In Vitro Evaluation of Acellular Collagen Matrices Derived from Porcine Pericardium: Influence of the Sterilization Method on Its Biological Properties

The aim of this study were characterize acellular collagen matrices derived from porcine pericardium (PP) and to evaluate their properties after sterilization by ethylene oxide and gamma ray. PP matrices were subjected to alkaline hydrolysis (AH), and samples were characterized for biological stability, membrane thickness measurements, differential scanning calorimetry (DSC) and scanning electron microscopy (SEM). Subsequently, the matrices were frozen, lyophilized and sterilized by ethylene oxide or gamma radiation. For in vitro assays, CHO-K1 cell culture was used and evaluated for cytotoxicity, clonogenic survival assay, genotoxicity and mutagenicity. Analysis of variance (ANOVA) was used, followed by Dunnett’s post-test, with a significance level of 5%. After AH, there was no significant change in matrix thickness. The relative biodegradability of the material after implantation was observed. Morphology and dimensions had small changes after AH. As for cell viability, none of the tested matrices showed a statistically significant difference (p > 0.05; Dunnett) regardless of the sterilization method. Furthermore, it was found that PP matrices did not interfere with the proliferation capacity of CHO-K1 cells (p > 0.05; Dunnett). As for genotoxicity, when sterilized with ethylene oxide (NP, P12 and P24), it showed genotoxic potential, but it was not genotoxic when sterilized by gamma radiation. No mutagenic effects were observed in either group. PP-derived collagen matrices hydrolyzed at different times were not cytotoxic. It is concluded that the best method of sterilization is through gamma radiation, since no significant changes were observed in the properties of the PP matrices.

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Computational and In-Vitro Validation of Natural Molecules as Potential Acetylcholinesterase Inhibitors and Neuroprotective Agents

Current Alzheimer Research ◽

10.2174/1567205016666181212155147 ◽

2019 ◽

Vol 16 (2) ◽

pp. 116-127 ◽

Cited By ~ 2

Author(s):

Ashwani Kumar ◽

Vineet Mehta ◽

Utkarsh Raj ◽

Pritish Kumar Varadwaj ◽

Malairaman Udayabanu ◽

...

Keyword(s):

In Silico ◽

Hippocampal Neurons ◽

Symptomatic Relief ◽

In Vitro Assays ◽

Disease Modifying Drugs ◽

In Silico Docking ◽

Ache Inhibitors ◽

Significant Difference ◽

Disease Modifying

Background: Cholinesterase inhibitors are the first line of therapy for the management of Alzheimer’s disease (AD), however, it is now established that they provide only temporary and symptomatic relief, besides, having several inherited side-effects. Therefore, an alternative drug discovery method is used to identify new and safer ‘disease-modifying drugs’. Methods: Herein, we screened 646 small molecules of natural origin having reported pharmacological and functional values through in-silico docking studies to predict safer neuromodulatory molecules with potential to modulate acetylcholine metabolism. Further, the potential of the predicted molecules to inhibit acetylcholinesterase (AChE) activity and their ability to protect neurons from degeneration was determined through in-vitro assays. Results: Based on in-silico AChE interaction studies, we predicted quercetin, caffeine, ascorbic acid and gallic acid to be potential AChE inhibitors. We confirmed the AChE inhibitory potential of these molecules through in-vitro AChE inhibition assay and compared results with donepezil and begacestat. Herbal molecules significantly inhibited enzyme activity and inhibition for quercetin and caffeine did not show any significant difference from donepezil. Further, the tested molecules did not show any neurotoxicity against primary (E18) hippocampal neurons. We observed that quercetin and caffeine significantly improved neuronal survival and efficiently protected hippocampal neurons from HgCl2 induced neurodegeneration, which other molecules, including donepezil and begacestat, failed to do. Conclusion: Quercetin and caffeine have the potential as “disease-modifying drugs” and may find application in the management of neurological disorders such as AD.

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Effect of Titanium Implants Coated with Radiation-Crosslinked Collagen on Stability and Osseointegration in Rat Tibia

Materials ◽

10.3390/ma11122520 ◽

2018 ◽

Vol 11 (12) ◽

pp. 2520 ◽

Cited By ~ 4

Author(s):

Eun-Bin Bae ◽

Ji-Hyun Yoo ◽

Sung-In Jeong ◽

Min-Su Kim ◽

Youn-Mook Lim ◽

...

Keyword(s):

Gamma Radiation ◽

Human Mesenchymal Stem Cells ◽

Region Of Interest ◽

Bone Volume ◽

Titanium Implants ◽

Control Group ◽

Gene Expressions ◽

Significant Difference

This study aimed to evaluate the titanium (Ti) implants coated with collagen type Ⅰ crosslinked using gamma-irrigation or glutaraldehyde (GA). The in vitro surface observations, quantification assay, and cell studies using human mesenchymal stem cells (hMSCs) were conducted. For in vivo experiments, the implants were divided into three groups and inserted into the rat tibias: control group (non-treated Ti implant), GA group (Ti implants coated with GA-crosslinked collagen) and 25 kGy group (Ti implants coated with gamma-radiation-crosslinked collagen at dose of 25 kGy). The animals were sacrificed at 4 weeks after implantation and the tissue sections were obtained. New bone volume (mm3) and bone-to-implant contact (BIC, %) within the region of interest (ROI) was measured. The in vitro results showed the highest osteogenic differentiation and levels of osteogenesis-related gene expressions in the 25 kGy group without cytotoxicity. The new bone volume of GA group was significantly higher than the control (p < 0.05). In the result of the BIC, the 25 kGy group was significantly higher than the control (p < 0.05). However, there was no significant difference between the experimental groups. Within the limitations of this study, Ti implant coated with gamma-radiation-crosslinked collagen has potential utility without side effects from chemical agents.

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In vitro and in vivo toxicity of fungicides and biofungicides for the control of verticillium and fusarium wilt of pepper

Pesticidi i fitomedicina ◽

10.2298/pif2101023m ◽

2021 ◽

Vol 36 (1) ◽

pp. 23-34

Author(s):

Milica Mihajlovic ◽

Emil Rekanovic ◽

Jovana Hrustic ◽

Mila Grahovac ◽

Brankica Tanovic

Keyword(s):

Antifungal Activity ◽

Verticillium Dahliae ◽

Antagonistic Activity ◽

In Vitro Assays ◽

Thiophanate Methyl ◽

Significant Difference ◽

By Products ◽

Pepper Plants

A survey of in vitro and in vivo sensitivity of Verticillium dahliae and Fusarium oxysporum to several commercial fungicides and biofungicides was undertaken. In in vitro assays, the tested isolate of V. dahliae proved to be very sensitive to difenoconazole (EC50 = 0.02 mg/l). However, under greenhouse conditions, the highest efficacy in V. dahliae control on inoculated pepper plants was recorded for a product based on thiophanate-methyl (83.10% compared to control). Among the tested fungicides, the lowest efficacy was recorded for a product based on azoxystrobin (23.10 %) with no significant difference compared to control (p > 0.05). In in vitro assays, the tested F. oxysporum isolate was the most sensitive to prochloraz (EC50 = 0.07 mg/l) and the least sensitive to fluopyram (EC50 = 1075.01 mg/l). In in vivo assay, the highest efficacy was achieved by products based on captan (95.60%), and the lowest by a product based on thiophanate-methyl (54.40%). Antagonistic activity of the bacterium B. subtilis under laboratory conditions was not satisfying. Also, the antifungal activity and spectrum of a tested product based on tee tree oil was not efficient in suppressing pepper wilting caused by V. dahliae and F. oxysporum.

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In Vivo Recovery of Factor VIII: A Comparison of One-Stage and Two-Stage Assay Methods

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657018 ◽

1979 ◽

Vol 42 (04) ◽

pp. 1230-1239 ◽

Cited By ~ 46

Author(s):

I M Nilsson ◽

T B L Kirkwood ◽

T W Barrowcliffe

Keyword(s):

Factor Viii ◽

Half Life ◽

In Vitro Assays ◽

Two Stage ◽

One Stage ◽

Significant Difference ◽

Assay Methods ◽

The One ◽

Fraction I

SummaryThe recovery and half-life of VIII: C in the plasma of severely haemophilic patients was measured by one-stage and two-stage assays after injection of two Factor VIII concentrates (Hemofil, Hyland and Fraction I-O, Kabi). Plasma volumes were measured with an Evans� Blue technique, and both concentrates and post-infusion samples were measured against the same plasma standard.There was a highly significant difference in recoveries estimated by the two assay methods. The one-stage assays gave the most consistent results, in that the average recovery was 100%, whereas the two-stage assays gave only about 80% of the value expected from in vitro assays. There was no difference in recoveries between the two concentrates.The two-stage assays gave a slightly shorter half-life than the one-stage assays, and the half-life of Hemofil was also shorter than that of Fraction I-O.

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Comparative studies of in vitro transformation by ethylene oxide and gamma-radiation of C3H/10T1/2 cells

Mutagenesis ◽

10.1093/mutage/4.1.58 ◽

1989 ◽

Vol 4 (1) ◽

pp. 58-61 ◽

Cited By ~ 9

Author(s):

Ada Kolman ◽

Maria Näslund ◽

Siv Osterman-Golkar ◽

Gian-Paolo Scalia-Tomba ◽

Anthony Meyer

Keyword(s):

Ethylene Oxide ◽

Gamma Radiation ◽

Comparative Studies ◽

In Vitro Transformation

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Effect of gamma radiation on morphological and genetic variation in regenerated plantlets Catharanthus roseus L. (G) Don

Genetika ◽

10.2298/gensr2001015n ◽

2020 ◽

Vol 52 (1) ◽

pp. 15-28

Author(s):

Zahra Noormohammadi ◽

Narges Asghari-Mooneghi ◽

Farah Farahani

Keyword(s):

Gamma Radiation ◽

Catharanthus Roseus ◽

Genetic Difference ◽

Morphological Characteristics ◽

Stem Length ◽

Madagascar Periwinkle ◽

Genetic Changes ◽

Significant Difference ◽

Regenerated Plantlets

Catharanthus roseus L. (G) Don, commonly known as Madagascar periwinkle, is an important medicinal plant receiving much attention from researchers. This plant is a rich source of terpenoid indole alkaloids (TIAs). The gamma radiation and somaclonal variation may provide suitable methods for genetic changes to increase medicinal components of C. roseus. In the present study, 30 and 60 (Gy) gamma rays were irradiated on seeds. C. roseus seeds were cultured in Murashige and Skoog (MS) medium in three different groups, 0 Gy (control) 30, and 60 Gy gamma irradiations. Morphological characteristics of regenerated plantlets were measured for surface area of leaves, stem length, root length, and fresh weight. The genetic variations were evaluated using nine inter-simple sequence repeats and 25 sequences related amplified polymorphism markers. Morphological results showed a significant difference between studied groups in leaf surface and twisted leaf in 60 Gy irradiated plantlets. The analysis of molecular variance revealed the significant genetic difference among groups. STRUCTURE analysis showed genetic variability both within and among groups. Using the NJ tree analysis, the plants studied were categorized into three distinct groups, which were supported by principal coordinate analysis (PCoA). In conclusion, gamma irradiation produced morphological and genetic changes in C. roseus regenerated in vitro plantlets providing to be a suitable method for further mutation breeding in periwinkle.

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Fabrication, Characterization, and Cytotoxicity of Thermoplastic Polyurethane/Poly(lactic acid) Material Using Human Adipose Derived Mesenchymal Stromal Stem Cells (hASCs)

Polymers ◽

10.3390/polym10101073 ◽

2018 ◽

Vol 10 (10) ◽

pp. 1073 ◽

Cited By ~ 8

Author(s):

Anna Lis-Bartos ◽

Agnieszka Smieszek ◽

Kinga Frańczyk ◽

Krzysztof Marycz

Keyword(s):

Tissue Engineering ◽

Lactic Acid ◽

Thermoplastic Polyurethane ◽

Tensile Tests ◽

In Vitro Assays ◽

Poly Lactic Acid ◽

Scanning Calorimetry ◽

Casting Technique ◽

Wide Range

Thermoplastic polyurethane (TPU) and poly(lactic acid) are types of biocompatible and degradable synthetic polymers required for biomedical applications. Physically blended (TPU+PLA) tissue engineering matrices were produced via solvent casting technique. The following types of polymer blend were prepared: (TPU+PLA) 7:3, (TPU+PLA) 6:4, (TPU+PLA) 4:6, and (TPU+PLA) 3:7. Various methods were employed to characterize the properties of these polymers: surface properties such as morphology (scanning electron microscopy), wettability (goniometry), and roughness (profilometric analysis). Analyses of hydrophilic and hydrophobic properties, thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC) of the obtained polymer blends were conducted. Tensile tests demonstrated that the blends exhibited a wide range of mechanical properties. Cytotoxicity of polymers was tested using human multipotent stromal cells derived from adipose tissue (hASC). In vitro assays revealed that (TPU+PLA) 3:7 matrices were the most cytocompatible biomaterials. Cells cultured on (TPU+PLA) 3:7 had proper morphology, growth pattern, and were distinguished by increased proliferative and metabolic activity. Additionally, it appeared that (TPU+PLA) 3:7 biomaterials showed antiapoptotic properties. hASC cultured on these matrices had reduced expression of Bax-α and increased expression of Bcl-2. This study demonstrated the feasibility of producing a biocompatible scaffold form based on (TPU+PLA) blends that have potential to be applied in tissue engineering.

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Biochemical and Computational Studies of the Interaction between a Glucosamine Derivative, NAPA, and the IKKα Kinase

International Journal of Molecular Sciences ◽

10.3390/ijms22041643 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1643

Author(s):

Mariangela Lopreiato ◽

Samuele Di Cristofano ◽

Rossana Cocchiola ◽

Alessia Mariano ◽

Libera Guerrizio ◽

...

Keyword(s):

Mass Spectrometry ◽

Binding Site ◽

Kinase Activity ◽

In Vitro Assays ◽

Inhibition Mechanism ◽

Atomic Interaction ◽

Vitro Assay ◽

Inflammatory Status ◽

Significant Difference

The glucosamine derivative 2-(N-Acetyl)-L-phenylalanylamido-2-deoxy-β-D-glucose (NAPA), was shown to inhibit the kinase activity of IKKα, one of the two catalytic subunits of IKK complex, decreasing the inflammatory status in osteoarthritis chondrocytes. In the present work we have investigated the inhibition mechanism of IKKα by NAPA by combining computational simulations, in vitro assays and Mass Spectrometry (MS) technique. The kinase in vitro assay was conducted using a recombinant IKKα and IKKtide, a 20 amino acid peptide substrate derived from IkBα kinase protein and containing the serine residues Ser32 and Ser36. Phosphorylated peptide production was measured by Ultra Performance Liquid Chromatography coupled with Mass Spectrometry (UPLC-MS), and the atomic interaction between IKKα and NAPA has been studied by molecular docking and Molecular Dynamics (MD) approaches. Here we report that NAPA was able to inhibit the IKKα kinase activity with an IC50 of 0.5 mM, to decrease the Km value from 0.337 mM to 0.402 mM and the Vmax from 0.0257 mM·min−1 to 0.0076 mM·min−1. The computational analyses indicate the region between the KD, ULD and SDD domains of IKKα as the optimal binding site explored by NAPA. Biochemical data indicate that there is a non-significant difference between Km and Ki whereas there is a statistically significant difference between the two Vmax values. This evidence, combined with computational results, consistently indicates that the inhibition is non-competitive, and that the NAPA binding site is different than that of ATP or IKKtide.

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In vitro Assays ofStaphylococcus epidermidisCharacteristics and Outcome in an Endocarditis Model

Canadian Journal of Infectious Diseases ◽

10.1155/1993/832920 ◽

1993 ◽

Vol 4 (4) ◽

pp. 203-208 ◽

Cited By ~ 2

Author(s):

Betty Herndon ◽

Lawrence Dall ◽

William Barnes

Keyword(s):

Animal Model ◽

Prosthetic Valve Endocarditis ◽

In Vitro Assays ◽

Bacterial Surface ◽

Time To Death ◽

Death Time ◽

Colony Forming Units ◽

Significant Difference ◽

Patient Groups

Objective:Staphylococcus epidermidisadherence to indwelling polymers is important in prosthetic valve endocarditis. Earlier studies have related streptococcal endocarditis to isolates with high levels of cell-associated hexoses. The objective of the present study was to determine if a relationship exists between anS epidermidisisolate assay score and production/severity of experimental endocarditis.Design: Groups of patientS epidermidisisolates were screened for surface hexoses and an animal model of endocarditis with isolates testing highest and lowest on the screen was produced. Disease severity produced by ‘high hexose’ versus ‘low hexose’ organisms was evaluated. Endocarditis responding variables were bacterial vegetation weight and log10colony forming units (cfu) and in survival tests, comparative time to death with different isolates. Bacterial characteristics were not measured. Baseline data showed a vegetation weight difference so that with a β error of 0.20 and a two-tailed α error of 0.05, a significant difference would be noted using 30 animals. A total of 64 animals was used.Population Studded: Bacterial isolates from two patient groups (n=42 and n=68) on which in vitro assays were run. An animal model of endocarditis (n=64) was used to evaluate four selected isolates for vegetation size, log10cfu/g, and survival time.Main Results: In a group ofS epidermidisendocarditis animals evaluated for time of death, a significantly more rapid death time resulted in the group dosed with the high hexose-scoring organism (P<0.025). Vegetations and log10cfu produced by test high hexose isolates averaged larger but were not significantly different.Conclusions: A significantly more rapid death rate occurs in untreated endocarditis using a high hexose isolate than withS epidermidiswith low surface hexoses. Using bacterial vegetation and cfu as endpoints, however, experimental endocarditis using patient isolates ofS epidermidisdoes not show the same strong correlation to bacterial surface hexoses as does streptococcal endocarditis.

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In vitro release of cytotoxic nucleoside analogs from lactide-caprolactone and lactide-glycolide copolymers.

Acta Biochimica Polonica ◽

10.18388/abp.2002_3837 ◽

2002 ◽

Vol 49 (1) ◽

pp. 205-210 ◽

Cited By ~ 5

Author(s):

Tomasz Kryczka ◽

Maciej Bero ◽

Janusz Kasperczyk ◽

Piotr Dobrzyński ◽

Barbara Marciniec ◽

...

Keyword(s):

Ethylene Oxide ◽

Gamma Radiation ◽

Local Treatment ◽

In Vitro Release ◽

Nucleoside Analogs ◽

Further Development ◽

Kinetics Of ◽

Vitro Release ◽

Observation Period

The aims of our study were to assess the release of cytotoxic nucleoside analogs 5-fluorouracil and 2-chloro-2'-deoxyadenosine from different lactide-glycolide or lactide-caprolactone biodegradable copolymers and the effects of sterilization on this release. The polymers were sterilized either with ethylene oxide at 37 degrees C, or with gamma radiation (15 kGy, 20 kGy, or 25 kGy). The kinetics of nucleoside release from the copolymers were measured over 50 days. Four copolymers exhibited relatively constant release of nucleosides in micromolar concentrations during the entire observation period. Sterilization with either ethylene oxide or gamma radiation only slightly influenced nucleoside release. Further development of these copolymers as an intracerebral nucleoside delivery system for local treatment of brain tumors is indicated.

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