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Rectal and vaginal temperatures are utilised in both in vivo and in vitro models to study the effects of heat stress on oocyte competence and embryo viability in cattle. However, uterine temperature increases by only 0.5 °C in heat-stressed cows, significantly lower than simulated increases in in vitro models. Temperature variations within oviducts and ovarian follicles during heat stress are poorly understood or unavailable, and evidence is lacking that oocytes and pre-implantation embryos experience mild (40 °C) or severe (41 °C) heat stress inside the ovarian follicle and the oviduct and uterus, respectively. Gathering detailed temperature data from the reproductive tract and follicles is crucial to accurately assess oocyte competence and embryo viability under realistic heat stress conditions. Potential harm from heat stress on oocytes and embryos may result from reduced nutrient availability (e.g., diminished blood flow to the reproductive tract) or other unidentified mechanisms affecting tissue function rather than direct thermal effects. Refining in vivo stress models in cattle is essential to accurately identify animals truly experiencing heat stress, rather than assuming heat stress exposure as done in most studies. This will improve model reliability and aid in the selection of heat-tolerant animals.
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Flavonoids are compounds that result from the secondary metabolism of plants and play a crucial role in plant development and mitigating biotic and abiotic stresses. The highest levels of flavonoids are found in legumes such as soybean. Breeding programs aim to increase desirable traits, such as higher flavonoid contents and vigorous seeds. Soybeans are one of the richest sources of protein in the plant kingdom and the main source of flavonoid derivatives for human health. In view of this, the hypothesis of this study is based on the possibility that the concentration of isoflavones in soybean seeds contributes to the physiological quality of the seeds. The aim of this study was to analyze the content of flavonoids in soybean genotypes and their influence on the physiological quality of the seeds. Seeds from thirty-two soybean genotypes were obtained by carrying out a field experiment during the 2021/22 crop season. The experimental design was randomized blocks with four replications and thirty-two F3 soybean populations. The seeds obtained were subjected to germination, first germination counting, electrical conductivity and tetrazolium vigor and viability tests. After drying and milling the material from each genotype, liquid chromatography analysis was carried out to obtain flavonoids, performed at UPLC level. Data were submitted to analysis of variance and, when significant, the means were compared using the Scott-Knott test at 5% probability. The results found here show the occurrence of genotypes with higher amounts of flavonoids when compared to their peers. The flavonoid FLVD_G2 had the highest concentration and differed from the others. Thus, we can assume that the type and concentration of flavonoids does not influence the physiological quality of seeds from different soybean genotypes, but it does indirectly contribute to viability and vigor, since the genotypes with the highest FLVD_G2 levels had better FGC values. The findings indicate that there is a difference between the content of flavonoids in soybean genotypes, with a higher content of genistein. The content of flavonoids does not influence the physiological quality of seeds, but contributes to increasing viability and vigor.
Assuntos
Flavonoides , Genótipo , Germinação , Glycine max , Sementes , Glycine max/genética , Glycine max/metabolismo , Glycine max/crescimento & desenvolvimento , Sementes/genética , Flavonoides/análise , Flavonoides/metabolismo , Isoflavonas/análise , Isoflavonas/metabolismoRESUMO
Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) control antral follicular growth by regulating several processes, such as the synthesis of hormones and signaling molecules, proliferation, survival, apoptosis, luteinization, and ovulation. To exert these effects, gonadotropins bind to their respective Gs protein-coupled receptors, activating the protein kinase A (PKA) pathway or recruiting Gq proteins to activate protein kinase C (PKC) signaling. Although the action mechanism of FSH and LH is clear, recently, it has been shown that both gonadotropins promote the synthesis of sphingosine-1-phosphate (S1P) in granulosa and theca cells through the activation of sphingosine kinase 1. Moreover, the inhibition of SPHKs reduces S1P synthesis, cell viability, and the proliferation of follicular cells in response to gonadotropins, and the addition of S1P to the culture medium increases the proliferation of granulosa and theca cells without apparent effects on sexual steroid synthesis. Therefore, we consider that S1P is a crucial signaling molecule that complements the canonical gonadotropin pathway to promote the proliferation and viability of granulosa and theca cells.
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Gonadotropinas , Lisofosfolipídeos , Folículo Ovariano , Esfingosina , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Esfingosina/farmacologia , Lisofosfolipídeos/metabolismo , Lisofosfolipídeos/farmacologia , Feminino , Animais , Humanos , Gonadotropinas/metabolismo , Folículo Ovariano/metabolismo , Folículo Ovariano/efeitos dos fármacos , Hormônio Luteinizante/metabolismo , Hormônio Foliculoestimulante/metabolismo , Hormônio Foliculoestimulante/farmacologia , Transdução de Sinais/efeitos dos fármacos , Células da Granulosa/metabolismo , Células da Granulosa/efeitos dos fármacosRESUMO
The objective of this work was to prepare and characterize liposomes containing co-encapsulated ascorbic acid (AA) and ascorbyl palmitate (AP), as well as to evaluate their stability, cytotoxicity, antioxidant, and antimicrobial activity. Through the pre-formulation studies, it was possible to improve the formulation, as leaving it more stable and with a greater antioxidant activity, resulting in a formulation designated LIP-AAP, with 161 nm vesicle size, 0.215 polydispersity index, -31.7 mV zeta potential, and pH of 3.34. Encapsulation efficiencies were 37% for AA and 79% for AP, and the content was 1 mg/mL for each compound. The optimized liposomes demonstrated stability under refrigeration for 60 days, significant antioxidant activity (31.4 µMol of TE/mL), and non-toxicity, but no antimicrobial effects against bacteria and fungi were observed. These findings confirm that the co-encapsulated liposomes are potent, stable antioxidants that maintain their physical and chemical properties under optimal storage conditions.
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Anti-Infecciosos , Antioxidantes , Ácido Ascórbico , Estabilidade de Medicamentos , Lipossomos , Ácido Ascórbico/química , Ácido Ascórbico/farmacologia , Ácido Ascórbico/análogos & derivados , Lipossomos/química , Antioxidantes/química , Antioxidantes/farmacologia , Anti-Infecciosos/farmacologia , Anti-Infecciosos/química , Humanos , Bactérias/efeitos dos fármacos , Tamanho da Partícula , Fungos/efeitos dos fármacos , Fungos/crescimento & desenvolvimento , Composição de MedicamentosRESUMO
PURPOSE: This study aimed to investigate the efficiency of antimicrobial photodynamic therapy (aPDT) on Streptococcus mutans biofilm in the oral cavity using the photosensitizer chloroaluminum phthalocyanine encapsulated in chitosan nanoparticles (ClAlPc/Ch) at three preirradiation times. METHODS: Biofilms of Streptococcus mutans strains (ATCC 25,175) were cultivated on bovine tooth blocks and exposed to a 10% sucrose solution three times a day for 1 min over three consecutive days. The samples were randomly distributed into five treatment groups (n = 5): (I) aPDT with ClAlPc/Ch with a preirradiation time of 5 min (F5), (II) aPDT with ClAlPc/Ch with a preirradiation time of 15 min (F15), (III) aPDT with ClAlPc/Ch with a preirradiation time of 30 min (F30), (IV) 0.12% chlorhexidine digluconate (CHX), and (V) 0.9% saline solution (NaCl). After treatment, the S. mutans biofilms formed on each specimen were collected to determine the number of viable bacteria (colony-forming units (CFU)/mL). Data were analyzed for normality using the Shapiro-Wilk test and the analysis of variance (ANOVA) and Tukey HSD tests to analyze the number of viable bacteria (α = 0.05). RESULTS: The one-way ANOVA showed a difference between the groups (p = 0.0003), and the Tukey HSD posttest showed that CHX had the highest microbial reduction of S. mutans, not statistically different from the F5 and F15 groups, whereas the NaCl group had the lowest microbial reduction statistically similar to the F30 group. CONCLUSION: The results demonstrate that aPDT mediated by ClAlPc/Ch when used at preirradiation times of 5-15 min can be an effective approach in controlling cariogenic biofilm of S. mutans, being an alternative to 0.12% CHX.
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Biofilmes , Quitosana , Nanopartículas , Fotoquimioterapia , Fármacos Fotossensibilizantes , Streptococcus mutans , Streptococcus mutans/efeitos dos fármacos , Streptococcus mutans/efeitos da radiação , Streptococcus mutans/fisiologia , Fotoquimioterapia/métodos , Quitosana/farmacologia , Quitosana/química , Nanopartículas/química , Biofilmes/efeitos dos fármacos , Biofilmes/efeitos da radiação , Animais , Bovinos , Fármacos Fotossensibilizantes/farmacologia , Técnicas In Vitro , Indóis/farmacologia , Boca/microbiologia , Clorexidina/farmacologia , Clorexidina/análogos & derivados , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/efeitos da radiação , Compostos OrganometálicosRESUMO
Background: Leprosy is a chronic infectious disease caused by Mycobacterium leprae, which can lead to a disabling neurodegenerative condition. M. leprae preferentially infects skin macrophages and Schwann cells-glial cells of the peripheral nervous system. The infection modifies the host cell lipid metabolism, subverting it in favor of the formation of cholesterol-rich lipid droplets (LD) that are essential for bacterial survival. Although researchers have made progress in understanding leprosy pathogenesis, many aspects of the molecular and cellular mechanisms of host-pathogen interaction still require clarification. The purinergic system utilizes extracellular ATP and adenosine as critical signaling molecules and plays several roles in pathophysiological processes. Furthermore, nucleoside surface receptors such as the adenosine receptor A2AR involved in neuroimmune response, lipid metabolism, and neuron-glia interaction are targets for the treatment of different diseases. Despite the importance of this system, nothing has been described about its role in leprosy, particularly adenosinergic signaling (AdoS) during M. leprae-Schwann cell interaction. Methods: M. leprae was purified from the hind footpad of athymic nu/nu mice. ST88-14 human cells were infected with M. leprae in the presence or absence of specific agonists or antagonists of AdoS. Enzymatic activity assays, fluorescence microscopy, Western blotting, and RT-qPCR analysis were performed. M. leprae viability was investigated by RT-qPCR, and cytokines were evaluated by enzyme-linked immunosorbent assay. Results: We demonstrated that M. leprae-infected Schwann cells upregulated CD73 and ADA and downregulated A2AR expression and the phosphorylation of the transcription factor CREB (p-CREB). On the other hand, activation of A2AR with its selective agonist, CGS21680, resulted in: 1) reduced lipid droplets accumulation and pro-lipogenic gene expression; 2) reduced production of IL-6 and IL-8; 3) reduced intracellular M. leprae viability; 4) increased levels of p-CREB. Conclusion: These findings suggest the involvement of the AdoS in leprosy neuropathogenesis and support the idea that M. leprae, by downmodulating the expression and activity of A2AR in Schwann cells, decreases A2AR downstream signaling, contributing to the maintenance of LD accumulation and intracellular viability of the bacillus.
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OBJECTIVE: To evaluate the effect of the deterioration of computer aided design/computer aided manufacturing (CAD/CAM) burs during zirconia milling, on surface roughness, contact angle, and fibroblast viability. MATERIALS AND METHODS: Ceramic blocks were milled and 75 ceramic disks (8 × 1.5 mm) made and allocated into three groups (n = 25): G1-brand new 2L and 1L burs, G2-2L bur at the end of lifetime and brand new 1L bur and G3-both burs at the end of their lifetimes. Roughness (Ra, Rq, and Rz) was evaluated using a 3D optical profilometer, the contact angle by the sessile drop method and the cell viability of the mouse NIH/3T3 fibroblast, using the Alamar Blue assay at intervals of 24, 48, and 72 h (ISO 10993-5). Data were analyzed by one-way ANOVA and Kruskal-Wallis tests (p ≤ 0.05). RESULTS: Roughness increased as the burs deteriorated and G3 (0.27 ± 0.04) presented a higher value for Ra (p < 0.001). The highest contact angle was observed in G3 (86.2 ± 2.66) when compared with G1 (63.7 ± 12.49) and G2 (75.3 ± 6.36) (p < 0.001). Alamar Blue indicated an increase in cell proliferation, with no significant differences among the groups at 24 and 72 h (p > 0.05). CONCLUSIONS: The deterioration of the burs increased the surface roughness and decreased the wettability, but did not interfere in cell viability and proliferation. CLINICAL SIGNIFICANCE: The use of custom zirconia abutments represents an effective strategy for single crowns restorations. Our findings suggest that these abutments can be efficiently milled using CAD/CAM burs within their recommended lifetime.
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The high consumption of dietary supplements was a fundamental driver for the creation of the regulatory framework by the Brazilian governmental authorities. However, the regulatory agencies lack official low-cost methodologies to evaluate the quality of food supplements. A preliminary screening method by HPLC-DAD was proposed and validated for screening and quantification of adulterants in dietary supplements. The limits of detection and quantification were <0.11 and 0.37 µg.g-1, respectively. The method was applied for the investigation of ten unauthorized substances (spironolactone, hydrochlorothiazide, furosemide, clenbuterol, testosterone, testosterone propionate, yohimbine, vardenafil, tadalafil, and sildenafil) with a time of analysis of <5 min. Sixteen percent of the 44 samples analyzed had at least one adulterant at or above therapeutic concentrations. Subsequently, in vitro evaluations were performed of the potential cytotoxicity to evaluate the cell viability, DNA damage, determination of nitric oxide levels, and quantification of reactive oxygen species. Despite the necessity of further studies, the results indicate a relationship between the presence of adulterants in food supplements and a potential cytotoxic effect.
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Integrating in vitro embryo production with embryonic microsurgery facilitates the generation of monozygotic twins. However, despite their potential benefits, these methods have not been widely adopted in commercial settings because of their substantial costs. Hence, there is a need to streamline the bisection procedure while ensuring efficient production of viable demi-embryos. In this study, we investigated the impact of different orientations of microsurgical incisions in relation to inner cell mass on embryonic development, morphology, viability, and expression of cell fate protein markers using a simplified microsurgery approach. Ovaries were transported from the slaughterhouse to the laboratory and aspirated to obtain oocytes that were selected and subjected to in vitro embryo production. The selected expanded blastocysts (n = 204) underwent microsurgery. The blastocysts were immobilized to facilitate incision using an adapted microblade, yielding demi-embryos (vertical incision) and viable embryonic fragments (transverse incision). The structures were then re-cultured for 12 h. Viability was assessed by measuring the re-expansion rate after re-culture, followed by immunofluorescence analysis of proteins (CDX2 and NANOG) and apoptosis analysis using terminal deoxynucleotyl transferase dUTP nick end-labeling (TUNEL). Microsurgically derived embryos exhibited remarkable plasticity, as evidenced by a slight reduction (P < 0.05) in the re-expansion rate (transverse 64.2 % and vertical 57.2 %) compared to that of the control group (blastocysts without microsurgery) (86.7 %). They also demonstrated the ability of morphological reconstitution after culturing. Despite the anticipated decrease (P < 0.05) in the total number of cells and embryo volume, microsurgery did not result in a significant increase (P > 0.05) in the number of apoptotic cells. Furthermore, microsurgery led to higher (P < 0.05) expression of markers associated with pluripotency, indicating its efficiency in preserving regenerative capacity. Moreover, microsurgery, whether followed by immunosurgery or not, made the isolation of embryonic cells easier. In conclusion, both transverse and vertical microsurgery incisions enabled the production of identical demi-embryos and served as tools for isolating embryonic cells without compromising the resumption of development and the apoptotic index.
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Técnicas de Cultura Embrionária , Microcirurgia , Animais , Microcirurgia/métodos , Microcirurgia/veterinária , Técnicas de Cultura Embrionária/veterinária , Fertilização in vitro/veterinária , Desenvolvimento Embrionário , Feminino , Embrião de Mamíferos/fisiologia , Blastocisto/fisiologia , Bovinos/embriologiaRESUMO
Amantadine (AMA) is a useful drug in neuronal disorders, but few studies have been performed to access its toxicological profile. Conversely, doxorubicin (Dox) is a well-known antineoplastic drug that has shown neurotoxic effects leading to cognitive impairment. The aims of this study are to evaluate the cytotoxic, genotoxic, and mutagenic effects of AMA, as well as its possible protective actions against deleterious effects of Dox. The Salmonella/microsome assay was performed to assess mutagenicity while cytotoxicity and genotoxicity were evaluated in SH-SY5Y cells using MTT and comet assays. Possible modulating effects of AMA on the cytotoxicity, genotoxicity, and mutagenicity induced by Dox were evaluated through cotreatment procedures. Amantadine did not induce mutations in the Salmonella/microsome assay and decreased Dox-induced mutagenicity in the TA98 strain. AMA reduced cell viability and induced DNA damage in SH-SY5Y cells. In cotreatment with Dox, AMA attenuated the cytotoxicity of Dox and showed an antigenotoxic effect. In conclusion, AMA does not induce gene mutations, although it has shown a genotoxic effect. Furthermore, AMA decreases frameshift mutations induced by Dox as well as the cytotoxic and genotoxic effects of Dox in SH-SY5Y cells, suggesting that AMA can interfere with Dox mutagenic activity and attenuate its neurotoxic effects.