scholarly journals Addition of Popular Exogenous Antioxidant Agent, PBN, to Culture Media May Be an Important Step to Optimization of Myogenic Stem/Progenitor Cell Preparation Protocol

Antioxidants ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 959
Author(s):  
Magdalena Nowaczyk ◽  
Agnieszka Malcher ◽  
Agnieszka Zimna ◽  
Wojciech Łabędź ◽  
Łukasz Kubaszewski ◽  
...  
Keyword(s):  
Gene Expression ◽  
Progenitor Cell ◽  
Human Skeletal Muscle ◽  
Culture Medium ◽  
Culture Media ◽  
Cell Preparation ◽  
Phenotypic Marker ◽  
Hypoxic Conditions ◽  
Genes Expression ◽  
Antioxidant Agent

The aim of the study was to modify human skeletal muscle-derived stem/progenitor cells (SkMDS/PCs) and demonstrate the optimal cell preparation protocol for application in post-infarction hearts. We used conditioned SkMDS/PC culture medium with α-phenyl-N-tert-butyl nitrone (PBN). SkMDS/PCs were cultured under hypoxic conditions and the results were compared to the standard ones. We observed a significant increase of CD-56 positive phenotypic marker the ability to form functional myotubes, increase in the proportion of young cells in cell primary suspensions, and a decrease in the percentage of apoptotic cells among PBN-conditioned cells in normoxia an hypoxia. We also observed significantly higher levels of SOD3 expression; maintained expression of SOD1, SOD2, and CAT; a higher level of BCL2 gene expression; and a rather significant decrease in Hsp70 gene expression in PBN-conditioned SkMDS/PCs compared to the WT population under hypoxic conditions. In addition, significant increase of myogenic genes expression was observed after PBN addition to culture medium, compared to WT population under hypoxia. Interestingly, PBN addition significantly increased the lengths of telomeres under hypoxia. Based on the data obtained, we can postulate that PBN conditioning of human SkMDS/PCs could be a promising step in improving myogenic cell preparation protocol for pro-regenerative treatment of post-infarction hearts.

scholarly journals Triiodothyronine Improves Morphology and Up-regulates seladin-1 of neurospheres extracted from Subventricular Zone in Streptozotocin-induced rat model of Alzheimer's disease

2020 ◽  
Vol 6 (1) ◽  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Rat Model ◽  
Subventricular Zone ◽  
Wistar Rats ◽  
Culture Medium ◽  
Culture Media ◽  
Model Of Alzheimer’S Disease ◽  
Sh Group ◽  
Male Wistar Rats

Objectives: In this study, the effects of triiodothyronine (T3) on neurospheres isolated from SVZ of AD induced rats were examined. Methods: Eighteen male Wistar rats were classified into two groups: Sham (Sh) and STZ (Streptozotocin injected, 1.5 mg/kg in each lateral ventricle on days 1 and 3 after recovery). On day 21, the SVZ was extracted and neurospheres were cultured. T3 (50 nM) was added to the culture medium (STZ+T3 group) and then, the morphology and seladin-1 gene expression of neurospheres were evaluated. Results: The diameter and the number of neurospheres along with the gene expression of seladin-1 were significantly decreased in the STZ group compared to Sh group (P˂0.05) while the administration of T3 significantly (P˂0.05) increased all these parameters in the STZ group. Conclusion: STZ decreases the proliferation of stem cells extracted from SVZ and administration of T3 to the culture media improves the morphology and up-regulates the gene expression of seladin-1 of neurospheres.

scholarly journals OPTIMIZATION OF FACTORS AFFECTING THE Agrobacterium tumefaciens- MEDIATED TRANSFORMATION OF Eucalyptus saligna

2018 ◽  
Vol 41 (3) ◽  
Author(s):  
Yohana de Oliveira-Cauduro ◽  
Lais Gomes Adamuchio ◽  
João Carlos Bespalhok Filho ◽  
Isabel Rodrigues Gerhardt ◽  
Juliana Degenhardt-Goldbach ◽  
...  
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Culture Medium ◽  
Culture Media ◽  
Gus Gene ◽  
Eucalyptus Saligna ◽  
Factors Affecting ◽  
Gus Gene Expression ◽  
Positive Effect ◽  
Selection Of

ABSTRACT This study aimed to evaluate the effect of factors that may affect the genetic transformation of cotiledonary explants of Eucalyptus saligna mediated by EHA105 strain of Agrobacterium tumefaciens. The vector pBI121 carrying gus gene under control of 35S CaMV promoter was used. The effect of the following factors was evaluated: explant pre-culture, use of different antibiotics and presence of acetosyringone (AS) in co-culture media. An antioxidant solution was also used during excision, containing ascorbic acid (250mg.L-1), citric acid (25mg.L-1) and PVP-40 (1g.L-1). Pre-culture of the explants before the co-culture with bacteria was done over a 4-day period in MS culture medium supplemented with 4.4µM BAP and 2.7ìM NAA. After theco-culture period, three concentrations of kanamycin (12.5;25 and 50mg.L-1) combined with 300mg.L-1 Augmentin® in the culture medium were tested The influence of the antibiotic was also evaluated by keeping the explants in a medium containing 50mg.L-1 Km and 300mg.L-1 Augmentin® or 500mg.L-1 cefotaxime. It was concluded that Augmentin® stimulates organogenesis, that a Km concentration of 12.5mg.L-1 allows selection of explants transformed with gus gene and, finally, the addition of AS (50ìM) to the liquid and solid co-culture media has a positive effect on gus gene expression. Moreover, the use of an antioxidant solution during cotyledon excision is dispensable and the pre-culture of the explants has no effect on bud regeneration or gus gene expression. A transformation efficiency of 1.5% was reached.

293 EFFECT OF MEDIUM TYPE FOR CULTURE OF ADIPOSE-DERIVED MESENCHYMAL STEM CELLS ON PRE-IMPLANTATION DEVELOPMENT OF CLONED EMBRYOS

2013 ◽  
Vol 25 (1) ◽  
pp. 294
Author(s):  
G. A. Kim ◽  
H. J. Oh ◽  
J. Kim ◽  
T. H. Lee ◽  
J. H. Lee ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Culture Medium ◽  
Culture Media ◽  
Formation Rate ◽  
Donor Cells

Mesenchymal stem cells (MSC) have been known as useful donor cells for somatic cell nuclear transfer (SCNT). It has been suggested that the culture condition of donor cells causes different results on preimplantation development of SCNT embryos. In this study, we investigated the patterns of gene expression of adipose-derived mesenchymal stem cells (ad-MSC) in different culture media (DMEM and RKME), and examined the effect of ad-MSC, with the gene expression changed, used as donor cells on the preimplantation development of cloned embryos. Canine ad-MSC were isolated from fat tissue of 3-year-old female beagle and were cultured in DMEM supplemented with 10% fetal bovine serum (MSC-DMEM) and RKME (MSC-MSC) provided from RNL Bio Corp. (Seoul, Korea). Total RNA was extracted from ad-MSC cultured in each culture medium. After synthesising cDNA of each sample, quantitative RT-PCR was done according to the Takara Bio Inc. guidelines and using the 7300 Real Time PCR Cycler System (Applied Biosystems, Carlsbad, CA, USA). The level of all tested gene transcription was normalized to β-actin expression levels. The relative quantification of gene expression was analysed by the 2–ΔΔCt method. The data from all experiments were analysed by Student’s t-test using a statistical analysis GraphPad Prism 4.02 (GraphPad Software Inc., San Diego, CA, USA). Significance was determined at P < 0.05. The stemness, the reprogramming-related gene expression level of donor cells of MSC-DMEM and MSC-MSC were compared. In order to confirm the effect of MSC cultured in 2 different culture media on somatic cell nuclear transfer, we performed interspecies somatic cell nuclear transfer (iSCNT). The enucleated bovine oocytes were injected, respectively, with donor cells of MSC-DMEM and MSC-MSC, and were fused by electrofusion. The iSCNT embryos were cultured in modified SOF at 38.5°C for 7 days in an atmosphere of 5% CO2 and 5% O2, and the developmental ability of iSCNT embryos was observed under the microscope. The MSC-MSC contained a significantly higher amount of Sox2, Nanog, Oct4, Stella, HDAC1, DNMT1, and MeCP2 than the MSC-DMEM, whereas the amount of Rex1 was not different in either MSC-MSC or MSC-DMEM. In the development ability of iSCNT embryos, MSC-DMEM embryos resulted in a 16-cell embryo formation rate that was higher than that of MSC-MSC embryos (9.09 and 5.30%, respectively; P < 0.05). However, the blastocyst formation rate was not different between MSC-DMEM embryos and MSC-MSC embryos (4.5 and 3.2%, respectively; P > 0.05). These results demonstrate that the gene expression of ad-MSC can be modified, by culture media, into a state where reprogramming is easily done. Even so, ad-MSC with gene expression changed by culture medium did not influence the developmental ability of blastocysts. In conclusion, the alteration of gene-related stemness and reprogramming in canine ad-MSC would not be able to effectively control reprogramming in SCNT. This study was supported by RDA (#PJ0089752012), RNL Bio (#550-20120006), IPET (#311062-04-1-SB010), Research Institute for Veterinary Science, and Nestlé Purina Korea.

scholarly journals Improvement of Cell Culture Methods for the Successful Generation of Human Keratinocyte Primary Cell Cultures Using EGF-Loaded Nanostructured Lipid Carriers

Biomedicines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1634
Author(s):  
Jesús Chato-Astrain ◽  
David Sánchez-Porras ◽  
Óscar Darío García-García ◽  
Claudia Vairo ◽  
María Villar-Vidal ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Skin ◽  
Cell Cultures ◽  
Culture Medium ◽  
Culture Media ◽  
Nanostructured Lipid Carriers ◽  
Primary Cell ◽  
Primary Cell Cultures ◽  
Human Keratinocyte ◽  
Skin Keratinocytes

Human skin keratinocyte primary cultures can be established from skin biopsies with culture media containing epithelial growth factor (EGF). Although current methods are efficient, optimization is required to accelerate the procedure and obtain these cultures in less time. In the present study, we evaluated the effect of novel formulations based on EGF-loaded nanostructured lipid carriers (NLC). First, biosafety of NLC containing recombinant human EGF (NLC-rhEGF) was verified in immortalized skin keratinocytes and cornea epithelial cells, and in two epithelial cancer cell lines, by quantifying free DNA released to the culture medium. Then we established primary cell cultures of human skin keratinocytes with basal culture media (BM) and BM supplemented with NLC-rhEGF, liquid EGF (L-rhEGF), or NLC alone (NLC-blank). The results showed that cells isolated by enzymatic digestion and cultured with or without a feeder layer had a similar growth rate regardless of the medium used. However, the explant technique showed higher efficiency when NLC-rhEGF culture medium was used, compared to BM, L-rhEGF, or NLC-blank. Gene expression analysis showed that NLC-rhEGF was able to increase EGFR gene expression, along with that of other genes related to cytokeratins, cell–cell junctions, and keratinocyte maturation and differentiation. In summary, these results support the use of NLC-rhEGF to improve the efficiency of explant-based methods in the efficient generation of human keratinocyte primary cell cultures for tissue engineering use.

scholarly journals Subculture human skeletal muscle cells to produce the cells with different Culture medium compositions

2021 ◽  
Vol 3 (4) ◽  
pp. 01-03
Author(s):  
Ying Balch
Keyword(s):  
Skeletal Muscle ◽  
Growth Medium ◽  
Human Skeletal Muscle ◽  
Culture Medium ◽  
Culture Media ◽  
Muscle Cells ◽  
Skeletal Muscle Cells ◽  
Human Skeletal Muscle Cells ◽  
Group 3 ◽  
Group 2

This study aimed to subculture human skeletal muscle cells (HSkMC) using a culture medium with different compositions to determine the most efficient medium for the growth of the human skeletal muscle cells. The culture media was divided into three groups: Group1. An HSkMC growth medium. Group 2. An HSkMC growth medium + with 10% high glucose (GH). Group 3. An HSkMC growth medium + 10% fetal bovine serum (FBS). HSkMC from groups 1 to 3 gradually became round in shape and gathered in clusters. These changes differed between the groups. In group 3, the HSkMC clusters were more in numbers and gathered as significantly more prominent than in the other groups under the EVOS-Microscope shown. We concluded that by manipulating the composition of the culture medium, it is possible to induce HSkMC to promote the best growth.

scholarly journals Influence of sucrose concentration in the culture medium on the condition of the photosynthetic apparatus of grapes cultured in vitro

2020 ◽  
Vol 25 ◽  
pp. 04003
Author(s):  
Maria Sundyreva ◽  
Anton Rebrov ◽  
Alisa Mishko
Keyword(s):  
Culture Medium ◽  
Culture Media ◽  
Sucrose Concentration ◽  
Photosynthetic Apparatus ◽  
Photosynthetic Parameters ◽  
Photosynthetic Genes ◽  
Genes Expression ◽  
Oxidative Processes ◽  
Stress Formation

An influence of different sucrose concentrations in the culture media on the photosynthetic parameters, photosynthetic apparatus related genes expression, oxidative processes and acclimation of grape plants cultured in vitro was examined in this article. An increase of the sucrose concentration in the culture media resulted in a reduced expression of several photosynthetic genes. The most effective functioning of the photosynthetic apparatus was discovered by a decreased amount of surcose in culture media. An increase of the sucrose concentration in the culture media disrupts pigments synthesis, particularly carotenoids, which can be a cause of the secondary oxidative stress formation and grape plants growth reduction during acclimation.

Pertumbuhan Dan Perkembangan Anggrek Dendrobium anosmum Pada Media Kultur In Vitro Dengan Beberapa Konsentrasi Air Kelapa

Agrologia ◽  
2018 ◽  
Vol 1 (1) ◽  
Author(s):  
S. Tuhuteru ◽  
Meity L Hehanussa ◽  
Simon H.T Raharjo
Keyword(s):  
Culture Medium ◽  
Culture Media ◽  
Water Concentration ◽  
Medium Composition ◽  
Coconut Water ◽  
Orchid Species ◽  
Randomized Design ◽  
Wet Weight ◽  
Completely Randomized Design

Dendrobium anosmum is one of natural orchids in Indonesia. Optimization of medium composition for orchid propagation through in vitro culture is necessary to enhance propagule multiplication capabilities and quality. This study was aimed to study the influence of concentration of coconut water in culture medium on in vitro growth and development of D. anosmum orchid species and to determine the optimal coconut water concentration in culture media.  The experiment were arranged in a Completely Randomized Design with four treatments and eight replications. The treatments consisted of the addition of coconut water with concentrations: 0 ml•l -1 (control), 50 ml•l-1, 100 ml•l-1 and 150 ml•l-1. The results showed that addition of coconut water in culture medium gave different effect on shoot growth and multiplication of D. anosmum orchids.  Coconut water concentration of 100 ml•l-1 was the best concentration for growth and multiplication of D. anosmum orchids, based on both shoots and roots growth, plantlet height and wet weight.

scholarly journals Untargeted Metabolomics Approach for the Discovery of Environment-Related Pyran-2-ones Chemodiversity in a Marine-Sourced Penicillium restrictum

Marine Drugs ◽  
2021 ◽  
Vol 19 (7) ◽  
pp. 378
Author(s):  
Van-Tuyen Le ◽  
Samuel Bertrand ◽  
Thibaut Robiou du Pont ◽  
Fabrice Fleury ◽  
Nathalie Caroff ◽  
...  
Keyword(s):  
Culture Medium ◽  
Blue Mussel ◽  
Culture Media ◽  
Principal Component ◽  
Chemical Diversity ◽  
Untargeted Metabolomics ◽  
High Chemical ◽  
Medium Mass ◽  
Metabolomics Study ◽  
Penicillium Restrictum

Very little is known about chemical interactions between fungi and their mollusc host within marine environments. Here, we investigated the metabolome of a Penicillium restrictum MMS417 strain isolated from the blue mussel Mytilus edulis collected on the Loire estuary, France. Following the OSMAC approach with the use of 14 culture media, the effect of salinity and of a mussel-derived medium on the metabolic expression were analysed using HPLC-UV/DAD-HRMS/MS. An untargeted metabolomics study was performed using principal component analysis (PCA), orthogonal projection to latent structure discriminant analysis (O-PLSDA) and molecular networking (MN). It highlighted some compounds belonging to sterols, macrolides and pyran-2-ones, which were specifically induced in marine conditions. In particular, a high chemical diversity of pyran-2-ones was found to be related to the presence of mussel extract in the culture medium. Mass spectrometry (MS)- and UV-guided purification resulted in the isolation of five new natural fungal pyran-2-one derivatives—5,6-dihydro-6S-hydroxymethyl-4-methoxy-2H-pyran-2-one (1), (6S, 1’R, 2’S)-LL-P880β (3), 5,6-dihydro-4-methoxy-6S-(1’S, 2’S-dihydroxy pent-3’(E)-enyl)-2H-pyran-2-one (4), 4-methoxy-6-(1’R, 2’S-dihydroxy pent-3’(E)-enyl)-2H-pyran-2-one (6) and 4-methoxy-2H-pyran-2-one (7)—together with the known (6S, 1’S, 2’S)-LL-P880β (2), (1’R, 2’S)-LL-P880γ (5), 5,6-dihydro-4-methoxy-2H-pyran-2-one (8), (6S, 1’S, 2’R)-LL-P880β (9), (6S, 1’S)-pestalotin (10), 1’R-dehydropestalotin (11) and 6-pentyl-4-methoxy-2H-pyran-2-one (12) from the mussel-derived culture medium extract. The structures of 1-12 were determined by 1D- and 2D-MMR experiments as well as high-resolution tandem MS, ECD and DP4 calculations. Some of these compounds were evaluated for their cytotoxic, antibacterial, antileishmanial and in-silico PTP1B inhibitory activities. These results illustrate the utility in using host-derived media for the discovery of new natural products.

scholarly journals Rheological and Microstructural Features of Plant Culture Media Doped with Biopolymers: Influence on the Growth and Physiological Responses of In Vitro-Grown Shoots of Thymus lotocephalus

2021 ◽  
Vol 2 (2) ◽  
pp. 538-553
Author(s):  
Natacha Coelho ◽  
Alexandra Filipe ◽  
Bruno Medronho ◽  
Solange Magalhães ◽  
Carla Vitorino ◽  
...  
Keyword(s):  
In Vitro Culture ◽  
Culture Medium ◽  
Culture Media ◽  
Average Pore Size ◽  
Physiological Responses ◽  
Gum Arabic ◽  
Shoot Elongation ◽  
Hydroxyethyl Cellulose ◽  
Plant Culture

In vitro culture is an important biotechnological tool in plant research and an appropriate culture media is a key for a successful plant development under in vitro conditions. The use of natural compounds to improve culture media has been growing and biopolymers are interesting alternatives to synthetic compounds due to their low toxicity, biodegradability, renewability, and availability. In the present study, different culture media containing one biopolymer (chitosan, gum arabic) or a biopolymer derivative [hydroxyethyl cellulose (HEC), carboxymethyl cellulose (CMC)], at 100 or 1000 mg L−1, were tested regarding their influence on the growth and physiological responses of Thymus lotocephalus in vitro culture. Cellulose-based biopolymers (HEC and CMC) and gum arabic were used for the first time in plant culture media. The results showed that CMC at 100 mg L−1 significantly improved shoot elongation while chitosan, at the highest concentration, was detrimental to T. lotocephalus. Concerning only the evaluated physiological parameters, all tested biopolymers and biopolymer derivatives are safe to plants as there was no evidence of stress-induced changes on T. lotocephalus. The rheological and microstructural features of the culture media were assessed to understand how the biopolymers and biopolymer derivatives added to the culture medium could influence shoot growth. As expected, all media presented a gel-like behaviour with minor differences in the complex viscosity at the beginning of the culture period. Most media showed increased viscosity overtime. The surface area increased with the addition of biopolymers and biopolymer derivatives to the culture media and the average pore size was considerably lower for CMC at 100 mg L−1. The smaller pores of this medium might be related to a more efficient nutrients and water uptake by T. lotocephalus shoots, leading to a significant improvement in shoot elongation. In short, this study demonstrated that the different types of biopolymers and biopolymer derivatives added to culture medium can modify their microstructure and at the right concentrations, are harmless to T. lotocephalus shoots growing in vitro, and that CMC improves shoot length.

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