Supplementation Of Culture Medium Research Articles

Effects of Melatonin, GM-CSF, IGF-1, and LIF in Culture Media on Embryonic Development: Potential Benefits of Individualization.

The implantation of good-quality embryos to the receptive endometrium is essential for successful live birth through in vitro fertilization (IVF). The higher the quality of embryos, the higher the live birth rate per cycle, and so efforts have been made to obtain as many high-quality embryos as possible after fertilization. In addition to an effective controlled ovarian stimulation process to obtain high-quality embryos, the composition of the embryo culture medium in direct contact with embryos in vitro is also important. During embryonic development, under the control of female sex hormones, the fallopian tubes and endometrium create a microenvironment that supplies the nutrients and substances necessary for embryos at each stage. During this process, the development of the embryo is finely regulated by signaling molecules, such as growth factors and cytokines secreted from the epithelial cells of the fallopian tube and uterine endometrium. The development of embryo culture media has continued since the first successful human birth through IVF in 1978. However, there are still limitations to mimicking a microenvironment similar to the reproductive organs of women suitable for embryo development in vitro. Efforts have been made to overcome the harsh in vitro culture environment and obtain high-quality embryos by adding various supplements, such as antioxidants and growth factors, to the embryo culture medium. Recently, there has been an increase in the number of studies on the effect of supplementation in different clinical situations such as old age, recurrent implantation failure (RIF), and unexplained infertility; in addition, anticipation of the potential benefits from individuation is rising. This article reviews the effects of representative supplements in culture media on embryo development.

MicroRNAs associated with the quality of follicular fluids affect oocyte and early embryonic development.

Oocyte and embryo quality differs significantly among individuals. Follicular fluid (FF) is a solo environment of oocyte maturation and may flux into the oviduct. Supplementation of invitro maturation (IVM) and culture (IVC) medium with extracellular vesicles of FFs supports oocyte maturation and embryonic development. We addressed a hypothesis that miRNA profiles in FFs are crucial background of oocyte maturation and embryonic development. FFs were collected from the ovaries of individual cows, and the FFs were classified into Good or Poor FF based on the developmental rate to the blastocyst stage of enclosed oocytes. miRNAs associated with the Good FFs were explored using small RNA sequencing. In addition, FFs were classified using the concentration of Good-FF-associated miRNAs. These classified FFs or miRNA were added to the IVM or IVC mediums. Supplementation of IVM and IVC medium with Good FF improved embryonic development. Good FFs contained miR-151-3p and miR-425-5p at a high concentration compared with those in Poor FFs. FFs selected by the concentration of miR-151-3p and miR-425-5p improved oocyte maturation and embryonic development. Supplementation of IVM or IVC medium with either miR-151-3p or miR-425-5p improved embryonic development to the blastocyst stage. miRNAs were associated with the Good FFs determined oocyte maturation and embryonic development.

Optimization of Some Culture Conditions and Medium Composition for the Production of Bacitracin from Lactobacillus helveticus DF

The objective of this study was to evaluate the effects of some culture conditions and components of the culture medium on the production of bacteriocin from Lactobacillus helveticus DF to get maximum bacteriocin yield. The results showed that the best culture conditions for maximum bacteriocin production were; pH 6, incubation temperature 37°C, incubation period 48 hours and the best culture medium MRS at anaerobic condition. Optimization of the medium ingredients was also investigated.The production of active antimicrobial peptide bacteriocin, active against Gram positive and Gram negative microorganisms including P.aeruginosa (B) and E. Feacalis (A), by L. helveticus DF isolated from feces of infants is influenced by complex nitrogen sources and carbon sources in the production medium. Medium components, especially pepton, beef extract and yeast extract, and their concentration contributed to increase in the production during the stationary phase. The Optimal nitrogen sources for their production were 1.5% , 2%, 2% (w/v) of pepton, beef extract and yeast extract respectively, accordingly, the present result evidenced that the increment in bacteriocin production was attributed with nitrogen source. Bacteriocin production by L. helveticus DF emphasized that, the higher bacteriocin yield was attained in the medium supplied with Tween 80 compared to medium without addition of tween 80, optimal concentration for their production 0.75%. Carbon source supplementation in culture media favored the maximum bacteriocin yield by L. helveticus DF, maximum bacteriocin yield when cultured in MRS supplemented with glucose and mannose (2%w/v). Bacteriocin production was affected differently by the presence of different concentrations of K2HPO4 or KH2PO4. Optimal bacteriocin production was recorded in the presence of 0.25% K2HPO4. The results of this study showed that the best production medium was named modified MRS medium which composed of 2% glucose, 2% mannose, 1.5% peptone, 2% yeast extract, 2% beef extract, 0.25% K2HPO4 and 0.75% tween 80, adjusted at pH 6 and incubated at 37°C for 48 hours (under anaerobic conditions) and with inoculum size of 4 % of cells number 1×109 cell/ml. After optimization, Bacteriocin activity was increase to 80 and 160 AU\ML against E. Feacalis (A) and P.aeruginosa (B) respectively.

High throughput production and characterization of primary rat hepatocyte spheroids

Developing in vitro models to explore cell biology and physiology holds significant importance in biotechnology, cancer research, drug discovery, toxicity testing, and the emerging fields of tissue engineering and regenerative medicine. The conventional two-dimensional (2D) approaches of mammalian cell culture (2D) have limitations in replicating all of the mechanical and biochemical signals in vivo. Culturing cells as spheroids provides a three-dimensional environment that more accurately mimics physiological conditions compared to 2D culture. In this study, we established a method for high-throughput spheroid formation using primary rat hepatocytes (PRHs). Using a high throughput platform, we investigated the effect of varying concentrations of cell culture media supplements on spheroid formation. Additionally, we assessed different cell seeding densities and characterized their properties for 7 days. The average diameter and circularity of PRH spheroids remained stable on days 2, 4, and 7, regardless of the initial seeding density. Moreover, PRH spheroids demonstrated high viability (> 90 %) for up to 7 days across all seeding cell densities. Our results demonstrated that this technique enables straightforward, large scale, consistent, and repeatable spheroid manufacturing and presents an alternative approach for future applications.

85 Impact of nobiletin supplementation of culture medium on gene expression patterns of in vitro-produced pig embryos

85 Impact of nobiletin supplementation of culture medium on gene expression patterns of in vitro-produced pig embryos

Towards standardized human platelet lysate production in Europe: An initiative of the European Blood Alliance.

Human platelet lysate (hPL) is a supplement for cell culture media that can be derived from platelet concentrates. As not-for-profit blood establishments, we endorse the evolution of maximally exploiting the potential of donated blood and its derived components, including platelets. The decision to use platelet concentrates to supply hPL as a cell culture supplement should align with the principles and values that blood establishments hold towards the use of donated blood components in transfusion. As a consequence, questions on ethics, practical standardization of hPL production and logistics as well as on assuring hPL quality and safety need careful consideration. We therefore propose an opinion on some of these matters based on available literature and on discussions within the proceedings of the Working Group on Innovation and New Products of the European Blood Alliance. In addition, we propose collaboration among European blood establishments to streamline efforts of hPL supply to maximize the potential of hPL and its application in the wider field of medicine.

Optimisation of hormonal treatment to improve follicular development in one-day-old mice ovaries cultured under in vitro condition.

Base medium containing knock-out serum replacement (KSR) has been found to support formation and maintenance of follicles in one-day-old mice ovaries, but has not been shown to properly support activation and growth of primordial follicles. The present study was conducted to tailor the hormonal content of base medium containing KSR to enhance development of primordial follicles in neonatal ovaries. One-day-old mice ovaries were initially cultured with base medium for four days, and then, different hormonal treatments were added to the culture media and the culture was proceeded for four additional days until day eight. Ovaries were collected for histological and molecular assessments on days four and eight. In experiment I, the main and interactive effects of FSH and testosterone were investigated and FSH promoted activation of primordial follicles and development of primary and preantral follicles, and upregulated genes of phosphoinositide 3-kinase (Pi3k ), KIT ligand (Kitl ), growth differentiation factor 9 (Gdf9 ) and follicle stimulating hormone receptor (Fshr ) (P Bmp15 ), Connexin-43 (Cx43 ) and luteinising hormone and choriogonadotropin receptor (Lhcgr ) (P P Lhcgr (P P >0.05). Supplementation of culture medium containing KSR with gonadotropins, particularly hMG, could improve follicular growth and expression of factors regulating follicular development. This study was a step forward in formulating an optimal medium for development of follicles in cultured one-day-old mice ovaries.

Seed priming with plant growth-promoting rhizobacteria and supplementation of culture medium with biochar alleviated salinity damages in Prosopis koelziana seedlings

The Prosopis koelziana genus, a member of the Fabaceae family, is crucial to the ecology and economy of arid areas. It is commonly used to restore deserts, forests, and degraded soil with low fertility and high salt concentration. However, recent studies found that the seedlings of P. koelziana are highly susceptible to salinity stress despite having a high tolerance to it. To address this issue, an experiment was conducted to determine if plant growth-promoting rhizobacteria (PGPR) pretreatment and biochar application could enhance the seedlings’ tolerance to saline conditions. The research showed that salinity stress decreased seedling growth but increased proline content, soluble sugar content, and antioxidant enzyme activity. Na+ ions increased in plants under salt stress, while K+ ions decreased. The results indicated that the pretreatment of plants with biochar and bacteria increased the fresh weight of seedlings by about 23%, 58%, and 50% in 200, 400, and 600 mM NaCl, respectively, compared with non-pretreated plants. Biochar and bacteria application also increased the proline content by about 62%, 46%, 32%, and 45% in 0, 200, 400, and 600 mM NaCl. This experiment showed that bacteria and biochar pretreatment of plants reduced oxidative stress and sodium uptake (20%, 28%, and 10% decrement of root Na uptake in 200, 400, and 600 mM NaCl). Therefore, these two bio stimulators should be used together for more significant effects on the salinity mitigation of P. Koelziana seedlings and the production of resistant seedlings that have a high ability to transfer to the field.

Improving genetic engineering of adoptive cell therapies by a novel culture media supplement

Improving genetic engineering of adoptive cell therapies by a novel culture media supplement

Scalable expansion of human pluripotent stem cells under suspension culture condition with human platelet lysate supplementation.

The large-scale production of human pluripotent stem cells (hPSCs), including both embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs), shows potential for advancing the translational realization of hPSC technology. Among multiple cell culture methods, suspension culture, also known as three-dimensional (3D) culture, stands out as a promising method to fulfill the large-scale production requirements. Under this 3D culture condition, cell expansion and the preservation of pluripotency and identity during long-term culture heavily relies on the culture medium. However, the xenogeneic supplements in culture medium remains an obstacle for the translation of cell and gene therapy applications from bench to bedside. Here, we tested human platelet lysate (hPL), a xeno-free and serum-free biological material, as a supplement in the 3D culture of hPSCs. We observed reduced intercellular variability and enhanced proliferation in both hESC and hiPSC lines. These cells, after extended culture in the hPL-supplemented system, maintained pluripotency marker expression, the capacity to differentiate into cells of all three germ layers, and normal karyotype, confirming the practicability and safety of hPL supplementation. Furthermore, through RNA-sequencing analysis, we found an upregulation of genes associated with cell cycle regulations in hPL-treated cells, consistent with the improved cellular division efficiency. Taken together, our findings underscore the potential of hPL as a xeno-free and serum-free supplement for the large-scale production of hPSCs, which holds promise for advancing clinical applications of these cells.

Evaluation of Mitochondrial Function on Pyruvate Dehydrogenase Complex Deficient Patient-derived Cell Lines.

Pyruvate Dehydrogenase Complex (PDC) is a pivotal gatekeeper between cytosolic glycolysis and mitochondrial oxidative phosphorylation, playing important role in aerobic energy metabolism. Most PDC deficiency, cases being caused by mutations in PDHA1 encoding the α subunit of the rate-limiting E1 enzyme, which is characterized by abnormal phenotypes caused by energy deprivation at peripheral/central nervous systems and muscular tissues. This study aims to evaluate the potential therapeutic effect of arginine and thiamine in ameliorating mitochondrial function in patient-derived cultured cells. PDC-deficient cell lines, carrying three different PDHA1 variants, were cultured in the absence and presence of arginine and/or thiamine at therapeutical levels, 4 mM and 100 μM, respectively. Mitochondrial bioenergetics profile was evaluated using the Seahorse extracellular flux analyzer. In physiological conditions, control cells presented standard values for all parameters evaluating the mitochondrial function, no differences being observed after supplementation of culture medium with therapeutic levels of arginine and/or thiamine. However, PDC-PDHA1 deficient cell lines consumed less oxygen than the control cells, but arginine and thiamine supplementation increased the basal respiration for values similar or higher than the control cell line. Moreover, arginine and thiamine treatment highlighted an inefficient oxidative phosphorylation carried out by PDC-deficient cell lines. Finally, this treatment showed an increased oxygen consumption by enzymes other than those in the respiratory chain, thus proving the dependence of these mutant cell lines on cytosolic sources for ATP production, namely glycolysis. This study showed that arginine and thiamine, at therapeutical levels, increase the basal oxygen consumption rate of PDC-deficient cell lines, as well as their ATP-linked respiration. This parameter measures the capacity of the cell to meet its energetic demands and, therefore, its increase reveals a higher electron flow through the respiratory chain, which is coupled to elevated oxidative phosphorylation, thus indicating an overall increased robustness in mitochondrial- related bioenergetics.

Nutrient supplementation of culture media improves the detection of Cutibacterium acnes in platelet components by an automated culture system.

Platelet concentrates (PCs) contaminated with Cutibacterium acnes are often transfused prior to detection by the BACT/ALERT system. Though C. acnes is implicated in mild transfusion reactions, delayed clinical effects are unknown. This study assessed the ability to enhance C. acnes detection by supplementing culture media with Tween 80 (T80, an oleic acid source) and a commercial nutrient supplement. Anaerobic culture bottles (BPN) were supplemented with T80 or oleic acid. T80-supplemented BPN bottles were inoculated with four C. acnes isolates (10 or 100 colony-forming units [CFU]/bottle) or other transfusion-relevant bacteria (10 CFU/bottle). Samples of plasma containing SSP+ (platelet additive solution [PAS]) (PAS-plasma) at different concentrations, plasma-PCs and PAS-PCs, spiked with two C. acnes isolates (10 CFU/bottle), were inoculated into T80-supplemented BPN bottles. Furthermore, plasma-PCs were spiked with C. acnes and tested in BPN bottles supplemented with the BD Difco Supplement VX (BDVx). Bottles were incubated in the BACT/ALERT system and times to detection (TtoD) were compared (N = 3). A reduction in TtoD of ≤3.5 days was observed for C. acnes in T80-supplemented BPN, while other species did not show the same effect. However, false positives were observed when T80-supplemented BPN was inoculatedwith PAS-plasma (except for 70% PAS:30% plasma), plasma-PCs or PAS-PCs. Oleic acid supplementation also resulted in false positives. Interestingly, BDVx-supplemented BPN reduced the TtoD of C. acnes in PCs by ≤1.2 days (p < 0.05), with no false-positive results. BDVx supplementation for detection of C. acnes from PCs could result in timely unit retrieval, preventing the transfusion of contaminated products. In clinical settings, T80 supplementation could significantly enhance C. acnes detection from non-blood-derived samples.

In Vitro Plant Growth Promoting Effect of Fucoidan Fractions of Turbinaria decurrens for Seed Germination, Organogenesis, and Adventitious Root Formation in Finger Millet and Eggplant

Bioactive ingredients derived from brown algae have been extensively used in the food, medicine, and cosmetic industries. In this study, fucoidans of low and high molecular fractions (LMF and HMF) extracted and isolated from brown alga Turbinaria decurrens were analyzed for their efficacy on seed germination, seedling growth, callus induction, direct organogenesis, and adventitious root formation in eggplant and finger millet. The yield and sugar content of LMF were higher than HMF. FTIR confirmed that the isolated fractions containing fucoidan has more sulfate groups in HMF than in LMF. The results showed an enhanced seed germination and seedling growth in both crops. In eggplant, 1 mg/L LMF treatment showed the maximum germination (91.6%), whereas, in finger millet, 0.1 and 0.5 mg/L LMF recorded a substantial increase in germination percentage (41.6 and 46%). Maximum fresh weight (FW) was noted with 1.0 mg/L LMF, and 1.0 mg/L LMF and 0.5 mg/L HMF showed maximum dry weight (DW) in eggplant. In finger millet, maximum DW was observed in 0.5 mg/L LMF and 1.0 mg/L HMF. Maximum biomass was noted in the 0.1 mg/L LMF treated group in the case of callus growth in eggplant. Similarly, the shoot tip initiation, proliferation, and plantlet regeneration were significantly improved with fucoidan LMF (0.1 mg/L). In conclusion, fucoidan extracted from T. decurrens exhibiting natural growth promoter property is reported for the first time in this study. These fucoidan fractions, LMF and HMF, can be utilized as cost-effective supplements in plant tissue culture media replacing the commercial PGRs for micropropagation.

Longitudinal characterization of TK6 cells sequentially adapted to animal product-free, chemically defined culture medium: considerations for genotoxicity studies.

Introduction: In vitro approaches are an essential tool in screening for toxicity of new chemicals, products and therapeutics. To increase the reproducibility and human relevance of these in vitro assessments, it is advocated to remove animal-derived products such as foetal bovine serum (FBS) from the cell culture system. Currently, FBS is routinely used as a supplement in cell culture medium, but batch-to-batch variability may introduce inconsistency in inter- and intra-lab assessments. Several chemically defined serum replacements (CDSR) have been developed to provide an alternative to FBS, but not every cell line adapts easily and successfully to CDSR-supplemented medium, and the long-term effect on cell characteristics remains uncertain. Aim: The aim of this study was to adapt the TK6 cell line to animal-product free CDSR-supplemented medium and evaluate the long-term effects on cell health, growth, morphology, phenotype, and function. This included a provisional assessment to determine the suitability of the transitioned cell line for standardised genotoxicity testing using the "in vitro mammalian cell micronucleus test" (OECD TG 487). Materials and methods: Gradual adaptation and direct adaptation methodologies were compared by assessing the cell proliferation, size and viability every passage until the cells were fully adapted to animal-free CDSR. The metabolic activity and membrane integrity was assessed every 4-8 passages by PrestoBlue and CytoTox-ONE™ Homogeneous Membrane Integrity Assay respectively. A detailed morphology study by high content imaging was performed and the expression of cell surface markers (CD19 and CD20) was conducted via flow cytometry to assess the potential for phenotypic drift during longer term culture of TK6 in animal-free conditions. Finally, functionality of cells in the OECD TG 487 assay was evaluated. Results: The baseline characteristics of TK6 cells cultured in FBS-supplemented medium were established and variability among passages was used to set up acceptance criteria for CDSR adapted cells. TK6 were adapted to CDSR supplemented medium either via direct or gradual transition reducing from 10% v/v FBS to 0% v/v FBS. The cell growth rate was compromised in the direct adaptation and therefore the gradual adaptation was preferred to investigate the long-term effects of animal-free CDSR on TK6 cells. The new animal cells showed comparable (p > 0.05) viability and cell size as the parent FBS-supplemented cells, with the exception of growth rate. The new animal free cells showed a lag phase double the length of the original cells. Cell morphology (cellular and nuclear area, sphericity) and phenotype (CD19 and CD20 surface markers) were in line (p > 0.05) with the original cells. The new cells cultured in CDSR-supplemented medium performed satisfactory in a pilot OECD TG 487 assay with compounds not requiring metabolic activation. Conclusion: TK6 cells were successfully transitioned to FBS- and animal product-free medium. The new cell cultures were viable and mimicked the characteristics of FBS-cultured cells. The gradual transition methodology utilised in this study can also be applied to other cell lines of interest. Maintaining cells in CDSR-supplemented medium eliminates variability from FBS, which in turn is likely to increase the reproducibility of in vitro experiments. Furthermore, removal of animal derived products from cell culture techniques is likely to increase the human relevance of in vitro methodologies.

P-191 The impact of resveratrol supplementation as antioxidant on embryonic development in older women (over 40years) undergoing IVF

Abstract Study question Does the resveratrol supplementation in human embryo culture medium improve embryo quality and pregnancy outcomes in older women? Summary answer Resveratrol significantly improved good quality embryo rates. These results are thought to be due to the biological activities of resveratrol as an antioxidant. What is known already Reproductive aging involves age-related declines in ovarian function and development capacity. After initiating meiosis, germ cells arrest at the dictyate stage of prophase I in the ovary for 10-50 years in women. Accumulated reactive oxygen species (ROS) affect oocytes adversely. Antioxidants reduce ROS-induced damage to oocytes and resveratrol is an important antioxidant polyphenolic compound. Resveratrol has positive effects on fertility, it improves the development of blastocysts in aged animal and it could enhance the maturation and quality of oocytes from aged women. However, there are just a few studies there are clarified relationship between resveratrol and embryos in older women. Study design, size, duration The study retrospectively included 966 patients with aged over 40 years to evaluate the impact of resveratrol on embryo quality and IVF pregnancy outcomes between January 2019 and December 2021. Participants/materials, setting, methods The oocytes from aged female (over 40years) were divided into 2 groups, fertilized and cultured in MRC media (MRC#D13 (day 1 ∼ 3) and MRC#D46 (day 4 ∼ 6 culture medium); Maria Medical Foundation, South Korea) in the presence or absence of resveratrol (0.5 μM; Sigma-Aldrich, USA). Fertilization, good quality embryo, clinical pregnancy, and implantation rate after embryo transfer were compared among between the two groups. Data analyzed using the SPSS and T-test p value &amp;lt;0.05 considered statistically significant. Main results and the role of chance There was no significant difference in female age (42.5 ± 2.2 vs. 42.3 ± 2.0, P = 0.358), No. of previous ART cycles (3.6 ± 2.3 vs. 3.7 ± 2.5, P = 0.680), thickness of endometrium (9.2 ± 1.5 vs. 9.4 ± 1.4, P = 0.312), maturation rate per oocytes (74.6% vs. 75.8%, P = 0.290), fertilization rate (81.5% vs. 80.0%, P = 0.165), and No. of transferred embryos (2.4 ± 0.6 vs. 2.4 ± 0.6, P = 0.818). However, good quality embryo rate (44.0% vs. 36.4%, p &amp;lt; 0.05) in the resveratrol group was 7.6% higher than the control group. Pregnancy rate (26.8% vs. 22.7%, P = 0.167) and implantation rate (11.9% vs. 9.9%, P = 0.131) were slightly higher than control group but this was not statistically significant. Limitations, reasons for caution This is a retrospective study that generates strong conclusions, but it could be interesting to test it in a prospective study. The study was conducted at a single IVF center. Embryo transfer result is based on implantation rate, while the live birth data for all pregnancies are not yet available. Wider implications of the findings This is the first study to investigate the effects of resveratrol on the embryonic developmental competence and pregnancy potential of older women (over 40years). Trial registration number not applicable

Surfactants reduce aggregation of monoclonal antibodies in cell culture medium with improvement in performance of mammalian cell culture.

Therapeutic monoclonal antibodies (mAbs) are biologics produced using mammalian cells and represent an important class of biotherapeutics. Aggregation in mAbs is a major challenge that can be mitigated by rigorous and reproducible upstream and downstream approaches. The impact of frequently used surfactants, like polysorbate 20, polysorbate 80, poloxamer 188, and 2-hydroxypropyl-beta-cyclodextrin, on aggregation of mAbs during cell culture was investigated in this study. Their impact on cell proliferation, viability, and mAb titer was also investigated. Polysorbate 20 and polysorbate 80 at the concentration of 0.01 g/L and poloxamer 188 at the concentration of 5 g/L were found to be effective in reducing aggregate formation in cell culture medium, without affecting the cell growth or viability. Furthermore, their presence in culture media resulted in increased cell proliferation as compared to the control group. Addition of these surfactants at the specified concentrations increased monomer production while decreasing high molecular weight species in the medium. After mAbs were separated, using protein "A" chromatography, flasks with surfactant exhibited improved antibody stability, when analyzed by DLS. Thus, while producing aggregation-prone mAbs via mammalian cell culture, these excipients may be employed as cell culture medium supplements to enhance the quality and yield of functional mAbs.

Expression of Apoptosis-Related Genes in Cultured Ovarian Tissue of Mouse in Presence of Coenzyme Q10.

Oxidative stress affects the development of ovarian follicles during in vitro culture thus applying an antioxidant is necessary. The aim of this study was to investigate the effect of coenzyme Q10 (CoQ10) on the expression of apoptosis-related genes of mouse ovaries during in vitro culture. The immature mouse ovaries were cultured in the presence of 50 μM CoQ10 for 7 days. Histological examinations were performed and the 17 beta-estradiol concentration was measured on the seventh day of culture. The relative expression of Caspase 3, Bax, Bad, and Bcl 2 genes were investigated by real-time RT-PCR. The rates of normal follicles in the presence of CoQ10 was significantly increased compared to the control group. Also, in CoQ10-trated group a significant increase in the level of 17 beta-estradiol was seen compared to the control group. The mRNA expression of anti-apoptotic gene Bcl2 was significantly increased while the expression of pro-apoptotic genes (Caspase3, Bax and Bad) significantly declined in CoQ10 treated group compared to those of control group. The supplementation of the ovarian culture media with CoQ10 improved the follicular development through alteration in expression of apoptosis-related genes and stimulated the production of estradiol.

Novel Stimulants of Medicinal Basidiomycetes Growth Based on Nanoparticles of N-monosubstituted Amino Acid Derivatives of Fullerene C60

The influence of nanoparticles of hydrated C60 fullerene and its N-monoamino acid derivatives on the oxidative metabolism and growth of the mycelial biomass of basidiomycetes during their submerged cultivation was studied. It was found that the supplementation of culture media with nanoparticles of the studied compounds at their final concentration range of 10−7 to 10−11 M significantly increased the resulting biomass, while the severity of the effect in this concentration range changed slightly. That prompted the use of nanomolar concentrations of compounds as reasonable. The most pronounced stimulating effect (an increase in biomass of about 240% with respect to control) was observed when culturing Laetiporus sulphureus, the intrinsically high level of oxidative metabolism of which was significantly lowered by the presence of the studied additives. It was shown that the growth-enhancing action of nanoparticles of fullerene C60 and its derivatives could not be attributed to photochemical reactions, particularly fullerene photoexcitation. Fullerene and its derivatives manifest a growth regulatory effect on bio-objects from different kingdoms of the living world (plants and fungi), which is indicative of these compounds’ mechanism of action based on a direct impact on fundamental, universal for all living beings, biophysical processes, primarily chain free-radical oxidation.

Determination of Vaccine Immunogenicity Using Bovine Monocyte-Derived Dendritic Cells

Dendritic cells (DCs) are the most potent antigen-presenting cells (APCs) within the immune system. They patrol the organism looking for pathogens and play a unique role within the immune system by linking the innate and adaptive immune responses. These cells can phagocytize and then present captured antigens to effector immune cells, triggering a diverse range of immune responses. This paper demonstrates a standardized method for the in vitro generation of bovine monocyte-derived dendritic cells (MoDCs) isolated from cattle peripheral blood mononuclear cells (PBMCs) and their application in evaluating vaccine immunogenicity. Magnetic-based cell sorting was used to isolate CD14+ monocytes from PBMCs, and the supplementation of complete culture medium with interleukin (IL)-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) was used to induce the differentiation of CD14+ monocytes into naive MoDCs. The generation of immature MoDCs was confirmed by detecting the expression of major histocompatibility complex II (MHC II), CD86, and CD40 cell surface markers. A commercially available rabies vaccine was used to pulse the immature MoDCs, which were subsequently co-cultured with naive lymphocytes. The flow cytometry analysis of the antigen-pulsed MoDCs and lymphocyte co-culture revealed the stimulation of T lymphocyte proliferation through the expression of Ki-67, CD25, CD4, and CD8 markers. The analysis of the mRNA expression of IFN-γ and Ki-67, using quantitative PCR, showed that the MoDCs could induce the antigen-specific priming of lymphocytes in this in vitro co-culture system. Furthermore, IFN-γ secretion assessed using ELISA showed a significantly higher titer (**p < 0.01) in the rabies vaccine-pulsed MoDC-lymphocyte co-culture than in the non-antigen-pulsed MoDC-lymphocyte co-culture. These results show the validity of this in vitro MoDC assay to measure vaccine immunogenicity, meaning this assay can be used to identify potential vaccine candidates for cattle before proceeding with in vivo trials, as well as in vaccine immunogenicity assessments of commercial vaccines.

Perspective: Can early embryonic losses be reduced in lactating dairy cows?

Perspective: Can early embryonic losses be reduced in lactating dairy cows?