on Vero 76 cells. Eight solvents, namely acetone, methanol, ethanol, dimethyl sulfoxide (DMSO), polyvinylpyrrolidone, nicotinamide, L-arginine, and sodium benzoate, were used to dissolve 1600 µM each of compound Z214 and compound Z276, which were chosen as the representatives of novel N-substituted 5-(phenylamino)uracil derivatives. Only L-arginine (700 mM), sodium benzoate (1500 mM), and DMSO (128 mM) were able to solubilise both compounds. Cytotoxicity assays on Vero 76 cells have shown that the maximum concentrations of L-arginine, sodium benzoate, and DMSO that demonstrated 100% cell viability were 108 mM, 10 mM, and 211 mM respectively. L-arginine at concentrations ranged from 215 mM to 860 mM have shown to significantly increased cell proliferation; while both sodium benzoate and DMSO have significantly reduced cell viability at concentrations ≥ 10 mM and ≥ 211 mM respectively. CC50 values were 23.22 mM and 214.92 mM for sodium benzoate and DMSO respectively. The findings in this study revealed that DMSO at a concentration of 211 mM was found to be the most appropriate solvent to solubilise 1600 µM and below of novel N-Substituted 5-(phenylamino)uracil derivatives.

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In vivo studies with an intravascular and intracardiac reflection oximeter

Journal of Applied Physiology ◽

10.1152/jappl.1962.17.3.552 ◽

1962 ◽

Vol 17 (3) ◽

pp. 552-558 ◽

Cited By ~ 49

Author(s):

Y. Enson ◽

W. A. Briscoe ◽

M. L. Polanyi ◽

A. Cournand

Keyword(s):

Oxygen Saturation ◽

Glass Fibers ◽

Blood Stream ◽

In Vivo Studies ◽

Pulsatile Blood Flow ◽

Cardiac Catheter ◽

Reflected Light

An oximeter is described which employs two bundles of flexible glass fibers to conduct appropriately filtered light into, and that light diffusely reflected by the blood out of, the blood stream for the determination of oxygen saturation or dye concentration within blood flowing past the tip of either an arterial needle or a cardiac catheter which contains both bundles. The ratio of intensities of the reflected light at two wavelengths is linearly related to oxygen saturation (IRR805/ IRR660) and dye concentration (IRR900/IRR805). Data is reported in vivo and in vitro with respect to accuracy of the determinations (± 1.9%). The effect of patient-to-patient variation in hematocrit and other factors, and of pulsatile blood flow, is described. Application of the technique to physiologic study is illustrated, and theoretical aspects of reflection oximetry, as they apply to the instrument, are discussed. Submitted on November 17, 1961

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Lack of Evidence for In Vivo Transformation of Zidovudine Triphosphate to Stavudine Triphosphate in Human Immunodeficiency Virus-Infected Patients

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.50.3.835-840.2006 ◽

2006 ◽

Vol 50 (3) ◽

pp. 835-840 ◽

Cited By ~ 4

Author(s):

Margarita Meléndez ◽

Raúl Blanco ◽

Wilfredo Delgado ◽

Rosario García ◽

Jorge Santana ◽

...

Keyword(s):

Mass Spectrometry ◽

Human Immunodeficiency Virus ◽

High Performance ◽

In Vivo Studies ◽

Tandem Mass ◽

Limit Of Quantitation ◽

Immunodeficiency Virus

ABSTRACT The in vivo and in vitro determination of significant intracellular stavudine (d4T) triphosphate (d4TTP) concentrations in human immunodeficiency virus (HIV)-infected subjects and NS-1 cells treated with zidovudine (ZDV) has recently been reported. This study was conducted to corroborate these findings with in vivo samples from HIV-infected subjects taking ZDV and in vitro CEMSS cells incubated with different ZDV concentrations. Previously, we have reported on our validated high-performance liquid chromatography coupled with tandem mass spectrometry methodology for the simultaneous determination of d4TTP, lamivudine triphosphate, and ZDV triphosphate (ZDVTP) concentrations. Using this methodology, we monitored the d4TTP concentration in more than 100 samples from HIV-infected subjects treated with d4T. In addition, we simultaneously monitored the concentrations of d4TTP and ZDVTP in more than 500 samples from HIV-infected individuals who were taking ZDV. Finally, we performed in vitro studies by incubating CEMSS cells with 10 μM, 50 μM, and 100 μM ZDV and monitored the formation of d4TTP at 24 and 48 h. We could measure d4TTP concentrations from HIV-infected individuals with a limit of quantitation (LOQ) of 2.7 fmol/106 cells (total injection, 54 fmol). In the in vivo studies, we measured the d4TTP concentrations among patients receiving d4T treatment, but the samples from patients taking ZDV did not provide d4TTP concentrations above the LOQ. Furthermore, in vitro samples did not produce any signal for d4TTP, despite the detection of substantial ZDVTP concentrations in CEMSS cells. Thus, contrary to the previous report, we found no evidence for the in vivo or in vitro transformation of ZDVTP to d4TTP in HIV-infected subjects or CEMSS cells.

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Metabolism of Thrombopoietin (TPO) In Vivo: Determination of the Binding Dynamics for TPO in Mice

Blood ◽

10.1182/blood.v89.11.4063 ◽

1997 ◽

Vol 89 (11) ◽

pp. 4063-4070 ◽

Cited By ~ 40

Author(s):

Eric Stefanich ◽

Tauri Senn ◽

Ramon Widmer ◽

Christine Fratino ◽

Gilbert-André Keller ◽

...

Keyword(s):

Blood Cell ◽

Platelet Rich Plasma ◽

In Vivo Studies ◽

In Vivo Metabolism ◽

In Vivo Binding ◽

Curve Fit ◽

Degradation Pattern

AbstractPrevious in vivo studies have established that plasma thrombopoietin (TPO) levels are regulated by binding to c-Mpl on platelets and that, in vitro, platelets bind and degrade TPO. To determine if the in vivo metabolism of TPO was specific and saturable, we injected normal CD-1 mice IV with trace amounts of 125I-rmTPO with or without a saturating concentration of rmTPO. The amount of radioactivity present in the spleen, blood cell fraction, platelet fraction, tibia/fibula, and femur was significantly greater in the mice receiving 125I-rmTPO alone. Conversely, the amount of radioactivity present in the plasma was significantly greater in the mice receiving both 125I-rmTPO and rmTPO, thus suggesting the uptake of rmTPO by the spleen, platelets, and bone marrow in vivo was saturable. Platelet and spleen homogenates from animals receiving 125I-rmTPO alone showed a degradation pattern of 125I-rmTPO similar to that observed in vitro using mouse platelet rich plasma. To determine the in vivo binding dynamics for rmTPO, mice were injected with 125I-rmTPO alone or with increasing concentrations of rmTPO; spleen and blood cell-associated radioactivity was determined at 2 hours postinjection. A 4-parameter curve fit of the data indicated that the “in vivo binding affinity” for rmTPO was approximately 6.4 μg/kg. These data indicate that after a dose of approximately 6.4 μg/kg, 50% of all c-Mpl receptors will be saturated with rmTPO. Electron microscopy indicated that radioactivity was present bound to and within megakaryocytes and platelets in both sternum and spleen and platelets in circulation. Together these data demonstrate that in vivo, 125I-rmTPO is mainly metabolized by platelets and to a small extent by cells of the megakaryocyte lineage, via a specific and saturable mechanism.

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Development and Validation of Methods for the Determination of Anthocyanins in Physiological Fluids via UHPLC-MSn

Molecules ◽

10.3390/molecules25030518 ◽

2020 ◽

Vol 25 (3) ◽

pp. 518 ◽

Cited By ~ 1

Author(s):

Michael Kaiser ◽

Lisa Müller-Ehl ◽

Maike Passon ◽

Andreas Schieber

Keyword(s):

Sample Preparation ◽

Preparation Method ◽

In Vivo Studies ◽

Parameter Determination ◽

Consecutive Reaction ◽

Health Promoting ◽

Development And Validation

Recent in vitro and in vivo studies on anthocyanins confirmed numerous health-promoting effects in humans. Daily anthocyanin intake can be estimated via food databases, but the amount absorbed by the organism still remains uncertain because anthocyanin bioavailability is yet to be elucidated in its entirety. For this purpose, suitable and validated methods of sample preparation and analysis are required. Therefore, a sample preparation method for anthocyanin metabolite analysis in plasma was successfully established and validated. The validation yielded acceptable results for the anthocyanins in terms of recovery (54–108%) and precision (coefficient of variation (CV) < 15%). The UHPLC-MS method used in the consecutive reaction monitoring (CRM) mode was sufficiently sensitive, resulting in limits of detection <2.3 ng/mL and limits of quantification < 8.1 ng/mL with associated repeatability of the MS system with CVs of <5.1%. In addition, a method for the sum parameter determination of anthocyanidins in urine comprising solely the evaporation of acidified samples was developed, validated, and successfully applied to real samples. The results showed that this method is applicable for the methylated anthocyanidins, but not for the hydroxylated anthocyanidins, due to the chosen CRM modes required for optimum selectivity.

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Molecular Mechanisms of Canine Osteosarcoma Metastasis

International Journal of Molecular Sciences ◽

10.3390/ijms22073639 ◽

2021 ◽

Vol 22 (7) ◽

pp. 3639

Author(s):

Sylwia S. Wilk ◽

Katarzyna A. Zabielska-Koczywąs

Keyword(s):

Growth Factor ◽

Molecular Mechanisms ◽

Growth Factor Receptor ◽

In Vivo Studies ◽

Epidermal Growth ◽

Metastatic Process ◽

Transcription 3

Osteosarcoma (OSA) represents the most common bone tumor in dogs. The malignancy is highly aggressive, and most of the dogs die due to metastasis, especially to the lungs. The metastatic process is complex and consists of several main steps. Assessment of the molecular mechanisms of metastasis requires in vitro and especially in vivo studies for a full evaluation of the process. The molecular and biological resemblance of canine OSA to its human counterpart enables the utilization of dogs as a spontaneous model of this disease in humans. The aim of the present review article is to summarize the knowledge of genes and proteins, including p63, signal transducer and activator of transcription 3 (STAT3), Snail2, ezrin, phosphorylated ezrin-radixin-moesin (p-ERM), hepatocyte growth factor-scatter factor (HGF-SF), epidermal growth factor receptor (EGFR), miR-9, and miR-34a, that are proven, by in vitro and/or in vivo studies, to be potentially involved in the metastatic cascade of canine OSA. The determination of molecular targets of metastatic disease may enhance the development of new therapeutic strategies.

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