scholarly journals Graphene oxide nanofilm and the addition of L-glutamine can promote development of embryonic muscle cells

2020 ◽  
Author(s):  
Marlena Zielińska-Górska ◽  
Anna Hotowy ◽  
Mateusz Wierzbicki ◽  
Jaśmina Bałaban ◽  
Malwina Sosnowska ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Surface Properties ◽  
Satellite Cells ◽  
Chicken Embryo ◽  
Culture Medium ◽  
Muscle Cells ◽  
Control Group ◽  
Protein Levels ◽  
Myogenic Potential

Abstract Background: Formation of muscular pseudo-tissue depends on muscle precursor cells, the extracellular matrix (ECM)-mimicking structure and factors stimulating cell differentiation. These three things cooperate and can create a tissue-like structure, however, their interrelationships are relatively unknown. The objective was to study the interaction between surface properties, culture medium composition and heterogeneous cell culture. We would like to demonstrate that changing the surface properties by coating with graphene oxide nanofilm (nGO) can affect cell behaviour and especially their need for the key amino acid L-glutamine (L-Glu). Results: Chicken embryo muscle cells and their precursors, cultured in vitro, were used as the experimental model. The mesenchymal stem cell, collected from the hind limb of the chicken embryo at day 8 were divided into 4 groups; the control group and groups treated with nGO, L-Glu and nGO supplied with L-Glu (nGOxL-Glu). The roughness of the surface of the plastic plate covered with nGO was much lower than a standard plate. The test of nGO biocompatibility demonstrated that the cells were willing to settle on the nGO without any toxic effects. Moreover, nGO by increasing hydrophilicity and reducing roughness stimulated the colonisation of primary stromal cells that promote embryonic satellite cells. The viability significantly increased in cells cultured on nGOxL-Glu. Observations of cell morphology showed that the most mature state of myogenesis was characteristic for the group nGOxL-Glu. This result was confirmed by increasing the expression of MYF5 genes at mRNA and protein levels. nGO also increased the expression of MYF5 and also very strongly the expression of PAX7 at mRNA and protein levels. However, when analysing the expression of PAX7 , a positive link was observed between the nGO surface and the addition of L-Glu. Conclusions: The use of nGO and L-Glu supplement may improve myogenesis and also the myogenic potential of myocytes and their precursors by promoting the formation of satellite cells. Studies have, for the first time, demonstrated positive cooperation between surface properties nGO and L-Glu supplementation to the culture medium regarding the myogenic potential of cells involved in muscle formation.

scholarly journals Graphene oxide nanofilm and the addition of L-glutamine can promote development of embryonic muscle cells

2020 ◽  
Author(s):  
Marlena Zielińska-Górska ◽  
Anna Hotowy ◽  
Mateusz Wierzbicki ◽  
Jaśmina Bałaban ◽  
Malwina Sosnowska ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Surface Properties ◽  
Satellite Cells ◽  
Chicken Embryo ◽  
Culture Medium ◽  
Muscle Cells ◽  
Control Group ◽  
Protein Levels ◽  
Myogenic Potential

Abstract Background: Formation of muscular pseudo-tissue depends on muscle precursor cells, the extracellular matrix (ECM)-mimicking structure and factors stimulating cell differentiation. These three things cooperate and can create a tissue-like structure, however, their interrelationships are relatively unknown. The objective was to study the interaction between surface properties, culture medium composition and heterogeneous cell culture. We would like to demonstrate that changing the surface properties by coating with graphene oxide nanofilm (nGO) can affect cell behaviour and especially their need for the key amino acid L-glutamine (L-Glu).Results: Chicken embryo muscle cells and their precursors, cultured in vitro, were used as the experimental model. The mesenchymal stem cell, collected from the hind limb of the chicken embryo at day 8 were divided into 4 groups; the control group and groups treated with nGO, L-Glu and nGO supplied with L-Glu (nGOxL-Glu). The roughness of the surface of the plastic plate covered with nGO was much lower than a standard plate. The test of nGO biocompatibility demonstrated that the cells were willing to settle on the nGO without any toxic effects. Moreover, nGO by increasing hydrophilicity and reducing roughness and presumably through chemical bonds available on the GO surface stimulated the colonisation of primary stromal cells that promote embryonic satellite cells. The viability significantly increased in cells cultured on nGOxL-Glu. Observations of cell morphology showed that the most mature state of myogenesis was characteristic for the group nGOxL-Glu. This result was confirmed by increasing the expression of MYF5 genes at mRNA and protein levels. nGO also increased the expression of MYF5 and also very strongly the expression of PAX7 at mRNA and protein levels. However, when analysing the expression of PAX7, a positive link was observed between the nGO surface and the addition of L-Glu.Conclusions: The use of nGO and L-Glu supplement may improve myogenesis and also the myogenic potential of myocytes and their precursors by promoting the formation of satellite cells. Studies have, for the first time, demonstrated positive cooperation between surface properties nGO and L-Glu supplementation to the culture medium regarding the myogenic potential of cells involved in muscle formation.

scholarly journals Calcium Carbonate Nanoparticles—Toxicity and Effect of In Ovo Inoculation on Chicken Embryo Development, Broiler Performance and Bone Status

Animals ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 932
Author(s):  
Arkadiusz Matuszewski ◽  
Monika Łukasiewicz ◽  
Jan Niemiec ◽  
Maciej Kamaszewski ◽  
Sławomir Jaworski ◽  
...  
Keyword(s):  
Calcium Carbonate ◽  
Embryo Development ◽  
Bone Quality ◽  
Chicken Embryo ◽  
Broiler Chickens ◽  
Selection Procedure ◽  
High Specificity ◽  
Control Group ◽  
In Ovo

The use of intensive selection procedure in modern broiler chicken lines has led to the development of several skeletal disorders in broiler chickens. Therefore, current research is focused on methods to improve the bone quality in birds. In ovo technology, using nanoparticles with a high specificity to bones, is a potential approach. The present study aimed to evaluate the effect of in ovo inoculation (IOI) of calcium carbonate nanoparticles (CCN) on chicken embryo development, health status, bone characteristics, and on broiler production results and bone quality. After assessing in vitro cell viability, the IOI procedure was performed with an injection of 500 μg/mL CCN. The control group was not inoculated with CCN. Hatchability, weight, and selected bone and serum parameters were measured in embryos. Part of hatchlings were reared under standard conditions until 42 days, and production results, meat quality, and bone quality of broilers were determined. CCN did not show cytotoxicity to cells and chicken embryo and positively influenced bone parameters of the embryos and of broilers later (calcification) without negatively affecting the production results. Thus, the IOI of CCN could modify the molecular responses at the stage of embryogenesis, resulting in better mineralization, and could provide a sustained effect, thereby improving bone quality in adult birds.

YAP1 Overexpression Is Associated with Kidney Dysfunction in Lupus Nephritis

Pathobiology ◽  
2021 ◽  
pp. 1-12
Author(s):  
Ying Xie ◽  
Yuanyuan Ruan ◽  
Huimei Zou ◽  
Yixin Wang ◽  
Xin Wu ◽  
...  
Keyword(s):  
Lupus Nephritis ◽  
Epithelial Mesenchymal Transition ◽  
Kidney Tissue ◽  
Mouse Kidney ◽  
Experimental Comparison ◽  
Control Group ◽  
Mrna Levels ◽  
Mesenchymal Transition ◽  
Protein Levels

<b><i>Objective:</i></b> The goal of the present study was to determine the expression of yes-associated protein 1 (YAP1) in renal tissues of mice with lupus nephritis (LN) and elucidate its role in the progression of renal fibrosis. <b><i>Methods:</i></b> C57BL/6 mice and MRL/lpr mice were selected for experimental comparison. Mouse kidney tissues were removed and sectioned for hematoxylin and eosin staining, Masson’s trichome staining, Sirius staining, and immunohistochemistry. The mRNA and protein levels of YAP1 in mouse kidney tissues were detected, and the correlation between YAP1 and fibronectin (FN) mRNA levels was analyzed. Mouse renal epithelial cells were used for in vitro experiments. After transfection and stimulation, the cells were divided into 4 groups, namely the C57BL/6 serum group (group 1), the MRL/lpr serum group (group 2), the MRL/lpr serum + siRNA-negative control group (group 3), and the MRL/lpr serum + siRNA-YAP1 group (group 4). Epithelial-mesenchymal transition (EMT) markers in each group were detected by Western blotting and immunofluorescence staining. Serum creatinine, blood urea nitrogen, and urinary protein levels were detected and assessed for their correlation with YAP1 mRNA levels by Spearman’s analysis. <b><i>Results:</i></b> Compared to C57BL/6 mice, MRL/lpr mice exhibited obvious changes in fibrosis in renal tissues. In addition, YAP1 expression was significantly higher in the renal tissues of MRL/lpr mice than in those of C57BL/6 mice, and YAP1 mRNA levels were positively correlated with those of FN. YAP1 silencing in lupus serum-stimulated cells could effectively relieve serum-induced EMT. Finally, we observed that YAP1 mRNA levels in mouse kidney tissue were significantly and positively correlated with the degree of renal function injury. <b><i>Conclusion:</i></b> YAP1 expression in the kidney tissues of LN mice was higher than that observed in normal mice, indicating that YAP1 may play an important role in the occurrence and development of LN.

scholarly journals Effects of temperature on proliferation of myoblasts from donor piglets with different thermoregulatory maturities

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Katharina Metzger ◽  
Dirk Dannenberger ◽  
Armin Tuchscherer ◽  
Siriluck Ponsuksili ◽  
Claudia Kalbe
Keyword(s):  
Muscle Development ◽  
Muscle Growth ◽  
Extreme Temperature ◽  
Growth Inhibitor ◽  
Muscle Cells ◽  
The Body ◽  
Farm Animals ◽  
Neonatal Piglets ◽  
Protein Levels

Abstract Background Climate change and the associated risk for the occurrence of extreme temperature events or permanent changes in ambient temperature are important in the husbandry of farm animals. The aim of our study was to investigate the effects of permanent cultivation temperatures below (35 °C) and above (39 °C, 41 °C) the standard cultivation temperature (37 °C) on porcine muscle development. Therefore, we used our porcine primary muscle cell culture derived from satellite cells as an in vitro model. Neonatal piglets have limited thermoregulatory stability, and several days after birth are required to maintain their body temperature. To consider this developmental step, we used myoblasts originating from thermolabile (five days of age) and thermostable piglets (twenty days of age). Results The efficiency of myoblast proliferation using real-time monitoring via electrical impedance was comparable at all temperatures with no difference in the cell index, slope or doubling time. Both temperatures of 37 °C and 39 °C led to similar biochemical growth properties and cell viability. Only differences in the mRNA expression of myogenesis-associated genes were found at 39 °C compared to 37 °C with less MYF5, MYOD and MSTN and more MYH3 mRNA. Myoblasts grown at 35 °C are smaller, exhibit higher DNA synthesis and express higher amounts of the satellite cell marker PAX7, muscle growth inhibitor MSTN and metabolic coactivator PPARGC1A. Only permanent cultivation at 41 °C resulted in higher HSP expression at the mRNA and protein levels. Interactions between the temperature and donor age showed that MYOD, MYOG, MYH3 and SMPX mRNAs were temperature-dependently expressed in myoblasts of thermolabile but not thermostable piglets. Conclusions We conclude that 37 °C to 39 °C is the best physiological temperature range for adequate porcine myoblast development. Corresponding to the body temperatures of piglets, it is therefore possible to culture primary muscle cells at 39 °C. Only the highest temperature of 41 °C acts as a thermal stressor for myoblasts with increased HSP expression, but it also accelerates myogenic development. Cultivation at 35 °C, however, leads to less differentiated myoblasts with distinct thermogenetic activity. The adaptive behavior of derived primary muscle cells to different cultivation temperatures seems to be determined by the thermoregulatory stability of the donor piglets.

scholarly journals Experimental conversion of virulent RH Toxoplasma gondii tachyzoites in vitro

2001 ◽  
Vol 7 (1-2) ◽  
pp. 181-188
Author(s):  
N. A. Hammouda ◽  
I. R. Ibrahim ◽  
E. D. Elkerdany ◽  
A. Y. Negm ◽  
S. R. Allam
Keyword(s):  
Toxoplasma Gondii ◽  
Culture Medium ◽  
Dna Analysis ◽  
Morphological Changes ◽  
Control Group ◽  
Image Analyser

We aimed to induce conversion of RH-stain tachyzoites to bradyzoites by changing the pH of the culture medium. Alkalization of the medium to pH 8 induced morphological changes in the cultured tachyzoites. The majority of the organism increased in size and changed from a regular crescent shape to a rounded or ovoid shape. Cyst-like structures were formed. Using a computerized image analyser, significant differences in the size of the whole organisms and in their nuclei were observed compared to the control group. The converted organisms also showed significant differences from the control group by quantitative DNA analysis, and did not infect mice.

scholarly journals Indirect Cold Atmospheric Pressure Plasma treatment on Melanoma and Fibroblast Cells

2020 ◽  
Vol 10 (2) ◽  
pp. 2117-2125
Keyword(s):  
Plasma Treatment ◽  
Cancer Cells ◽  
Culture Medium ◽  
Atmospheric Pressure Plasma ◽  
Control Group ◽  
Fibroblast Cells ◽  
Alizarin Red ◽  
Migration Assay ◽  
Melanoma Cancer

Cold atmosphere plasma has been shown as a promising technology for certain cancer treatments. In this paper, we report indirect plasma treatment using CAP discharged in cell culture medium and study the effect of identical plasma stimulated culture medium on melanoma cancer cells and fibroblast cells cultured in vitro. The results of MTT assay, migration assay, ROS detection, and alizarin red assay show that plasma-treated medium can have a strong negative effect on melanoma cancer cells compared with the control group. However, the plasma-treated medium has a less cytotoxic effect on fibroblast cells than that on melanoma cancer cells at the same treatment. This result is attributed to the production of reactive oxygen species in the plasma-treated medium to induce apoptosis and inhibit melanoma cell proliferation and further cell metastasis. According to the results, this study shows the potential of CAP plasma treatment for anti-cancer therapy.

The Cultivation of the Pituitary of Infantile Rats by the Glass-Rod Technique and the Influence of Grafted Explants on the Growth of Hypophysectomized Hosts

Development ◽  
1954 ◽  
Vol 2 (1) ◽  
pp. 14-25
Author(s):  
Petar N. Martinovitch
Keyword(s):  
Pituitary Gland ◽  
Chicken Embryo ◽  
Anterior Pituitary ◽  
Culture Medium ◽  
Embryo Extract ◽  
Rats And Mice ◽  
Initial Results

Just before the last war we made our first attempts to cultivate the anterior pituitary of rats and mice (Martinovitch, 1940). In those preliminary experiments we used pituitaries of animals 1 to 6 months old, and the culture medium was composed of chicken plasma and chicken embryo extract in equal proportions. The explants were grown by the watchglass technique and the incubating temperature was 33° to 34° C. Under these conditions some of our cultures survived for several months. Two months old cultures were grafted in the anterior eye chamber of normal animals and some of these grafts were successfully established. These initial results, and those obtained by Gaillard (1937,1942), seemed sufficiently promising to justify further research. Earlier work in vitro on the pituitary gland, e.g. the papers of Kasahara (1936), Anderson & Haymaker (1937), and Cutting & Lewis (1938), dealt mainly with unorganized growth.

scholarly journals Mechanism of KLF4 Protection against Acute Liver Injury via Inhibition of Apelin Signaling

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Weitao Ji ◽  
Hongyun Shi ◽  
Hailin Shen ◽  
Jing Kong ◽  
Jiayi Song ◽  
...  
Keyword(s):  
Liver Injury ◽  
Signaling Pathway ◽  
Acute Liver Injury ◽  
Control Group ◽  
Necrosis Factor Alpha ◽  
Protein Levels ◽  
Klf4 Expression ◽  
Mrna And Protein Expression

Krüppel-like factor 4 (KLF4) is a key transcription factor that regulates genes involved in the proliferation or differentiation in different tissues. Apelin plays roles in cardiovascular functions, metabolic disease, and homeostatic disorder. However, the biological function of apelin in liver disease is still ongoing. In this study, we investigated the mechanism of KLF4-mediated protection against acute liver injury via the inhibition of the apelin signaling pathway. Mice were intraperitoneally injected with carbon tetrachloride (CCl4; 0.2 mL dissolved in 100 mL olive oil, 10 mL/kg) to establish an acute liver injury model. A KLF4 expression plasmid was injected through the tail vein 48 h before CCl4 treatment. In cultured LX-2 cells, pAd-KLF4 or siRNA KLF4 was overexpressed or knockdown, and the mRNA and protein levels of apelin were determined. The results showed that the apelin serum level in the CCl4-injected group was higher than that of control group, and the expression of apelin in the liver tissues was elevated while KLF4 expression was decreased in the CCl4-injected group compared to the KLF4-plasmid-injected group. HE staining revealed serious hepatocellular steatosis in the CCl4-injected mice, and KLF4 alleviated this steatosis in the mice injected with KLF4 plasmid. In vitro experiments showed that tumor necrosis factor-alpha (TNF-α) could downregulate the transcription and translation levels of apelin in LX-2 cells and also upregulate KLF4 mRNA and protein expression. RT-PCR and Western blotting showed that the overexpression of KLF4 markedly decreased basal apelin expression, but knockdown of KLF4 restored apelin expression in TNF-α-treated LX-2 cells. These in vivo and in vitro experiments suggest that KLF4 plays a key role in inhibiting hepatocellular steatosis in acute liver injury, and that its mechanism might be the inhibition of the apelin signaling pathway.

scholarly journals Inflamma-miR-21 Negatively Regulates Myogenesis during Ageing

Antioxidants ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 345 ◽  
Author(s):  
Maria Borja-Gonzalez ◽  
Jose C. Casas-Martinez ◽  
Brian McDonagh ◽  
Katarzyna Goljanek-Whysall
Keyword(s):  
Skeletal Muscle ◽  
Satellite Cells ◽  
Muscle Regeneration ◽  
Muscle Hypertrophy ◽  
Myogenic Potential ◽  
Age Related ◽  
Primary Myoblasts ◽  
Muscle Homeostasis ◽  
Old Mice

Ageing is associated with disrupted redox signalling and increased circulating inflammatory cytokines. Skeletal muscle homeostasis depends on the balance between muscle hypertrophy, atrophy and regeneration, however during ageing this balance is disrupted. The molecular pathways underlying the age-related decline in muscle regenerative potential remain elusive. microRNAs are conserved robust gene expression regulators in all tissues including skeletal muscle. Here, we studied satellite cells from adult and old mice to demonstrate that inhibition of miR-21 in satellite cells from old mice improves myogenesis. We determined that increased levels of proinflammatory cytokines, TNFα and IL6, as well as H2O2, increased miR-21 expression in primary myoblasts, which in turn resulted in their decreased viability and myogenic potential. Inhibition of miR-21 function rescued the decreased size of myotubes following TNFα or IL6 treatment. Moreover, we demonstrated that miR-21 could inhibit myogenesis in vitro via regulating IL6R, PTEN and FOXO3 signalling. In summary, upregulation of miR-21 in satellite cells and muscle during ageing may occur in response to elevated levels of TNFα and IL6, within satellite cells or myofibrillar environment contributing to skeletal muscle ageing and potentially a disease-related decline in potential for muscle regeneration.

286 INFLUENCE OF REVERSIBLE MEIOSIS INHIBITION ON SWINE EMBRYOS PRODUCED BY IN VITRO FERTILIZATION AND PATHENOGENETIC ACTIVATION

2006 ◽  
Vol 18 (2) ◽  
pp. 250
Author(s):  
M. G. Marques ◽  
A. B. Nascimento ◽  
V. P. Oliveira ◽  
A. R. S. Coutinho ◽  
M. E. O. A. Assumpção ◽  
...  
Keyword(s):  
Culture Medium ◽  
Cell Number ◽  
Control Group ◽  
Cleavage Rate ◽  
Vitro Fertilization ◽  
Electric Pulses ◽  
Cell Numbers ◽  
Blastocyst Rate ◽  
And Control

The present work evaluated the reversible meiosis inhibition effect on the development of swine embryos produced by in vitro fertilization (IVF) or parthenogenetic activation (PA). The efficiency of PZM3 and NCSU23 embryo culture media was also evaluated. Oocytes from ovaries collected at a slaughterhouse were subjected to IVM in two different groups: CHX (cycloheximide 5 µM for 10 h) and control, both with TCM-199 + 3.05 mM glucose + 0.91 mM sodium pyruvate + 10% porcine follicular fluid (pFF) + 0.57 mM cystein + 10 ng epidermal growth factor (EGF)/mL + 10 IU eCG/mL + 10 IU hCG/mL for the initial 22 h. In the remaining period (20 h for CHX and 22 h for control), medium without hormones was utilized. After IVM, oocytes were denuded and fertilized for 6 h (IFV) or the matured oocytes were submitted to activation by electric pulses (PA) (2 DC of 1.5 kV/cm for 30 µs), incubated for 1 h in culture medium with 10 μM of CHX, and again submitted to the same electric pulses for 60 µs. Embryo development was evaluated by cleavage rate on Day 3 and blastocyst rate and blastocyst cell number on Day 7 of culture. Cleavage and blastocyst rates were analyzed by the equality-of-two-ratios test and cell number by the Kruskal-Wallis and Mann-Whitney tests (P < 0.05). In relation to IVF, the PZM3 medium was more efficient than NCSU23 for cleavage rate in the CHX group (PZM3: 68.4%, NCSU23: 44.4%) and had a better blastocyst rate in the control group (PZM3: 13.4%, NCSU23: 5.6%). With reference to PA, NCSU23 presented better cleavage and blastocyst rates than PZM3 in the CHX group (NCSU23: 89.5%, PZM3: 78.5% and NCSU23: 20.4%, PZM3: 13.0%, respectively). In the control group, only the NCSU23 blastocyst rate was higher than that for PZM3 (NCSU23: 22.5%, PZM3: 10.8%). No culture medium effect on cell number mean of IVF and PA blastocysts was observed. Maturation block improved cleavage rates in IVF groups cultured with PZM3 (68.4% and 50.6%, respectively, for CHX and control) and in PA groups cultured with NCSU23 (89.5% and 80.3%, respectively, for CHX and control), but no improvement of blastocyst rates in both groups (IVF and PA) was verified. Table 1 below shows that maturation block decreased the IVF and increased the PA blastocyst cell numbers. As older oocytes are more effectively activated, oocytes blocked with CHX achieved the maturation stage faster than the control group, therefore resulting in high-quality PA blastocysts. In conclusion, PZM3 was more efficient for IVF embryo production in contrast to NCSU23, whereas NCSU23 can be indicated for PA embryo production. Moreover, maturation blockage with CHX influenced blastocyst cell number, decreasing in IVF embryos and increasing in PA embryos. Table 1. Mean (±SD) of blastocyst cell numbers for IVF or PA groups after in vitro maturation without (control) or with cycloheximide (CHX) and cultured in NCSU23 or PZM3 medium This work was supported by FAPESP 02/10747–1.

Sign in / Sign up

Export Citation Format

Share Document