scholarly journals Effect of Lignocaine Concentration on Human Fibroblasts Growth in Eyes Undergoing Trabeculectomy: An in vitro Study

2018 ◽  
Vol 3 (3) ◽  
pp. 1-10 ◽  
Author(s):  
Madhuravasal Krishnan Janani ◽  
Venkatakrishnan Jaichandran ◽  
Hajib Narahari Rao Madhavan ◽  
Lingam Vijaya ◽  
Ronnie Jacob George ◽  
...  
Keyword(s):  
Culture Medium ◽  
In Vitro Model ◽  
Morphological Changes ◽  
Human Fibroblasts ◽  
Annexin V ◽  
In Vitro Study ◽  
Tenon’S Capsule ◽  
Tenon's Capsule ◽  
Vitro Model

Purpose: To evaluate the effect of lignocaine on growth and apoptosis indication of primary human Tenon’s capsule fibroblast (HTFs) in an in vitro model. Patients and Methods: Tenon’s capsule tissue obtained from patients undergoing trabeculectomy were grown in cell culture medium. The effect of different concentrations of lignocaine (0.5, 1.0, 1.5, and 2%) on the morphology and growth of the fibroblasts was studied using microscopy, cell viability, and proliferation assay, and apoptosis was detected using the FITC Annexin V Apoptosis Kit. Results: Morphological changes similar to those of apoptotic cells, including cytoplasmic vacuolation, shrinkage, and rounding were visualized in the cells treated with concentrations greater than 1.0% (i.e., 1.5, 2.0%). Though proliferation inhibition was found with all four concentrations (0.5–2.0%), the viability of cells decreased from 1.0% lignocaine. Conclusion: 0.5% lignocaine prevents proliferation of fibroblasts without causing apoptosis in vitro.

Biochemical Characterisation of an In Vitro Model of Hepatocellular Apoptotic Cell Death

2009 ◽  
Vol 37 (2) ◽  
pp. 209-218 ◽  
Author(s):  
Mathieu Vinken ◽  
Elke Decrock ◽  
Elke De Vuyst ◽  
Luc Leybaert ◽  
Tamara Vanhaecke ◽  
...  
Keyword(s):  
Cell Death ◽  
In Vitro Model ◽  
Caspase 3 ◽  
Fas Ligand ◽  
Apoptotic Cell ◽  
Annexin V ◽  
Apoptotic Cell Death ◽  
Biochemical Characterisation ◽  
Vitro Model

This study was set up to critically evaluate a commonly-used in vitro model of hepatocellular apoptotic cell death, in which freshly isolated hepatocytes, cultured in a monolayer configuration, are exposed to a combination of Fas ligand and cycloheximide for six hours. A set of well-acknowledged cell death markers was addressed: a) cell morphology was studied by light microscopy; b) apoptotic and necrotic cell populations were quantified by in situ staining with Annexin-V, Hoechst 33342 and propidium iodide (PI); c) apoptotic and necrotic activities were monitored by probing caspase 3-like activity and measuring the extracellular leakage of lactate dehydrogenase (LDH), respectively; and d) the expression of apoptosis regulators was investigated by immunoblotting. The initiation of apoptosis was evidenced by the activation of caspase 8 and caspase 9, and increased Annexin-V reactivity. Progression through the apoptotic process was confirmed by the activation of caspase 3 and Bid, the enhanced expression of Bax, and the occurrence of nuclear fragmentation. Late transition to a necrotic appearance was demonstrated by an increased number of PI-positive cells and augmented extracellular release of LDH. Thus, the in vitro model allows the study of the entire course of Fas-mediated hepatocellular apoptotic cell death, which is not possible in vivo. This experimental system can serve a broad range of in vitro pharmaco-toxicological purposes, thereby directly assisting in the reduction of animal experimentation.

Studies in Cranial Suture Biology: In Vitro Cranial Suture Fusion

1996 ◽  
Vol 33 (2) ◽  
pp. 150-156 ◽  
Author(s):  
James P. Bradley ◽  
Jamie P. Levine ◽  
Christopher Blewett ◽  
Thomas Krummel ◽  
Joseph G. Mccarthy ◽  
...  
Keyword(s):  
Organ Culture ◽  
In Vitro Model ◽  
In Vitro Study ◽  
Cranial Base ◽  
Cranial Suture ◽  
Model Study ◽  
Suture Fusion ◽  
Vitro Model

The biology underlying craniosynostosis remains unknown. Previous studies have shown that the underlying dura mater, not the suture itself, signals a suture to fuse. The purpose of this study was to develop an in vitro model for cranial-suture fusion that would still allow for suture-dura interaction, but without the influence of tensional forces transmitted from the cranial base. This was accomplished by demonstrating that the posterior frontal mouse cranial suture, known to be the only cranial suture that fuses in vivo, fuses when plated with its dura in an organ-culture system. In such an organ-culture system, the sutures are free from both the influence of dural forces transmitted from the cranial base and from hormonal influences only available in a perfused system. For the cranial-suture fusion in vitro model study, the sagittal sutures (controls that remain patent in vivo) and posterior frontal sutures (that fuse in vivo) with the underlying dura were excised from 24-day-old euthanized mice, cut into 5 × 4 × 2-mm specimens, and cultured in a chemically defined, serum-free media. One hundred sutures were harvested at the day of sacrifice, then every 2 days thereafter until 30 days in culture, stained with H & E, and analyzed. A subsequent cranial-suture without dura in vitro study was performed in a similar fashion to the first study, but only the calvariae with the posterior frontal or sagittal sutures (without the underlying dura) were cultured. Results from the cranial-suture fusion in vitro model study showed that all sagittal sutures placed in organ culture with the underlying dura remained patent. More importantly, the posterior frontal sutures with the underlying dura, which were plated-down as patent at 24 days of age, demonstrated fusion after various growth periods in organ culture. In vitro posterior frontal mouse-suture fusion occurred in an anterior-to-posterior direction but in a delayed fashion, 4 to 7 days later than in vivo posterior frontal mouse-suture fusion. In contrast, the subsequent cranial-suture without dura in vitro study showed patency of all sutures, including the posterior frontal suture. These data from in vitro experiments indicate that: (1) mouse calvariae, sutures, and the underlying dura survive and grow in organ-culture systems for 30 days; (2) the local dura, free from external influences transmitted from the cranial base and hormones from distant sites, influences the cells of its overlying suture to cause fusion; and (3) without dura influence, all in vitro cranial sutures remained patent. By first identifying the factors involved in dural-suture signaling and then regulating these factors and their receptors, the biologic basis of suture fusion and craniosynostosis may be unraveled and used in the future to manipulate pathologic (premature) suture fusion.

Effect of Different EndoAnchor Configurations on Aortic Endograft Displacement Resistance: An Experimental Study

2019 ◽  
Vol 26 (5) ◽  
pp. 704-713 ◽  
Author(s):  
Seline R. Goudeketting ◽  
Jenske J. M. Vermeulen ◽  
Kim van Noort ◽  
Gerben te Riet o. g. Scholten ◽  
Henny Kuipers ◽  
...  
Keyword(s):  
In Vitro Model ◽  
Force Measurement ◽  
Current Model ◽  
In Vitro Study ◽  
Vascular Surgeon ◽  
Circumferential Distribution ◽  
Exact Moment ◽  
Displacement Force ◽  
Vitro Model

Purpose: This study investigated the effect of different EndoAnchor configurations on aortic endograft displacement resistance in an in vitro model. Materials and Methods: An in vitro model was developed and validated to perform displacement force measurements on different EndoAnchor configurations within an endograft and silicone tube. Five EndoAnchor configurations were created: (1) 6 circumferentially deployed EndoAnchors, (2) 5 EndoAnchors within 120° of the circumference and 1 additional, contralateral EndoAnchor, (3) 4 circumferentially deployed EndoAnchors, (4) 2 rows of 4 circumferentially deployed EndoAnchors, and (5) a configuration of 2 columns of 3 EndoAnchors. An experienced vascular surgeon deployed EndoAnchors under C-arm guidance at the proximal sealing zone of the endograft. A constant force with increments of 1 newton (N) was applied to the distal end of the endograft. The force necessary to displace a part of the endograft by 3 mm was defined as the endograft displacement force (EDF). Two video cameras recorded the measurements. Videos were examined to determine the exact moment 3-mm migration had occurred at part of the endograft. Five measurements were performed after each deployed EndoAnchor for each configuration. Measurements are given as the median and interquartile range (IQR) Q1, Q3. Results: Baseline displacement force measurement of the endograft without EndoAnchors resulted in a median EDF of 5.1 N (IQR 4.8, 5.2). The circumferential distribution of 6 EndoAnchors resulted in a median EDF of 53.7 N (IQR 49.0, 59.0), whereas configurations 2 through 5 demonstrated substantially lower EDFs of 29.0 N (IQR 28.5, 30.1), 24.6 N (IQR 21.9, 27.2), 36.7 N, and 9.6 N (IQR 9.4, 10.0), respectively. Decreasing the distance between the EndoAnchors over the circumference of the endograft increased the displacement resistance. Conclusion: This in vitro study demonstrates the influence EndoAnchor configurations have on the displacement resistance of an aortic endograft. Parts of the endograft where no EndoAnchor has been deployed remain sensitive to migration. In the current model, the only configuration that rivaled a hand-sewn anastomosis was the one with 6 EndoAnchors. A circumferential distribution of EndoAnchors with small distances between EndoAnchors should be pursued, if possible. This study provides a quantification of different EndoAnchor configurations that clinicians may have to adopt in clinical practice, which can help them make a measured decision on where to deploy EndoAnchors to ensure good endograft fixation.

Gelatin sponge insertion into tympanostomy tubes: An in-vitro study to evaluate postoperative obstruction

2009 ◽  
Vol 140 (5) ◽  
pp. 661-664
Author(s):  
Chad P. Secor ◽  
Robert E. Wilson ◽  
Paul M. Spring ◽  
Richard C. Haydon
Keyword(s):  
In Vitro Model ◽  
In Vitro Study ◽  
The Other ◽  
Gelatin Sponge ◽  
Pressure Equalization ◽  
Significant Difference ◽  
Tympanostomy Tubes ◽  
Ventilation Tubes ◽  
Vitro Model

Objective: To analyze the efficacy of gelatin sponge insertion into lumens of tympanostomy tubes to prevent obstruction in the presence of blood. Study Design: In vitro model. Methods: Absorbable gelatin sponge wicks were placed in the lumen of Ultrasil Collar Button ventilation tubes and Shepherd Grommet ventilation tubes. One half of each group was covered with blood, the other left untreated. Each tube was treated with ofloxacin solution three times daily for seven days. After treatment, the tubes were inspected. Reinspection was performed after brief suctioning. Numerical scores were given based on degree of obstruction. Results: A statistically significant difference in degree of obstruction ( P < 0.0001) was seen between all tubes with wicks alone versus those with blood added. After re-evaluation, there remained a statistically significant difference between tubes with wicks alone and tubes with wicks and blood ( P < 0.0001). Conclusions: Gelatin sponge insertion does not prevent, and may in fact, enhance, obstruction of pressure equalization tube lumens in the presence of blood.

An In Vivo/In Vitro Study of Allyl Alcohol Toxicity Using Enzyme Inhibitors

1993 ◽  
Vol 21 (1) ◽  
pp. 38-42
Author(s):  
Romana Pulci ◽  
Donatella Moneta ◽  
Philippe Dostert ◽  
Marco Brughera ◽  
Giovanna Scampini ◽  
...  
Keyword(s):  
Aldehyde Dehydrogenase ◽  
Allyl Alcohol ◽  
In Vitro Model ◽  
Enzyme Inhibitors ◽  
In Vitro Study ◽  
Rat Hepatocytes ◽  
Isolated Rat Hepatocytes ◽  
Vitro Model

The aim of this study was to verify an in vitro model of hepatotoxicity, designed to assess the production of reactive species from biologically-inert chemicals through their metabolic transformation. One example is allyl alcohol, which produces acrolein through the action of the enzyme alcohol dehydrogenase. Acrolein is a highly hepatotoxic aldehyde which is detoxified to acrylic acid by aldehyde dehydrogenase (ALDH). A deficiency of this enzyme, common in some Asian populations, can give rise to pathological conditions of hepatotoxicity. Isolated rat hepatocytes were incubated with allyl alcohol with and without cyanamide, a known inhibitor of ALDH. The toxicity of allyl alcohol, assessed on the basis of release of glutamic-oxaloacetic transaminase (GOT), glutamic-pyruvic transaminase (GPT) and lactate dehydrogenase (LDH) into the culture medium, was dramatically increased by the addition of cyanamide. In vivo, the same treatment scheme was used in rats treated with allyl alcohol with or without cyanamide pretreatment. It was also demonstrated that allyl alcohol toxicity is dramatically enhanced by the addition of an aldehyde dehydrogenase inhibitor, as shown by plasma levels of hepatic enzymes (GOT, GPT and LDH) and by histological findings. We believe that this in vitro model, involving the use of enzyme inhibitors, could be useful for verification of the hypothesis that hepatotoxins, such as acrolein, are produced from some pharmaceutical and other chemical compounds.

scholarly journals Pentagamavunon-0 (PGV-0) Enhance Cytotoxic Effect of Doxorubicin through Increasing of Apoptosis, Senescence and ROS Level on Triple Negative Breast Cancer 4T1

2020 ◽  
Vol 11 (1) ◽  
pp. 7
Author(s):  
Ismanurrahman Hadi ◽  
Riris Istighfari Jenie ◽  
Edy Meiyanto
Keyword(s):  
Combination Treatment ◽  
In Vitro Model ◽  
Annexin V ◽  
Positive Control ◽  
Breast Cancers ◽  
Single Treatment ◽  
Ic50 Values ◽  
Vitro Model ◽  
Better Than

TNBC, one of the sub type of breast cancers was widely known with high tumorigenic and poor prognosis than others. The development of combination agent (co-chemotherapy) with doxorubicin for chemotherapy of TNBC were carried out to decrease doxorubicin side effect and resistance in cancer. This present study aims to explore the co-chemotherapeutic properties of PGV-0 and investigate induction of doxorubicin on apoptosis, senescence and ROS against TNBC. 4T1 Cell line were used as a TNBC in vitro model. Cytotoxic measurement was performed using MTT assay resulting in IC50 values of 52 μM. Meanwhile, the combination of doxorubicin and PGV-0 showed synergistic effect which decreased cell viability of 4T1 better than single treatment of doxorubicin. Apoptosis analysis was performed using annexin V/PI assay indicated that the combination treatment of PGV-0 and doxorubicin increased apoptosis evidence. Senescence detection was carried out using senescence-associated-β galactosidase (SA-β-gal) assay. The results showed that a single treatment of PGV-0 induced cellular senescence and increased senescence cells in combination treatment. Moreover, DCFDA staining showed that PGV-0 increased ROS level at single treatment, whereas combination treatment increased ROS intracellular compared to the positive control of doxorubicin. Based on these results, PGV-0 has potential as a co-chemotherapeutic candidate on TNBC.Keyword: 4T1, PGV-0, Co-chemotherapy, Cytotoxic, Senescence, Apoptosis, ROS

Peptidergic properties expressed in vitro by embryonic neuroblasts after neural induction

Development ◽  
1989 ◽  
Vol 105 (3) ◽  
pp. 529-540
Author(s):  
F. Pituello ◽  
P. Deruntz ◽  
L. Pradayrol ◽  
A.M. Duprat
Keyword(s):  
Culture Medium ◽  
In Vitro Model ◽  
Neural Induction ◽  
Defined Medium ◽  
Neuronal Precursor Cells ◽  
Cell Extracts ◽  
Neuronal Precursors ◽  
Immunocytochemical Techniques ◽  
Vitro Model

As an immediate consequence of neural induction, some neuroectodermal cells acquire the ability to develop a number of characteristic neuronal features, without requiring any subsequent embryonic cues (Duprat et al. 1987). Thus, adrenergic, cholinergic and gabaergic traits are expressed in cultures of neural fold and neural plate isolated from amphibian embryos immediately after induction and grown in a defined medium. The aim of the present study was to determine, using the same in vitro model, their abilities to develop peptidergic phenotypes. Using immunocytochemical techniques, we show that substance P-, enkephalin- (leu-enkephalin, metenkephalin), and somatostatin- like immunoreactivities are expressed in subpopulations of neurones grown in vitro, whereas VIP (vasoactive intestinal polypeptide) is not detected under the same conditions. The appearance and development of the somatostatinergic phenotype has been quantified by RIA both in cell extracts and in the culture medium. Somatostatin-like immunoreactivity (SLI) undetectable at the late gastrula stage, can be measured in cells after 4 days of culture and continues to increase over the next 10 days. In culture medium, SLI is present at a constant level from day 4 up to day 14. These data reveal that some neuronal precursor cells acquire, during neural induction, the potentiality to biosynthesize, store and release neuropeptides. Furthermore, the expression of these peptidergic phenotypes in distinct subpopulations of neurones suggests that certain neuronal precursors become committed to different metabolic pathways at the earliest steps of neurogenesis.

Syringomyelia Hydrodynamics: An In Vitro Study Based on In Vivo Measurements

2005 ◽  
Vol 127 (7) ◽  
pp. 1110-1120 ◽  
Author(s):  
Bryn A. Martin ◽  
Wojciech Kalata ◽  
Francis Loth ◽  
Thomas J. Royston ◽  
John N. Oshinski
Keyword(s):  
Spinal Cord ◽  
Wall Motion ◽  
In Vitro Model ◽  
Fluid Velocity ◽  
In Vitro Study ◽  
Elastic Tube ◽  
Flow Waveform ◽  
Vitro Model

A simplified in vitro model of the spinal canal, based on in vivo magnetic resonance imaging, was used to examine the hydrodynamics of the human spinal cord and subarachnoid space with syringomyelia. In vivo magnetic resonance imaging (MRI) measurements of subarachnoid (SAS) geometry and cerebrospinal fluid velocity were acquired in a patient with syringomyelia and used to aid in the in vitro model design and experiment. The in vitro model contained a fluid-filled coaxial elastic tube to represent a syrinx. A computer controlled pulsatile pump was used to subject the in vitro model to a CSF flow waveform representative of that measured in vivo. Fluid velocity was measured at three axial locations within the in vitro model using the same MRI scanner as the patient study. Pressure and syrinx wall motion measurements were conducted external to the MR scanner using the same model and flow input. Transducers measured unsteady pressure both in the SAS and intra-syrinx at four axial locations in the model. A laser Doppler vibrometer recorded the syrinx wall motion at 18 axial locations and three polar positions. Results indicated that the peak-to-peak amplitude of the SAS flow waveform in vivo was approximately tenfold that of the syrinx and in phase (SAS∼5.2±0.6ml∕s,syrinx∼0.5±0.3ml∕s). The in vitro flow waveform approximated the in vivo peak-to-peak magnitude (SAS∼4.6±0.2ml∕s,syrinx∼0.4±0.3ml∕s). Peak-to-peak in vitro pressure variation in both the SAS and syrinx was approximately 6 mm Hg. Syrinx pressure waveform lead the SAS pressure waveform by approximately 40 ms. Syrinx pressure was found to be less than the SAS for ∼200ms during the 860-ms flow cycle. Unsteady pulse wave velocity in the syrinx was computed to be a maximum of ∼25m∕s. LDV measurements indicated that spinal cord wall motion was nonaxisymmetric with a maximum displacement of ∼140μm, which is below the resolution limit of MRI. Agreement between in vivo and in vitro MR measurements demonstrates that the hydrodynamics in the fluid filled coaxial elastic tube system are similar to those present in a single patient with syringomyelia. The presented in vitro study of spinal cord wall motion, and complex unsteady pressure and flow environment within the syrinx and SAS, provides insight into the complex biomechanical forces present in syringomyelia.

scholarly journals In vitro study after exposure to the aqueous extract of Piper amalago L. shows changes of morphology, proliferation, cytoskeleton and molecules of the extracellular matrix

2021 ◽  
Vol 10 (4) ◽  
pp. e0110413289
Author(s):  
Vera Lucia Pereira dos Santos ◽  
Célia Regina Cavichiolo Franco ◽  
Ricardo Wagner ◽  
Caroline Dadalt Silva ◽  
Célia Regina Cavichiolo Franco ◽  
...  
Keyword(s):  
Amino Acid ◽  
Cell Viability ◽  
Cell Body ◽  
In Vitro Model ◽  
In Vitro Study ◽  
Healing Agent ◽  
Vitro Model ◽  
Insect Bites ◽  
Number Of Cells

Piper amalago L. is a medicinal plant traditionally used as a healing agent for wounds, burns, abscesses, boils, and insect bites. The current study aimed to evaluate the possible effects of the aqueous crude extract obtained from P. amalago leaves, in different concentrations and in different incubation times, using the in vitro model of mouse fibroblasts (3T3). The extract was tested in different concentrations at the 24 h incubation time for analysis of cell viability, cytotoxicity, proliferation, cell morphology, immunostaining, adhesion and cell spreading assays, as well as to determine the hydroxyproline concentration and activity of the metalloproteinase MMP2. Morphologically, after exposure to the concentrations of 15 and 150 µg/mL, the cells maintained the morphology, yet a greater number of cells with more expansions of the cell body and larger than the control cells were observed. The treated cell culture also showed a greater number of cells, larger cells, a greater expansion of the cell body, adherent cells spread over the substrate, and a more juxtaposed, central and spherical nucleus. The treatment induced greater cell adhesion to the polymer, fibronectin, and collagen I. Biochemical results showed a significant increase in the hydroxyproline amino acid after exposure for 96 h. The extract did not induce loss of cell viability until the concentration reached 150 µg/mL, positively modulating proliferation, morphology, adhesion, degree of spreading, and organization of microfilaments. The extract also promoted a significant increase in the hydroxyproline amino acid.

scholarly journals From Blood to Lesioned Brain: An In Vitro Study on Migration Mechanisms of Human Nasal Olfactory Stem Cells

2017 ◽  
Vol 2017 ◽  
pp. 1-17 ◽  
Author(s):  
Stéphane D. Girard ◽  
Isabelle Virard ◽  
Emmanuelle Lacassagne ◽  
Jean-Michel Paumier ◽  
Hanae Lahlou ◽  
...  
Keyword(s):  
Stem Cells ◽  
Genomic Dna ◽  
Dna Microarrays ◽  
In Vitro Model ◽  
In Vitro Study ◽  
Cognate Receptor ◽  
Mouse Hippocampus ◽  
Vitro Model ◽  
Chemotactic Effect

Stem cell-based therapies critically rely on selective cell migration toward pathological or injured areas. We previously demonstrated that human olfactory ectomesenchymal stem cells (OE-MSCs), derived from an adult olfactory lamina propria, migrate specifically toward an injured mouse hippocampus after transplantation in the cerebrospinal fluid and promote functional recoveries. However, the mechanisms controlling their recruitment and homing remain elusive. Using an in vitro model of blood-brain barrier (BBB) and secretome analysis, we observed that OE-MSCs produce numerous proteins allowing them to cross the endothelial wall. Then, pan-genomic DNA microarrays identified signaling molecules that lesioned mouse hippocampus overexpressed. Among the most upregulated cytokines, both recombinant SPP1/osteopontin and CCL2/MCP-1 stimulate OE-MSC migration whereas only CCL2 exerts a chemotactic effect. Additionally, OE-MSCs express SPP1 receptors but not the CCL2 cognate receptor, suggesting a CCR2-independent pathway through other CCR receptors. These results confirm that OE-MSCs can be attracted by chemotactic cytokines overexpressed in inflamed areas and demonstrate that CCL2 is an important factor that could promote OE-MSC engraftment, suggesting improvement for future clinical trials.

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