In Vitro Tests for Detecting Chemicals Affecting the Embryo Implantation Process

Comprehensive understanding of the regulatory mechanism of the implantation process in pigs is crucial for reproductive success. The endometrium plays an important role in regulating the establishment and maintenance of gestation. The goal of the current study was to determine the effect of adiponectin on the global expression pattern of genes and relationships among differentially expressed genes (DE-genes) in the porcine endometrium during implantation using microarrays. Diverse transcriptome analyses including gene ontology (GO), biological pathway, networks, and DE-gene analyses were performed. Adiponectin altered the expression of 1286 genes with fold-change (FC) values greater than 1.2 (p < 0.05). The expression of 560 genes were upregulated and 726 downregulated in the endometrium treated with adiponectin. Thirteen genes were selected for real-time PCR validation of differential expression based on a known role in metabolism, steroid and prostaglandin synthesis, interleukin and growth factor action, and embryo implantation. Functional analysis of the relationship between DE-genes indicated that adiponectin interacts with genes that are involved in the processes of cell proliferation, programmed cell death, steroid and prostaglandin synthesis/metabolism, cytokine production, and cell adhesion that are critical for reproductive success. The presented results suggest that adiponectin signalling may play a key role in the implantation of pig.

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P–403 Sodium tungstate increases embryo adhesion through a direct effect on endometrial cells

Human Reproduction ◽

10.1093/humrep/deab130.402 ◽

2021 ◽

Vol 36 (Supplement_1) ◽

Author(s):

I Canals ◽

D Cotán ◽

R Torres ◽

J A Horcajadas ◽

A Arbat

Keyword(s):

Direct Effect ◽

Sodium Tungstate ◽

Embryo Implantation ◽

In Vivo Models ◽

Endometrial Cells ◽

A Cells ◽

Adhesion Rate ◽

Implantation Process

Abstract Study question Does sodium tungstate treatment induce a change in endometrial cells’ capacity to implant trophoblasts? Summary answer Administration of sodium tungstate to endometrial cells increases trophoblast adhesion. What is known already Sodium tungstate (ST) has shown its capacity to modulate the activity of cytokines, such as leptin, an activator of an obligatory signalling cascade in the embryo-implantation process. STAT3, a signal transducer molecule critical for the embryo implantation process, is also known to be activated by ST. Still, ST’s effect on implantation using biological systems has never been studied. Embryo implantation process and endometrium roles are complicated to study in vivo due to a lack of animal models and appropriate techniques. In vitro techniques using immortalised cell lines allows a first approach to study early implantation stages, such as embryo adhesion. Study design, size, duration An in vitro study was carried out using a human endometrial carcinoma cell line (HEC–1-A) treated with sodium tungstate for 24 and 48h, and choriocarcinoma cell spheroids (JAr). Different times of treatment and concentrations were studied. Each experiment was performed in triplicate. Participants/materials, setting, methods Confluent endometrial HEC–1-A cultures were treated with ST at concentrations (0–150mM) and withaferin A (1mM), negative control for embryo adhesion. After the treatment period, HEC–1-A cultures were washed with ST-free culture medium to eliminate ST. Immediately, 15 JAr trophoblast spheroids were added to cultures and coincubated with gentle agitation for 30, 60 and 90 minutes. An inverted light microscope was used to count adhered and floating spheroids, and determine the trophoblast adherence ratio. Main results and the role of chance HEC–1-A cells treated with ST showed normal morphology and growth at all doses except 150mM. At the highest dose tested, the cells’ culture was still viable (negative blue trypan staining) and maintained morphology, but the adhesion to the plate surface was affected. Doses from 0.15 to 15mM were used to perform adhesion assays. HEC–1-A cells treated with ST for 24h showed an increased capacity to adhere JAr trophoblast spheroids. Adhesion rates reached significant differences at doses of 1.5 and 15mM after 60 and 90 minutes of coincubation. After 90 minutes, untreated cells reached 32.8% adhesion rate, while 1.5 and 15mM ST-treated cells reached 54.6% and 53.4% respectively (p < 0.05 ST vs untreated). Thus, the increment of trophoblast adhesion rate induced by ST reached 66%. Lower adhesion rates were observed after 60 minutes of coincubation but were also significant with a relative increase of 49.1% at 1.5mM and 50.5% at 1.5mM when compared with untreated cells (p < 0.05) Longer treatments (48h) showed similar trends to 24h-treatments, but with a lower extent of ST effect on HEC–1-A receptivity. Maximum adhesion rates were also observed at 90 minutes of coincubation and 1.5 and 15mM doses. The Mean adhesion rate increase was >40% with both doses. Limitations, reasons for caution: The current study is the first approach to evaluate sodium tungstate effect on endometrium using an in vitro model. Future research using in vivo models should be performed to assess sodium tungstate effect on endometrium receptivity and its potential as a fertility treatment. Wider implications of the findings: We conclude that the direct effect of sodium tungstate on endometrial cells increases embryo adhesion rate. These results open a new research line to a potential treatment in human reproduction management with sodium tungstate to solve the unmet need of inducing embryo implantation. Trial registration number Not applicable

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Anti-Diabetic Activity of a Mineraloid Isolate, in vitro and in Genetically Diabetic Mice

International Journal for Vitamin and Nutrition Research ◽

10.1024/0300-9831/a000048 ◽

2011 ◽

Vol 81 (1) ◽

pp. 34-42 ◽

Cited By ~ 1

Author(s):

Joel Deneau ◽

Taufeeq Ahmed ◽

Roger Blotsky ◽

Krzysztof Bojanowski

Keyword(s):

Dna Microarrays ◽

Human Fibroblasts ◽

Diabetic Animal ◽

In Vitro Tests ◽

Diabetic Mice ◽

Fossil Material ◽

Expression Of Genes ◽

Enhanced Expression ◽

Genetically Diabetic Mice

Type II diabetes is a metabolic disease mediated through multiple molecular pathways. Here, we report anti-diabetic effect of a standardized isolate from a fossil material - a mineraloid leonardite - in in vitro tests and in genetically diabetic mice. The mineraloid isolate stimulated mitochondrial metabolism in human fibroblasts and this stimulation correlated with enhanced expression of genes coding for mitochondrial proteins such as ATP synthases and ribosomal protein precursors, as measured by DNA microarrays. In the diabetic animal model, consumption of the Totala isolate resulted in decreased weight gain, blood glucose, and glycated hemoglobin. To our best knowledge, this is the first description ever of a fossil material having anti-diabetic activity in pre-clinical models.

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Comparison of High Purity Factor IX Concentrates and a Prothrombin Complex Concentrate in a Canine Model of Thrombogenicity

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646468 ◽

1991 ◽

Vol 66 (05) ◽

pp. 609-613 ◽

Cited By ~ 14

Author(s):

I R MacGregor ◽

J M Ferguson ◽

L F McLaughlin ◽

T Burnouf ◽

C V Prowse

Keyword(s):

High Purity ◽

Time Course ◽

Prothrombin Complex Concentrate ◽

Degradation Products ◽

Canine Model ◽

Factor Ix ◽

In Vitro Tests ◽

Albumin Infusion

SummaryA non-stasis canine model of thrombogenicity has been used to evaluate batches of high purity factor IX concentrates from 4 manufacturers and a conventional prothrombin complex concentrate (PCC). Platelets, activated partial thromboplastin time (APTT), fibrinogen, fibrin(ogen) degradation products and fibrinopeptide A (FPA) were monitored before and after infusion of concentrate. Changes in FPA were found to be the most sensitive and reproducible indicator of thrombogenicity after infusion of batches of the PCC at doses of between 60 and 180 IU/kg, with a dose related delayed increase in FPA occurring. Total FPA generated after 100-120 IU/kg of 3 batches of PCC over the 3 h time course was 9-12 times that generated after albumin infusion. In contrast the amounts of FPA generated after 200 IU/kg of the 4 high purity factor IX products were in all cases similar to albumin infusion. It was noted that some batches of high purity concentrates had short NAPTTs indicating that current in vitro tests for potential thrombogenicity may be misleading in predicting the effects of these concentrates in vivo.

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A Comparison of the in Vitro and in Vivo Thrombogenic Activity of Factor IX Concentrates Using Stasis (Wessler) and Non-Stasis Rabbit Models

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650089 ◽

1980 ◽

Vol 44 (02) ◽

pp. 081-086 ◽

Cited By ~ 8

Author(s):

C V Prowse ◽

A E Williams

Keyword(s):

Platelet Rich Plasma ◽

Thrombus Formation ◽

Factor Ix ◽

Coagulation System ◽

In Vitro Tests ◽

Clinical Use ◽

Low Activity

SummaryThe thrombogenic effects of selected factor IX concentrates were evaluated in two rabbit models; the Wessler stasis model and a novel non-stasis model. Concentrates active in either the NAPTT or TGt50 in vitro tests of potential thrombogenicity, or both, caused thrombus formation in the Wessler technique and activation of the coagulation system in the non-stasis model. A concentrate with low activity in both in vitro tests did not have thrombogenic effects in vivo, at the chosen dose. Results in the non-stasis model suggested that the thrombogenic effects of factor IX concentrates may occur by at least two mechanisms. A concentrate prepared from platelet-rich plasma and a pyrogenic concentrate were also tested and found to have no thrombogenic effect in vivo.These studies justify the use of the NAPTT and TGt50 in vitro tests for the screening of factor IX concentrates prior to clinical use.

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In Vitro Thrombogenicity Tests of Factor IX Concentrates

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657035 ◽

1979 ◽

Vol 42 (05) ◽

pp. 1355-1367 ◽

Cited By ~ 8

Author(s):

C V Prowse ◽

A Chirnside ◽

R A Elton

Keyword(s):

Factor Viii ◽

Partial Thromboplastin Time ◽

Generation Time ◽

Activated Partial Thromboplastin Time ◽

Factor Ix ◽

In Vitro Tests ◽

Factor Viii Inhibitor ◽

Factor Ixa ◽

Viii Inhibitor

SummaryVarious factor IX concentrates have been examined in a number of in vitro tests of thrombogenicity. The results suggest that some tests are superfluous as in concentrates with activity in any of these tests activation is revealed by a combination of the non-activated partial thromboplastin time, the thrombin (or Xa) generation time and factor VIII inhibitor bypassing activity tests. Assay of individual coagulant enzymes revealed that most concentrates contained more factor IXa than Xa. However only a small number of concentrates, chiefly those that had been purposefully activated, contained appreciable amounts of either enzyme.

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Inhibition of Fibrinolysis and Fibrinogenolysis in Man: Comparison of ε-Aminocaproic Acid and Kallikrein Inhibitor

Thrombosis and Haemostasis ◽

10.1055/s-0038-1660337 ◽

1963 ◽

Vol 10 (01) ◽

pp. 106-119 ◽

Cited By ~ 8

Author(s):

E Beck ◽

R Schmutzler ◽

F Duckert ◽

Keyword(s):

Aminocaproic Acid ◽

Side Reactions ◽

In Vitro Tests ◽

Factor V ◽

Clinical Use ◽

Strong Inhibitor ◽

Kallikrein Inhibitor ◽

Therapeutic Possibilities

SummaryInhibitor of kallikrein and trypsin (KI) extracted from bovine parotis was compared with ε-aminocaproic acid (EACA): both substances inhibit fibrinolysis induced with streptokinase. EACA is a strong inhibitor of fibrinolysis in concentrations higher than 0, 1 mg per ml plasma. The same amount and higher concentrations are not able to inhibit completely the proteolytic-side reactions of fibrinolysis (fibrinogenolysis, diminution of factor V, rise of fibrin-polymerization-inhibitors). KI inhibits well proteolysis of plasma components in concentrations higher than 2,5 units per ml plasma. Much higher amounts of KI are needed to inhibit fibrinolysis as demonstrated by our in vivo and in vitro tests.Combination of the two substances for clinical use is suggested. Therapeutic possibilities are discussed.

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Fruit Volatiles to Control Postharvest Rot of Stone Fruits and Pears

HortScience ◽

10.21273/hortsci.33.3.469a ◽

1998 ◽

Vol 33 (3) ◽

pp. 469a-469

Author(s):

L.J. Skog ◽

D.P. Murr ◽

B.E. Digweed

Keyword(s):

Volatile Compounds ◽

Isoamyl Acetate ◽

Penicillium Expansum ◽

Effective Control ◽

In Vitro Tests ◽

Stone Fruits ◽

In Situ Tests ◽

Highly Effective ◽

Postharvest Rot

Volatile compounds are ubiquitous in plants, giving fruits their characteristic aroma and flavor. There is increasing evidence that these compounds can protect plants from pathogenic organisms. In this trial ≈25 volatile compounds were tested for efficacy against Monilinia fructicola and Penicillium expansum. Both in vitro tests on agar plugs of actively growing pathogens and in situ tests on inoculated stone fruits and pears were conducted. The volatile compounds were grouped into three categories based upon fungicidal activity in vitro: highly effective (fungicidal concentration ≤100 M), moderately effective (fungicidal concentration between 100–200 M) and ineffective (fungicidal concentration >200 M). Highly effective compounds included: acetaldehyde, citral, 2-ethyl-1-hexanol, 2,exadienal, E-2-hexenal, 4-hexen-3-one, linalool, (E,E)2,4-nonadienal, E-2-nonenal, E-3-none-2-one, salicylaldehyde, and valeraldehyde. Moderately effective compounds included: (E,Z) 2,6-nonadienal, propionaldehyde, terpinene, butyl acetate, E-cinnamaldehde, hexanal, E-2-hexen-1-ol, Z-3-hexen-1-ol and isoamyl acetate. Ineffective compounds included: butyrolactone, ethanol, ethyl acetate, and methyl acetate. Effectiveness of the compounds varied with both strain and type of microorganism tested. Concentraions required for effective control were much higher when the compounds were tested on inoculated fruit. Phytotoxicity was a problem with some compounds.

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Role of in vivo and in vitro Tests in the Diagnosis of Severe Cutaneous Adverse Reactions (SCAR) to Drug

Current Pharmaceutical Design ◽

10.2174/1381612825666191107104126 ◽

2019 ◽

Vol 25 (36) ◽

pp. 3872-3880 ◽

Cited By ~ 3

Author(s):

Marcel M. Bergmann ◽

Jean-Christoph Caubet

Keyword(s):

Adverse Reactions ◽

Lymphocyte Transformation ◽

Stevens Johnson Syndrome ◽

In Vitro Tests ◽

High Morbidity ◽

Patch Tests ◽

Severe Cutaneous Adverse Reactions ◽

Cutaneous Adverse Reactions

Severe cutaneous adverse reactions (SCAR) are life-threatening conditions including acute generalized exanthematous pustulosis (AGEP), Stevens-Johnson Syndrome (SJS), toxic epidermal necrolysis (TEN) and drug reaction with eosinophilia and systemic symptoms (DRESS). Diagnosis of causative underlying drug hypersensitivity (DH) is mandatory due to the high morbidity and mortality upon re-exposure with the incriminated drug. If an underlying DH is suspected, in vivo test, including patch tests (PTs), delayed-reading intradermal tests (IDTs) and in vitro tests can be performed in selected patients for which the suspected culprit drug is mandatory, or in order to find a safe alternative treatment. Positivity of in vivo and in vitro tests in SCAR to drug varies depending on the type of reaction and the incriminated drugs. Due to the severe nature of these reactions, drug provocation test (DPT) is highly contraindicated in patients who experienced SCAR. Thus, sensitivity is based on positive test results in patients with a suggestive clinical history. Patch tests still remain the first-line diagnostic tests in the majority of patients with SCAR, followed, in case of negative results, by delayed-reading IDTs, with the exception of patients with bullous diseases where IDTs are still contra-indicated. In vitro tests have shown promising results in the diagnosis of SCAR to drug. Positivity is particularly high when the lymphocyte transformation test (LTT) is combined with cytokines and cytotoxic markers measurement (cyto-LTT), but this still has to be confirmed with larger studies. Due to the rarity of SCAR, large multi-center collaborative studies are needed to better study the sensitivity and specificity of in vivo and in vitro tests.

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