scholarly journals Stemness of Human Pluripotent Cells: Hypoxia-like Response Induced by Low Nitric Oxide

Antioxidants ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 1408
Author(s):  
Estefanía Caballano-Infantes ◽  
Irene Díaz ◽  
Ana Belén Hitos ◽  
Gladys Margot Cahuana ◽  
Antonio Martínez-Ruiz ◽  
...  
Keyword(s):  
Human Escs ◽  
Pluripotent Cells ◽  
No Donor ◽  
Hypoxia Response ◽  
Master Genes ◽  
Low Concentrations ◽  
Human Ipscs ◽  
Advanced Therapies ◽  
Lower Oxygen

The optimization of conditions to promote the stemness of pluripotent cells in vitro is instrumental for their use in advanced therapies. We show here that exposure of human iPSCs and human ESCs to low concentrations of the chemical NO donor DETA/NO leads to stabilization of hypoxia-inducible factors (HIF-1α and HIF-2α) under normoxia, with this effect being dependent on diminished Pro 402 hydroxylation and decreased degradation by the proteasome. Moreover, the master genes of pluripotency, NANOG and OCT-4, were upregulated. NO also induces a shift in the metabolic profile of PSCs, with an increased expression of hypoxia response genes in glycolysis. Furthermore, a reduction in the mitochondrial membrane potential with lower oxygen consumption and increased expression of mitochondrial fusion regulators, such as DRP1, was observed. The results reported here indicate that NO mimics hypoxia response in human PSCs and enhances their stemness properties when cultured under normoxic conditions.

The impact of culture on epigenetic properties of pluripotent stem cells and pre-implantation embryos

2013 ◽  
Vol 41 (3) ◽  
pp. 711-719 ◽  
Author(s):  
Kirsten R. McEwen ◽  
Harry G. Leitch ◽  
Rachel Amouroux ◽  
Petra Hajkova
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Culture Condition ◽  
Reproductive Technologies ◽  
Great Promise ◽  
Global Changes ◽  
Human Escs ◽  
Pluripotent Cells ◽  
The Impact

Cultured pluripotent stem cells hold great promise for regenerative medicine. Considerable efforts have been invested into the refinement and definition of improved culture systems that sustain self-renewal and avoid differentiation of pluripotent cells in vitro. Recent studies have, however, found that the choice of culture condition has a significant impact on epigenetic profiles of cultured pluripotent cells. Mouse and human ESCs (embryonic stem cells) show substantial epigenetic differences that are dependent on the culture condition, including global changes to DNA methylation and histone modifications and, in female human ESCs, to the epigenetic process of X chromosome inactivation. Epigenetic perturbations have also been detected during culture of pre-implantation embryos; limited research undertaken in mouse suggests a direct effect of the in vitro environment on epigenetic processes in this system. Widespread epigenetic changes induced by the culture condition in stem cells thus emphasize the necessity for extensive research into both immediate and long-term epigenetic effects of embryo culture during assisted reproductive technologies.

Kinetics of Various 99mTc-Sn-Pyrophosphate Compounds in the Rat

1977 ◽  
Vol 16 (04) ◽  
pp. 157-162 ◽  
Author(s):  
C. Schümichen ◽  
B. Mackenbrock ◽  
G. Hoffmann
Keyword(s):  
Normal Saline ◽  
Half Life ◽  
Compound A ◽  
Short Half Life ◽  
Low Concentrations ◽  
The Stability ◽  
Kinetics Of ◽  
In Vitro Stability

SummaryThe bone-seeking 99mTc-Sn-pyrophosphate compound (compound A) was diluted both in vitro and in vivo and proved to be unstable both in vitro and in vivo. However, stability was much better in vivo than in vitro and thus the in vitro stability of compound A after dilution in various mediums could be followed up by a consecutive evaluation of the in vivo distribution in the rat. After dilution in neutral normal saline compound A is metastable and after a short half-life it is transformed into the other 99mTc-Sn-pyrophosphate compound A is metastable and after a short half-life in bone but in the kidneys. After dilution in normal saline of low pH and in buffering solutions the stability of compound A is increased. In human plasma compound A is relatively stable but not in plasma water. When compound B is formed in a buffering solution, uptake in the kidneys and excretion in urine is lowered and blood concentration increased.It is assumed that the association of protons to compound A will increase its stability at low concentrations while that to compound B will lead to a strong protein bond in plasma. It is concluded that compound A will not be stable in vivo because of a lack of stability in the extravascular space, and that the protein bond in plasma will be a measure of its in vivo stability.

In Vitro Effects of Ethanol on Rabbit Platelet Aggregation, Secretion of Granule Contents, and Cyclic AMP Levels in the Presence of Prostacyclin

1989 ◽  
Vol 61 (02) ◽  
pp. 254-258 ◽  
Author(s):  
Margaret L Rand ◽  
Peter L Gross ◽  
Donna M Jakowec ◽  
Marian A Packham ◽  
J Fraser Mustard
Keyword(s):  
Cyclic Amp ◽  
Inhibitory Effect ◽  
Rabbit Platelet ◽  
Inhibitory Effects ◽  
Low Concentrations ◽  
Rabbit Platelets ◽  
High Concentrations ◽  
In Vitro Effects ◽  
Aggregation Of Platelets

SummaryEthanol, at physiologically tolerable concentrations, inhibits platelet responses to low concentrations of collagen or thrombin, but does not inhibit responses of washed rabbit platelets stimulated with high concentrations of ADP, collagen, or thrombin. However, when platelet responses to high concentrations of collagen or thrombin had been partially inhibited by prostacyclin (PGI2), ethanol had additional inhibitory effects on aggregation and secretion. These effects were also observed with aspirin- treated platelets stimulated with thrombin. Ethanol had no further inhibitory effect on aggregation of platelets stimulated with ADP, or the combination of ADP and epinephrine. Thus, the inhibitory effects of ethanol on platelet responses in the presence of PGI2 were very similar to its inhibitory effects in the absence of PGI2, when platelets were stimulated with lower concentrations of collagen or thrombin. Ethanol did not appear to exert its inhibitory effects by increasing cyclic AMP above basal levels and the additional inhibitory effects of ethanol in the presence of PGI2 did not appear to be brought about by further increases in platelet cyclic AMP levels.

Aggregation of Cat Platelets in Vitro

1970 ◽  
Vol 23 (03) ◽  
pp. 601-620 ◽  
Author(s):  
Th. B Tschopp
Keyword(s):  
Adenosine Monophosphate ◽  
Blood Collection ◽  
Base Line ◽  
Reversible Aggregation ◽  
Low Concentrations ◽  
Irreversible Aggregation ◽  
High Concentrations ◽  
Two Phases ◽  
Improved Technique

SummaryAggregation of cat platelets in the citrated plasma is examined by means of Born’s absorptiometer. A marked tendency of the platelets of this species to spontaneous aggregation necessitated first of all the development of an improved technique of blood collection.A hypothesis according to which 5-HT is released from the platelets, explains the absence of oscillations on the base line of the absorptiometer, the absence of platelet swelling, when ADP is added, and the effect of stirring on the aggregation curves in cat PRP. The average volume of cat platelets amounts to 10.46 μ3 when directly fixed in the blood, when fixed from PRP to 12.17 μ3, when fixed from stirred PRP to 13.51 μ3.In low concentrations (0.3-2 μM) ADP produce reversible aggregation; in narrowly restricted, individually dissimilar mean concentrations irreversible aggregation in two phases and in high concentrations, irreversible aggregation in one phase. Like ADP serotonin produces 2 phase irreversible aggregation in concentrations of 3-10 μM, but unlike ADP, the aggregation velocity decreases again with high 5-HT concentrations (>100 μM). Adrenaline does not produce aggregation and it is likely that adenosine and adenosine monophosphate inhibit the aggregation by serotonin but not by ADP. Species differences in the aggregation of human, rabbit and cat platelets are discussed.

DEVELOPMENT OF APPROACHES FOR DETECTION OF GENOME-INTEGRATED PROVIRAL DNA OF THE HUMAN IMMUNODEFICIENCY VIRUS (HIV-1) IN ULTRA LOW CONCENTRATIONS USING THE CRISPR/CAS SYSTEM

2020 ◽  
Author(s):  
M.A. Tyumentseva ◽  
◽  
A.I. Tyumentsev ◽  
V.G. Akimkin ◽  
◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Health Monitoring ◽  
Biological Samples ◽  
Proviral Dna ◽  
Guide Rnas ◽  
Ribonucleoprotein Complexes ◽  
Low Concentrations ◽  
Immunodeficiency Virus ◽  
Hiv 1

For the effective functioning of supervisory and health monitoring services, it is necessary to introduce modern molecular technologies into their practice. Therefore, the task of developing new effective methods for detecting pathogen, for example HIV, based on CRISPR/CAS genome editing systems, remains urgent. In the present work, guide RNAs and specific oligonucleotides were developed for preliminary amplification of highly conserved regions of the HIV-1 genome. The developed guide RNAs make it possible to detect single copies of HIV-1 proviral DNA in vitro as part of CRISPR/CAS ribonucleoprotein complexes in biological samples after preliminary amplification.

Effect of C6-Volatiles on Bioluminescent Plant Pathogens

HortScience ◽  
1998 ◽  
Vol 33 (3) ◽  
pp. 557d-557
Author(s):  
Jennifer Warr ◽  
Fenny Dane ◽  
Bob Ebel
Keyword(s):  
Biological Activity ◽  
Bacterial Growth ◽  
Xanthomonas Campestris ◽  
Plant Pathogens ◽  
Genetically Engineered ◽  
The Other ◽  
Computer Assisted ◽  
Signal Molecules ◽  
Low Concentrations

C6 volatile compounds are known to be produced by the plant upon pathogen attack or other stress-related events. The biological activity of many of these substances is poorly understood, but some might produce signal molecules important in host–pathogen interactions. In this research we explored the possibility that lipid-derived C6 volatiles have a direct effect on bacterial plant pathogens. To this purpose we used a unique tool, a bacterium genetically engineered to bioluminesce. Light-producing genes from a fish-associated bacterium were introduced into Xanthomonas campestris pv. campestris, enabling nondestructive detection of bacteria in vitro and in the plant with special computer-assisted camera equipment. The effects of different C6 volatiles (trans-2 hexanal, trans-2 hexen-1-ol and cis-3 hexenol) on growth of bioluminescent Xanthomonas campestris were investigated. Different volatile concentrations were used. Treatment with trans-2 hexanal appeared bactericidal at low concentrations (1% and 10%), while treatments with the other volatiles were not inhibitive to bacterial growth. The implications of these results with respect to practical use of trans-2 hexanal in pathogen susceptible and resistant plants will be discussed.

Uncertainties Associated with Assessing the Risk of an Endocrine Active Substance in the Canadian Environment

2001 ◽  
Vol 36 (2) ◽  
pp. 319-330 ◽  
Author(s):  
Mark Servos ◽  
Don Bennie ◽  
Kent Burnison ◽  
Philippa Cureton ◽  
Nicol Davidson ◽  
...  
Keyword(s):  
Risk Assessment ◽  
Endocrine Disruption ◽  
Endocrine System ◽  
Problem Formulation ◽  
Normal Function ◽  
Weight Of Evidence ◽  
Biological Responses ◽  
Low Concentrations ◽  
Multigenerational Effects

Abstract A number of biological responses and multigenerational effects, mediated through the disruption of endocrine systems, have been observed in biota exposed to relatively low concentrations of environmental contaminants. These types of responses need to be considered within a weight of evidence approach in our risk assessment and risk management frameworks. However, including endocrine responses in an environmental risk assessment introduces a number of uncertainties that must be considered. A risk assessment of nonylphenol and nonylphenol polyethoxylates (NP/NPE) is used as a case study to demonstrate the sources and magnitude of some of the uncertainties associated with using endocrine disruption as an assessment endpoint. Even with this relatively well studied group of substances, there are substantial knowledge gaps which contribute to the overall uncertainties, limiting the interpretation within the risk assessment. The uncertainty of extrapolating from in vitro or biochemical responses to higher levels of organization or across species is not well understood. The endocrine system is very complex and chemicals can interact or interfere with the normal function of endocrine systems in a number of ways (e.g., receptors, hormones) which may or may not result in an adverse responses in the whole organism. Using endocrine responses can lead to different conclusions than traditional endpoints due to a variety of factors, such as differences in relative potencies of chemicals for specific endpoints (e.g., receptor binding versus chronic toxicity). The uncertainties can also be considerably larger and the desirability of using endocrine endpoints should be carefully evaluated. Endocrine disruption is a mode of action and not a functional endpoint and this needs to be considered carefully in the problem formulation stage and the interpretation of the weight of evidence.

scholarly journals Biochemical, Ameliorative and Cytotoxic Effects of Newly Synthesized Curcumin Microemulsions: Evidence from In Vitro and In Vivo Studies

Nanomaterials ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 817
Author(s):  
Abbas Rahdar ◽  
Mohammad Reza Hajinezhad ◽  
Saman Sargazi ◽  
Maryam Zaboli ◽  
Mahmood Barani ◽  
...  
Keyword(s):  
Hepg2 Cells ◽  
Density Functional ◽  
Liver Enzymes ◽  
Elevated Serum ◽  
Ethyl Butyrate ◽  
Polymeric Chain ◽  
Prolonged Release ◽  
Advanced Therapies

Curcumin is known to exhibit antioxidant and tissue-healing properties and has recently attracted the attention of the biomedical community for potential use in advanced therapies. This work reports the formulation and characterization of oil-in-water F127 microemulsions to enhance the bioavailability of curcumin Microemulsions showed a high encapsulation efficiency and prolonged release. To investigate the interactions of curcumin with one unit of the polymeric chain of surfactant F127, ethyl butyrate, and sodium octanoate, as well as the interaction between ethyl butyrate and one unit of the F127 polymer chain, the Density Functional Theory (DFT) calculations at the M06-2X level of theory, were performed in water solution. The MTT assay was used to assess the cytotoxicity of free and encapsulated curcumin on non-malignant and malignant cell lines. Combination effects were calculated according to Chou-Talalay’s principles. Results of in vitro studies indicated that MCF7 and HepG2 cells were more sensitive to curcumin microemulsions. Moreover, a synergistic relationship was observed between curcumin microemulsions and cisplatin in all affected fractions of MCF7 and HepG2 cells (CI < 0.9). For in vivo investigation, thioacetamide-intoxicated rats received thioacetamide (100 mg/kg Sc) followed by curcumin microemulsions (30 mg/kg Ip). Thioacetamide-intoxicated rats showed elevated serum liver enzymes, blood urea nitrogen (BUN), and creatinine levels, and a significant reduction in liver superoxide dismutase (SOD) and catalase (CAT) activities (p < 0.05). Curcumin microemulsions reduced liver enzymes and serum creatinine and increased the activity of antioxidant enzymes in thioacetamide-treated rats in comparison to the untreated thioacetamide-intoxicated group. Histopathological investigations confirmed the biochemical findings. Overall, the current results showed the desirable hepatoprotective, nephroprotective, and anti-cancer effects of curcumin microemulsions.

Hepatoma Cell Cultures as In Vitro Models for the Hepatotoxicity of Xenobiotics

1988 ◽  
Vol 16 (1) ◽  
pp. 32-37
Author(s):  
Margherita Ferro ◽  
Anna Maria Bassi ◽  
Giorgio Nanni
Keyword(s):  
Cell Lines ◽  
Cell Cultures ◽  
Membrane Lipids ◽  
Hepatoma Cell ◽  
Positive Control ◽  
In Vitro Models ◽  
Low Concentrations ◽  
Formation Efficiency ◽  
Dose Dependent

Two hepatoma cell cultures were examined as in vitro models to be used in genotoxicity and cytotoxicity tests without the addition of bioactivating enzymes. The MH1C1, and HTC hepatoma lines were used in this study to establish their sensitivity to a number of xenobiotics, namely, cyclophosphamide (CP), the classical positive control in bioactivation tests; benzaldehyde (BA), a short-chain aldehyde; and 4-hydroxynonenal (HNE), a major toxic end-product of the peroxidative degradation of cell membrane lipids. As a first approach, we compared the following cytotoxicity tests: release of lactate dehydrogenase (LDH), and colony formation efficiency (CF). Colony-forming cells were exposed to the drugs according to different procedures, before or after the anchorage phase. The leakage of LDH into the medium following exposure of both cell lines to HNE, CP and BA for up to 24 hours was found not to be a good index of cytotoxicity. A better indicator of cytotoxicity was CF, as evaluated by exposure of the cells 24 hours after seeding. The effects were detectable at very low concentrations, corresponding to 10, 90 and 100μM for HNE, CP and BA, respectively. The impairment of CF efficiency was dose-dependent and time-dependent, and several differences between the two cell lines were observed.

scholarly journals The CAD risk locus 9p21 increases the risk of vascular calcification in an iPSC-derived VSMC model

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Anja Trillhaase ◽  
Beatrice Schmidt ◽  
Marlon Märtens ◽  
Undine Haferkamp ◽  
Jeanette Erdmann ◽  
...  
Keyword(s):  
Coronary Artery ◽  
Vascular Calcification ◽  
Alizarin Red ◽  
Vitro Assay ◽  
Human Ipscs ◽  
Increased Risk ◽  
Risk Locus ◽  
9P21 Locus ◽  
Cad Risk

Abstract Background Coronary artery disease (CAD) is the leading cause of death worldwide. Chromosome locus 9p21 was the first to be associated with increased risk of CAD and coronary artery calcification (CAC). Vascular calcification increases the risk for CAD. Vascular smooth muscle cells (VSMCs) are one of the major cell types involved in the development of vascular calcification. Methods So far, mainly animal models or primary SMCs have been used to model human vascular calcification. In this study, a human in vitro assay using iPSC-derived VSMCs was developed to examine vascular calcification. Human iPSCs were derived from a healthy non-risk (NR) and risk (R) donor carrying SNPs in the 9p21 locus. Additionally, 9p21 locus knockouts of each donor iPSC line (NR and R) were used. Following differentiation, the iPSC-derived VSMCs were characterized based on cell type, proliferation, and migration rate, along with calcium phosphate (CaP) deposits. CaP deposits were confirmed using Calcein and Alizarin Red S staining and then quantified. Results The data demonstrated significantly more proliferation, migration, and CaP deposition in VSMCs derived from the R and both KO iPSC lines than in those derived from the NR line. Molecular analyses confirmed upregulation of calcification markers. These results are consistent with recent data demonstrating increased calcification when the 9p21 murine ortholog is knocked-out. Conclusion Therefore, in conclusion, genetic variation or deletion of the CAD risk locus leads to an increased risk of vascular calcification. This in vitro human iPSC model of calcification could be used to develop new drug screening strategies to combat CAC.

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