•Melatonin receptors type 1A/B are expressed in bovine preantral and antral follicles.•Melatonin stimulates follicle growth through its membrane receptors.•Luzindole blocks the effects of melatonin ...on follicle growth.•Luzindole blocks the effects of melatonin on expression of antioxidant enzymes.
This study aims to investigate the (1) expression of melatonin receptors types 1A/B (MTNR1A/B) in bovine ovaries and (2) the in vitro effects of melatonin on secondary follicle development, antrum formation, viability, and expression of messenger ribonucleic acid (mRNA) for superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase-1 (GPX1) and peroxiredoxin 6 (PRDX6). The expression of MTNR1A/B in bovine ovarian follicles was demonstrated by immunohistochemistry. To choose the most effective concentration of melatonin on follicular growth and viability, isolated secondary follicles were cultured individually at 38.5°C, with 5% CO2 in air, for 18 d in TCM-199+ alone or supplemented with 10−11, 10−9, 10−7 or 10−5 M melatonin. Then, melatonin receptor antagonist, luzindole, was tested to further evaluate the mechanisms of actions of melatonin, that is, the follicles were cultured in control medium alone or supplemented with 10−7 M melatonin, 10 µM luzindole and both 10−7 M melatonin and 10 µM luzindole. Follicular growth, morphology and antrum formation were evaluated at days 6, 12 and 18. At the end of culture, viability of secondary follicles was analyzed by calcein-AM and ethidium homodimer-1, and the relative levels of mRNA for SOD, CAT, GPX1 and PRDX6 were evaluated by real time polymerase chain reaction. Immunohistochemistry results showed expression of MTNR1A/B in oocyte and granulosa cells of primordial, primary, secondary and antral follicles. Secondary follicles cultured in medium supplemented with melatonin at different concentrations had well preserved follicles after 18 d of culture. Furthermore, follicles cultured in presence of 10−7 M melatonin presented significantly higher diameters than those cultured in other treatments. The presence of melatonin receptor antagonist, luzindole, blocked the effects of melatonin on follicular growth and viability. In addition, follicles cultured in medium containing only melatonin had significantly higher rates of antrum formation. Follicles cultured in medium containing only melatonin had higher relative levels of mRNA for CAT, SOD and PRDX-6 than those cultured with both melatonin and luzindole. Follicles cultured with luzindole only or both melatonin and luzindole had lower relative levels of mRNA for PRDX6 and GPX1 than those cultured control medium. In conclusion, melatonin promotes growth of bovine secondary follicles through its membrane-coupled receptors, while luzindole blocks the effects of melatonin on follicle growth and reduces the expression of antioxidant enzymes in cultured follicles.
There are no reports of the systemic human pathology of the novel swine H1N1 influenza (S-OIV) infection.
The autopsy findings of 21 Brazilian patients with confirmed S-OIV infection are presented. ...These patients died in the winter of the southern hemisphere 2009 pandemic, with acute respiratory failure.
Lung tissue was submitted to virologic and bacteriologic analysis with real-time reverse transcriptase polymerase chain reaction and electron microscopy. Expression of toll-like receptor (TLR)-3, IFN-gamma, tumor necrosis factor-alpha, CD8(+) T cells and granzyme B(+) cells in the lungs was investigated by immunohistochemistry.
Patients were aged from 1 to 68 years (72% between 30 and 59 yr) and 12 were male. Sixteen patients had preexisting medical conditions. Diffuse alveolar damage was present in 20 individuals. In six patients, diffuse alveolar damage was associated with necrotizing bronchiolitis and in five with extensive hemorrhage. There was also a cytopathic effect in the bronchial and alveolar epithelial cells, as well as necrosis, epithelial hyperplasia, and squamous metaplasia of the large airways. There was marked expression of TLR-3 and IFN-gamma and a large number of CD8(+) T cells and granzyme B(+) cells within the lung tissue. Changes in other organs were mainly secondary to multiple organ failure.
Autopsies have shown that the main pathological changes associated with S-OIV infection are localized to the lungs, where three distinct histological patterns can be identified. We also show evidence of ongoing pulmonary aberrant immune response. Our results reinforce the usefulness of autopsy in increasing the understanding of the novel human influenza A (H1N1) infection.
During the last 10 to 15 yr, in vitro research to predict antral follicle growth and oocyte maturation has delivered interesting advances in the knowledge of processes regulating follicle growth and ...developmental competence of oocytes. This review discusses the contribution of cumulus and mural granulosa cells in the process of oocyte maturation and cumulus expansion in cumulus–oocyte complexes (COCs) from follicles of different sizes and shows that differences in gene expression in oocytes, granulosa, and theca cells of small and large follicles impact the success of in vitro blastocyst development. In addition, the molecular mechanisms by which COC metabolism and antioxidant defense provide oocyte competence are highlighted. Furthermore, new insights and perspectives on molecular and cellular regulation of in vitro oocyte maturation are emphasized.
•Role of hormones and growth factors on maturation of different-sized oocytes is discussed.•Maturation stage of granulosa cells is important for oocyte competence.•Cumulus–oocyte complex metabolism and antioxidant defense provide oocyte competence.•Perspectives are given on molecular and cellular regulation for in vitro oocyte maturation.
The aims of this study were to investigate the effects of epidermal growth factor (EGF) and progesterone on the development, viability and the gene expression of bovine secondary follicle culture ...in vitro for 18 days. Secondary follicles (∼0.2 mm) were isolated from ovarian cortex and individually cultured at 38.5 °C, with 5% CO2 in air, for 18 days, in TCM-199+ (n = 63) alone (control medium) or supplemented with 10 ng/mL progesterone (n = 64), 10 ng/mL EGF (n = 61) or both EGF and progesterone (n = 66). The effects of these treatments on growth, antrum formation, viability, ultrastructure and mRNA levels for GDF-9, c-MOS, H1foo and cyclin B1 were evaluated, significantly different (p < 0.05). The results showed that there was a progressive increase in follicular diameter in all treatments, but only follicles cultured in medium supplemented with EGF had increased significantly in diameter when compared to follicles cultured in the control medium at the end of the culture period, significantly different (p < 0.05). A positive interaction between EGF and progesterone was not observed. In addition, the presence of EGF, progesterone or both in culture medium did not influence the rate of follicle survival and antrum formation. However, the presence of only progesterone in cultured medium increased the expression of mRNAs for GDF9 and cyclin B1 in oocytes. EGF also significantly increased the levels of mRNAs for cMOS and GDF9 when compared to follicles cultured in control medium. Ultrastructural analyzes showed that cultured follicles in all treatments maintained the integrity of granulosa cells. In conclusion, the EGF promotes the development of secondary follicles cultured in vitro for 18 days and increases the expression of cMOS and GDF9, while progesterone alone or in association with EGF have not a positive effect on follicular growth. However, progesterone increases the expression of GDF9 and cyclin B1 in oocytes.
•EGF increases follicular growth and the expression of mRNA for GDF9 and cMOS .•Progesterone increases the expression of GDF9 and cyclin B1in cultured follicles.•EGF and progesterone have no synergic effect on bovine preantral follicles in vitro.
•TNF-α system members are expressed at different follicular stages in bovine ovaries.•TNF-α reduces follicular survival in cultured bovine ovarian tissue.•TNF-α increases the total number of ...apoptotic cells in cultured follicles.•Dexamethasone improves ultrastructure of bovine follicle cultured in vitro.
This study was conducted to detect the protein expression of TNF-α system members (TNF-α/TNFR1/TNFR2) in bovine ovarian follicles and to evaluate the effects of TNF-α or dexamethasone on the survival and growth of primordial follicles in vitro, as well as on gene expression in cultured ovarian tissue. It was hypothesized that TNF-α induces follicular atresia in ovarian tissues cultured in vitro, and that dexamethasone suppresses the production of endogenous TNF-α, which can improve follicle viability in vitro. Ovarian fragments were cultured for 6days in α-MEM+ supplemented with TNF-α (0, 1, 10, 100 or 200ng/ml) or dexamethasone (0, 1, 10, 100 or 200ng/ml). After culture, the expression of mRNAs for BCL-2, BAX, P53, TNF-α, and CASP3 and CASP6 were evaluated. Immunohistochemical results showed that the TNF-α system members, were detected in bovine preantral and antral follicles. After 6days, the TNF-α (10ng/ml) treatment reduced the percentage of normal preantral follicles and increased the number of TUNEL-positive cells in cultured tissue. Dexamethasone (10ng/ml) during 6days of culture did maintain the percentage of normal follicles and the ultrastructure of follicles, while the presence of TNF-α or dexamethasone did not influence primordial follicle activation. However, TNF-α or dexamethasone had no effect on the levels of mRNA for P53, BCL-2, BAX and CASP6, in cultured tissues, but the presence of dexamethasone reduced the levels of CASP3 compared to ovarian slices cultured in control medium (α-MEM+). In conclusion, proteins of the TNF-α system are expressed at different bovine follicle stages. The addition of TNF-α in culture reduces follicle survival and increases the number of apoptotic cells in ovarian tissue, while the presence of dexamethasone maintains follicle ultrastructure in cultured tissue.
This study evaluated (1) the effects of in vivo GnRH treatment on mRNA expression of TNF-α system (TNF-α, TNFR1 and TNFR2) in granulosa cells of bovine preovulatory follicles, (2) the in vitro ...influence of gonadotropins on mRNA expression of TNF-α system in cultured cumulus cells, (3) the protein expression of the TNF-α system in late antral follicles and, (4) the influence of TNF-α on cumulus cells expansion, ultrastructure and on expression of HAS2, CASP3 and CASP6 in follicular cells cultured for 24 h. An increased expression of TNF-α and TNFR1 was observed after 3, 6 and 12 h of GnRH treatment when compared to 0 and 24h. Higher TNFR2 mRNA levels were observed 3, 6 and 12 h after GnRH, when compared to 0 and 24 h. Proteins of TNF-α system were also expressed in late antral follicles. In vitro, TNF-α did not affect cumulus cells expansion, but reduced the HAS2, CASP3 and CASP6 mRNA levels in cumulus cells after 12 h. After 24 h of culture, TNF-α increased the mRNA levels for CASP6 in mural granulosa cells, while the TNF-α, TNFR1 and TNFR2 mRNA levels were increased in cumulus-oocyte complexes (COCs) cultured for 12 h with gonadotropins, but not after 24 h. Ultrastructural analysis confirmed the integrity of COCs cultured in presence of TNF-α. In conclusion, TNF-α system members are present in bovine antral follicles and expression of TNF-α is influenced by gonadotropins in vivo and in vitro. In vitro, TNF-α maintained cumulus cells ultrastructure during COC culture.
•The proteins by TNF-α and its receptors (TNFR1 and TNFR2), are present in antral follicles compartments.•GnRH treatment increases TNF-α mRNA level in granulosa cells of preovulatory follicles in vivo.•In vitro, expression of TNF-α mRNA in cumulus cells is also upregulated by gonadotropins.•TNF-α maintains ultrastructure and regulates gene expression during COC culture.
The aim of this study was to analyze sugarcane (Saccharum officinarum) biometric and technological data, obtained at different timepoints, using path analysis. The experiment was conducted in União, ...PI, Brazil, and evaluated 12 sugarcane genotypes (RB036066, RB9438, RB935744, RB021764, RB021754, RB021534, RB966229, RB977540, RB863129, and RB987935, and the varieties RB92579 and RB867515 as controls) in a randomized block design with four replications. Data were collected at six timepoints that were spaced 30 days apart (90, 120, 150, 180, 210, and 240 days). Direct and indirect effects of the following production components were compared: stalk length, stalk diameter, internode length, number of tillers, number of green leaves, and stalk dry matter. The technological variables evaluated were total recoverable sugar, degrees Brix, tons of polarization (pol, apparent sucrose content) per hectare, juice purity, fiber, juice pol, and tons of sugarcane per hectare. The coefficients of determination were high in all path analyses, suggesting that the components evaluated explained a large part of the variation in stalk production and in the technological variables. Stalk diameter was the trait that best correlated with stalk dry matter yield at all timepoints, with positive values that were higher than the residual effect. This demonstrates the possibility of obtaining significant gains via indirect selection for stalk dry matter yield via stalk diameter or via stalk diameter and number of tillers. The technological variables degrees brix and juice pol were the traits that best correlated with total recoverable sugar production, indicating that they could be used to indirectly select for total recoverable sugar.
Phytochemical investigation of the bark of Guatteria friesiana afforded 12 new aporphines (1–12), along with nine known alkaloids (13–21). The structures of the new alkaloids were determined on the ...basis of spectroscopic data interpretation. The cytotoxic activity of the isolated compounds against a small panel of tumor cell lines was assessed using the Alamar blue assay.
Myelodysplastic syndrome (MDS) is a hematological disorder characterized by abnormal stem cell differentiation and a high risk of acute myeloid leukemia transformation. Treatment options for MDS are ...still limited, making the identification of molecular signatures for MDS progression a vital task. Thus, we evaluated the proteome of bone marrow plasma from patients (n = 28) diagnosed with MDS with ring sideroblasts (MDS-RS) and MDS with blasts in the bone marrow (MDS-EB) using label-free mass spectrometry. This strategy allowed the identification of 1,194 proteins in the bone marrow plasma samples. Polyubiquitin-C (UBC), moesin (MSN), and Talin-1 (TLN1) showed the highest abundances in MDS-EB, and centrosomal protein of 55 kDa (CEP55) showed the highest relative abundance in the bone marrow plasma of MDS-RS patients. In a follow-up, in the second phase of the study, expressions of
UBC
,
MSN
,
TLN1
, and
CEP55
genes were evaluated in bone marrow mononuclear cells from 45 patients by using qPCR. This second cohort included only seven patients from the first study.
CEP55
,
MSN
, and
UBC
expressions were similar in mononuclear cells from MDS-RS and MDS-EB individuals. However,
TLN1
gene expression was greater in mononuclear cells from MDS-RS (p = 0.049) as compared to MDS-EB patients. Irrespective of the MDS subtype,
CEP55
expression was higher (p = 0.045) in MDS patients with abnormal karyotypes, while
MSN
,
UBC
, and
TALIN1
transcripts were similar in MDS with normal vs. abnormal karyotypes. In conclusion, proteomic and gene expression approaches brought evidence of altered TLN1 and CEP55 expressions in cellular and non-cellular bone marrow compartments of patients with low-risk (MDS-RS) and high-risk (MDS-EB) MDSs and with normal vs. abnormal karyotypes. As MDS is characterized by disrupted apoptosis and chromosomal alterations, leading to mitotic slippage, TLN1 and CEP55 represent potential markers for MDS prognosis and/or targeted therapy.
The present study deals with the ecological impacts of the introduction of two alien species of piscivorous fish in several lakes of the Middle Rio Doce lake district in Minas Gerais, Brazil. It was ...demonstrated that these effects were not restricted only to the fish community. The introduction of the predatory red piranha Pygocentrus nattereri and the tucunaré Cichla cf. ocellaris caused not only a sharp decrease in the number of native fish species, but also major shifts in other trophic levels. Just after the fish were introduced, most lakes began to show conspicuous changes in phytoplankton species composition, in which Cyanophyceae gradually came to dominate. The zooplankton community lost several species, and in some cases, such as Lake Carioca, all the cladoceran species disappeared. On the other hand, invertebrate predators, represented by the dipteran Chaoboridae, boomed in the lake, with higher densities of exotic species, probably as a result of the 'ecological release' by reduction of the original fish fauna. There was a general trend of species loss in different trophic levels. All these changes are apparently associated with decreases in water quality. The present situation in these lakes demands new approaches to the management and conservation of these ecosystems.
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