Factors associated with non-completion of TB preventive treatment in Brazil.

BACKGROUND: Among Brazilian initiatives to scale up TB preventive therapy (TPT) are the adoption of the 3HP regimen (12 weekly doses of rifapentine and isoniazid [INH]) in 2021 and the implementation in 2018 of the TPT surveillance information system. Since then, 63% of the 76,000 eligible individuals notified completed TPT. Recommended regimens in this period were 6H, 9H (6 or 9 months of INH) and 4R (4 months of rifampicin).OBJECTIVE: To analyse the factors associated with TPT non-completion.METHODS: We analysed the cohort of TPT notifications from 2018 to 2020. Robust variance Poisson regression model was used to verify the association of TPT non-completion with sociodemographic, clinical and epidemiological variables.RESULTS: Of the 39,973 TPT notified in the study period, 8,534 (21.5%) were non-completed, of which 7,858 (92.1%) were lost to follow-up. Age 15-60 years (relative risk [RR] 1.27, 95% confidence interval [95% CI] 1.20-1.35), TPT with isoniazid (RR 1.40, 95% CI 1.19-1.64) and Black/mixed race (RR 1.17, 95% CI 1.09-1.25) were associated with a higher risk of non-completion.CONCLUSION: Individuals in situations of social and financial vulnerability such as being Black/pardo race, younger and on longer TPT regimens were more likely to be associated with TPT incompletion.

Transcriptional downregulation of ABC transporters is related to follicular degeneration after vitrification and in vitro culture of ovine ovarian tissue.

ATP-binding cassette (ABC) transporters perform multiple functions in reproductive tissues. During ovarian tissue vitrification, the plasma membrane has important functions in the influx or efflux of water, and substances such as cryoprotectants and channel proteins that are required in this process. Thus, the present study aimed to verify the relative abundance of mRNA transcript of ABC transporters ABCB1, ABCG2, and MRP2 after vitrification and in vitro culture (IVC) of ovine ovarian tissue. For this study, the ovarian cortex fragments were proportioned into four groups as fresh control, vitrified control, fresh culture, and vitrified culture groups. After vitrification and in vitro culture, the ovarian tissue was evaluated using morphological procedures. Further, relative abundance of ABCB1, ABCG2, and MRP2 transporter mRNA transcripts in the ovarian cortex subjected to aforementioned treatment conditions were evaluated using qPCR. Our results showed a negative association between degenerated follicles and mRNA transcript abundances of ABCB1 and ABCG2. In addition, the percentage of growing follicles in the ovine ovarian cortex after vitrification was similar to that of the fresh control tissue without in vitro culture. The in vitro culture of fresh and vitrified tissue however, showed a significant decrease in the percentage of growing follicles. To the best of our knowledge, we believe that our data for the first time has studied the relative abundances of ABCB1 and ABCG2 mRNA transcripts in the ovine ovarian cortex. In addition, alterations of these protein channels may be indicative of a deleterious effect of osmotic stress on follicular survival during vitrification. Furthermore, these effects were detectable only after the IVC of the ovarian tissues. Nonetheless, further studies are required to investigate the functions of ABC transporters in ovine folliculogenesis, especially after in vitro culture of ovarian tissue.

Heterotopic autotransplantation of equine ovarian tissue using intramuscular versus subvulvar grafting sites: Preliminary results.

Ovarian tissue transplantation (OTT) is a technique well established and successfully applied in humans using mainly orthotopic or heterotopic transplantation sites. In livestock, OTT is still in its infancy and, therefore, different aspects of the technique, including the efficiency of different heterotopic OTT sites as well as the potential effect of age (i.e., young vs. old mares) in the ovarian graft quality, need to be investigated. The present study investigated the efficacy of the intramuscular (IM) or the novel subvulvar mucosa (SV) heterotopic autotransplantation sites to maintain the survivability of the grafts for 3 and 7 days post-OTT. Ovarian biopsy fragments were obtained in vivo and distributed to the following treatments: Fresh control group (ovarian fragments immediately fixed), SV-3, IM-3, SV-7, and IM-7. During and after graft harvesting, the macroscopic characteristics of the grafts (i.e., adherence, morphology, and bleeding) were scored, and the percentages of morphologically normal and developing preantral follicles as well as the follicular and stromal cell densities of the grafts were evaluated. The results were that similar (P > 0.05) macroscopic scores were observed between both transplantation sites 7 days post-OTT, with positive correlations (P < 0.01) found among adherence, morphology, and bleeding of the grafts. A lower (P < 0.05) percentage of morphologically normal follicles was found 7 days post-OTT in the SV site (82%) compared with the Fresh control group (99%) and IM site (95%); however, the percentages of developing follicles were similar (P > 0.05) between both transplantation sites 7 days post-OTT (30-43%). Although similar (P > 0.05) follicular densities were found in both transplantation sites in young and old mares at 3 and 7 days post-OTT, large individual variation in the follicular depletion rate was observed regardless of transplantation site. The Fresh control group and SV-7 treatments had higher (P < 0.05) stromal cell densities in young and old mares compared with both IM-7 treatments. When comparing transplant sites between young and old mares, the follicular density in old mares and the stromal cell density in young mares were greater (P < 0.05) in the SV than in the IM site. In conclusion, even though the transplantation sites differentially affected some end points, overall comparable findings of the OTT technique using both heterotopic autotransplantation sites (i.e., IM and SV) for equine ovarian tissue were observed.

Predicting the chemical composition of the body and the carcass of hair sheep using body parts and carcass measurements.

Determination of the chemical composition in the body and carcass of ruminants is important for both nutritional requirement studies and the meat industry. This study aimed to develop equations to predict the body and carcass chemical composition of hair sheep using the chemical composition of body parts, carcass measurements and shrunk BW as predictors. A database containing 107 individual records for castrated male hair sheep ranging from 24 to 43 kg BW was gathered from two body composition studies. The empty body, carcass and body parts were analyzed for water, ash, fat and protein contents (%). The body parts used to estimate body and carcass composition were fore leg, hind leg and 9-11th rib section. The carcass measurements used were leg length, thoracic circumference, hind circumference, hind width, thoracic width, thoracic depth and chest width. Each model performance was evaluated using a leave-one-out cross-validation. Multiple regression analysis considering the study as a random effect revealed that body parts in association with carcass measurements were significant for predicting the chemical composition in the body of castrate male sheep. However, the use of the chemical composition of hind leg produced the best models for predicting the ash and fat contents in the empty body, whereas the water and protein contents in the empty body were better predicted when using the chemical compositions of 9-11th rib section and fore leg, respectively. Multiple regression analysis also revealed that most body parts were suitable for predicting the carcass composition, except for 9-11th rib section whose chemical composition did not produce significant prediction equations for ash and protein carcass contents. The use of the chemical composition of hind leg in association with carcass measurements produced the best models for predicting the water and fat contents in the carcass, while the ash and protein contents in the carcass were better predicted when using the chemical composition of fore leg. In conclusion, precision, accuracy and goodness-of-fit of the equations drove the selection of the chemical composition of hind leg and carcass measurements in a multivariate approach, as the most suitable predictors of the chemical composition of the body and carcass of hair sheep. However, the chemical composition of fore leg may be used as well. The developed equations could improve the accuracy of the empty body and carcass composition estimations in sheep, optimizing the estimation of nutrient requirements, as well as the carcass quality evaluation for this species.

Harvesting, processing, and evaluation of in vitro-manipulated equine preantral follicles: A review.

The mammalian ovary is responsible for essential stages of folliculogenesis and hormonal production, regulating the female physiological functions during the menstrual/estrous cycles. The mare has been considered an attractive model for comparative studies due to the striking similarities shared with women regarding in vivo and in vitro folliculogenesis. The ovarian follicular population in horses contains a large number of oocytes enclosed in preantral follicles that are yet to be explored. Therefore, the in vitro manipulation of equine preantral follicles aims to avoid the process of atresia and promote the development of follicles with competent oocytes. In this regard, after ovarian tissue harvesting, the use of appropriate processing techniques, as well as suitable approaches to evaluating equine preantral follicles and ovarian tissue, are necessary. Although high-quality equine ovarian tissue can be obtained from several sources, some critical aspects, such as the age of the animals, ovarian cyclicity, reproductive phase, and the types of ovarian structures, should be considered. Therefore, this review will focus on providing an update on the most current advances concerning the critical factors able to influence equine preantral follicle quality and quantity. Also, the in vivo strategies used to harvest equine ovarian tissue, the approaches to manipulating ovarian tissue post-harvesting, the techniques for processing ovarian tissue, and the classical approaches used to evaluate preantral follicles will be discussed.

Supportive techniques to investigate in vitro culture and cryopreservation efficiencies of equine ovarian tissue: A review.

During the reproductive lifespan of a female, only a limited quantity of oocytes are naturally ovulated; therefore, the mammalian ovary possesses a substantial population of preantral follicles available to be handled and explored in vitro. Hence, the manipulation of preantral follicles enclosed in ovarian tissue aims to recover a considerable population of oocytes of high-value animals for potential application in profitable assisted reproductive technologies (ARTs). For this purpose, the technique of preantral follicle in vitro culture (IVC) has been the most common research tool, achieving extraordinary results with offspring production in the mouse model. Although promising outcomes have been generated in livestock animals after IVC of preantral follicles, the quantity and quality of embryo production with those oocytes are still poor. In recent years, the mare has become an additional model for IVC studies due to remarkable similarities with women and livestock animals regarding in vivo and in vitro ovarian folliculogenesis. For a successful IVC system, several factors should be carefully considered to provide an optimum culture environment able to support the viability and growth of preantral follicles enclosed in ovarian tissue. The cryopreservation of the ovarian tissue is another important in vitro manipulation technique that has been used to preserve the reproductive potential in humans and, in the future, may be used in highly valuable domestic animals or endangered species. Several improvements in cryopreservation protocols are necessary to support the utilization of ovarian tissue of different species in follow-up ARTs (e.g., ovarian fragment transplantation). This review aims to provide an update on the most current advances regarding supportive in vitro techniques used in equids to evaluate and manipulate preantral follicles and ovarian tissue, as well as methodological approaches used during IVC and cryopreservation techniques.

Betaine-loaded CaCO3 microparticles improve survival of vitrified feline preantral follicles through higher mitochondrial activity and decreased reactive oxygen species.

Ovary fragments from six sexually mature cats were vitrified in the presence or absence of betaine or ascorbic acid, loaded (7.4 or 74µM betaine; 20 or 200µM ascorbic acid) or not (1mM betaine or 0.3mM ascorbic acid) into CaCO3 microparticles, and assessed for follicular morphology, oxidative stress and mitochondrial activity Feline ovarian tissue was successfully preserved after vitrification in the presence of 74µM betaine loaded in CaCO3 microparticles, as confirmed by morphological analysis and the density of preantral follicles and stromal cells, as well as by the increased mitochondrial activity and decreased production of reactive oxygen species.

Cryopreservation of Bovine Semen Using Extract of Bertholletia excelsa (Brazil Nuts).

BACKGROUND: Semen cryopreservation is essential in animal breeding programs for improving the availability of genetic resources from animals with high breeding value. OBJECTIVE: To evaluate the addition of Brazil nut extract as a replacement for egg yolk in bovine semen cryopreservation. MATERIALS AND METHODS: Semen was collected from five Nelore bulls and cryopreserved with the addition (treatments) of 0, 25, 50, 75, or 100% Brazil nut extract in the cryoprotectant medium. After thawing, spermatic cells were evaluated for morphology, plasma membrane integrity, spermatic kinetics, and in vitro fertilization. The experimental design was in randomized blocks, and the data were submitted to regression analysis. RESULTS: The minor-type and total defects, and plasma membrane integrity were affected (P < 0.05) as a function of egg yolk substitution with Brazil nut extract. There was a significant effect (P < 0.05) of Brazil nut extract addition on the spermatic kinetics and cleavage rate. CONCLUSION: The addition of Brazil nut extract in the cryoprotective medium as a substitute of egg yolk for freezing bovine semen negatively affects sperm quality and fertility.

Expression of myo-inositol cotransporters in the sciatic nerve and dorsal root ganglia in experimental diabetes.

The transport of myo-inositol is the main mechanism for the maintenance of its high intracellular levels. We aimed to measure the mRNA and protein levels of myo-inositol cotransporters in the sciatic nerve (SN) and dorsal root ganglia (DRG) during experimental diabetes. Streptozotocin-induced (STZ; 4, 8, and 12 weeks; 65 mg/kg; ip) diabetic rats (DB) and age-matched euglycemic (E) rats were used for the analysis of mRNA and protein levels of sodium myo-inositol cotransporters 1, 2 (SMIT1, SMIT2) or H+/myo-inositol cotransporter (HMIT). There was a significant reduction in the mRNA levels for SMIT1 in the SN and DRG (by 36.9 and 31.0%) in the 4-week DB (DB4) group compared to the E group. SMIT2 was not expressed in SN. The mRNA level for SMIT2 was up-regulated only in the DRG in the DB4 group. On the other hand, the protein level of SMIT1 decreased by 42.5, 41.3, and 44.8% in the SN after 4, 8, and 12 weeks of diabetes, respectively. In addition, there was a decrease of 64.3 and 58.0% of HMIT in membrane and cytosolic fractions, respectively, in the SN of the DB4 group. In the DRG, there was an increase of 230 and 86.3% for SMIT1 and HMIT, respectively, in the DB12 group. The levels of the main inositol transporters, SMIT1 and HMIT, were greatly reduced in the SN but not in the DRG. SMIT-1 was selectively reduced in the sciatic nerve during experimental STZ-induced diabetes.

Performance and digestibility of confined lambs fed with Babassu cake (Orbignya speciosa) as a substitute for elephant grass silage / Desempenho e digestibilidade de cordeiros confinados alimentados com torta de babaçu (Orbignya speciosa) em substituição à silagem de capim-elefante

The objective of this study was to evaluate the effect of the partial replacement of elephant grass silage with babassu (Orbignya speciosa) on the performance, intake, digestibility, and carcass weight gain of sheep. Fifty-four castrated male lambs (19.08±2.76kg) were distributed in a completely randomized design and administered one of the following treatments: 0.0, 12.5, 25.0, 37.5 or 50% dry matter (%DM) replacement of elephant grass silage with babassu cake. No difference (P> 0.05) was observed in the dry matter, crude protein, and neutral detergent fiber intake, but the ether extract intake increased (P< 0.05). The digestibility of the dry matter, neutral detergent fiber, and total digestible nutrients were unaffected. The crude protein digestibility decreased (P< 0.05), and the ether extract digestibility increased with the replacement of the elephant grass. There were no changes in mean daily weight gain and carcass weight gain. In the carcasses, a linear increase was observed in the proportion of the ether extract, and the crude protein decreased. The replacement of the silage with the babassu cake by up to 50% did not change the performance of sheep, however it led to an increase in fat deposit in the carcass.

Objetivou-se avaliar a substituição parcial da silagem de capim elefante por torta de babaçu (Orbignya speciosa) sobre o desempenho, consumo, digestibilidade, composição química e ganho em peso de carcaça de ovinos. Cinquenta e quatro machos, castrados (19,08±2,76kg) foram distribuídos em delineamento inteiramente casualizado e administrado um dos seguintes tratamentos: 0.0, 12.5, 25.0, 37.5 e 50% (%MS) de substituição da silagem pela torta de babaçu. Não houve diferença (P> 0.05) na ingestão de matéria seca, proteína bruta e fibra em detergente neutro, porém o consumo de extrato etéreo aumentou (P< 0.05). A digestibilidade da matéria seca, fibra em detergente neutro e nutrientes digestíveis totais não foram alterados, no entanto, os coeficientes de digestibilidade da proteína bruta reduziu (P< 0.05) e do extrato etéreo aumentou (P< 0.05), com a substituição. Não houve alterações no ganho de peso médio diário e ganho de peso em carcaça. Nas carcaças, observou-se aumento linear na proporção do extrato etéreo, no entanto a porcentagem de proteína bruta diminuiu. O aumento no extrato etéreo da carcaça mostrou uma tendência linear. A substituição da silagem pela torta de babaçu em até 50% não alterou o desempenho de ovinos, porém levou a aumento na deposição de gordura na carcaça.

Agronomic performance of Marandu grass treated with plant growth biostimulants in the Amazon biome / Desempenho agronômico de capim-marandu tratado com bioestimulantes no bioma Amazônia

The aim was to evaluate the effect of different doses of two biostimulants on the productivity and canopy structure of Urochloa brizantha cv. Marandu grass in the establishment fase. The study was conducted in Universidade Federal Rural da Amazônia, Parauapebas, Pará, Brazil. One module of 35 plots of 25m² were established. A completely randomized experimental design was used, with seven treatments and five replicates. The treatments included a control, 0.5, 1, and 2kg/ha of biostimulant A (BIOST.A); and 0.25, 0.5, and 1L/ha of biostimulant B (BIOST.B). Three collections were performed. The data for application of the two biostimulants were analyzed separately, using the Dummy variable method and regression analysis. The application of 2kg/ha BIOST.A resulted in increases of 842kg/ha in the forage mass. The application of BIOST.B on Marandu grass resulted in a linear increase in stem mass. The application of 2kg/ha BIOST.A in the establishment of Marandu grass result in higher growth rates, forage accumulation, and stem proportion in the canopy.(AU)

Objetivou-se avaliar o efeito de diferentes doses de dois bioestimulantes sobre a produtividade e a estrutura do dossel do capim Urochloa brizantha cv. Marandu na fase de estabelecimento. O estudo foi realizado na Universidade Federal Rural da Amazônia, Parauapebas, Pará, Brasil. Um módulo de 35 parcelas de 25m² foi estabelecido. Utilizou-se delineamento experimental inteiramente ao acaso, composto de sete tratamentos e cinco repetições cada. Os tratamentos incluídos no controle foram: 0,5, 1 e 2kg/ha de bioestimulante A (BIOST.A); 0,25; 0,5 e 1L/ha de bioestimulante B (BIOST.B). Foram realizadas três coletas. Os dados para a aplicação dos dois bioestimulantes foram analisados separadamente, utilizando-se organização por método variável Dummy e análise de regressão. A aplicação de 2kg/ha BIOST.A resultou em incrementos de 842kg/ha na massa de forragem. A aplicação do BIOST.B na grama marandu resultou em aumento linear na produção de massa do caule. A aplicação de 2kg/ha BIOST.A no estabelecimento de erva de marandu resultou em maiores taxas de crescimento, acumulação de forragem e proporção de caule no dossel.(AU)

The Addition of AçaÍ Palm Fruit Extract in the Cryopreservation Diluent of Bull's Semen with Low Performance on Freezing.

BACKGROUND: Semen freezing is of great importance for animal production because it allows the use and the rapid diffusion of the genetic material from economically important animals. OBJECTIVE: To evaluate the effect of açai (Euterpe oleracea; Arecaceae family) extract addition to the semen cryopreservation diluent on the morphology, sperm motility parameters, and plasma membrane integrity of spermatozoa. MATERIALS AND METHODS: The ejaculates, obtained from five bulls with low performance on semen freezing, were fractionated and distributed according to the experimental group. The control samples did not have açaí extract added, whereas to the treated groups were added 5, 10, 15 or 20 mg ml-1 of açaí extract into the semen diluent. The sperm morphology was evaluated with a formalin-saline-buffered solution. The plasma membrane integrity was evaluated by the epifluorescent test, while the cellular kinetics was assessed by an automated analysis of the spermatic movement. RESULTS: The sperm defects showed a linearly decreasing effect (P < 0.05) with the addition of different concentrations of açaí extract. The plasma membrane integrity was higher (P < 0.05) after the açaí addition to the cryopreservation diluent. There was no significant effect (P > 0.05) of the açaí extract on the kinetics of spermatozoa. CONCLUSION: The addition of açaí extract to the cryopreservation diluent provided better preservation of the structural integrity of the sperm plasma membrane in the bull's semen with low tolerance to the cryopreservation process.

Expression of myo-inositol cotransporters in the sciatic nerve and dorsal root ganglia in experimental diabetes

The transport of myo-inositol is the main mechanism for the maintenance of its high intracellular levels. We aimed to measure the mRNA and protein levels of myo-inositol cotransporters in the sciatic nerve (SN) and dorsal root ganglia (DRG) during experimental diabetes. Streptozotocin-induced (STZ; 4, 8, and 12 weeks; 65 mg/kg; ip) diabetic rats (DB) and age-matched euglycemic (E) rats were used for the analysis of mRNA and protein levels of sodium myo-inositol cotransporters 1, 2 (SMIT1, SMIT2) or H+/myo-inositol cotransporter (HMIT). There was a significant reduction in the mRNA levels for SMIT1 in the SN and DRG (by 36.9 and 31.0%) in the 4-week DB (DB4) group compared to the E group. SMIT2 was not expressed in SN. The mRNA level for SMIT2 was up-regulated only in the DRG in the DB4 group. On the other hand, the protein level of SMIT1 decreased by 42.5, 41.3, and 44.8% in the SN after 4, 8, and 12 weeks of diabetes, respectively. In addition, there was a decrease of 64.3 and 58.0% of HMIT in membrane and cytosolic fractions, respectively, in the SN of the DB4 group. In the DRG, there was an increase of 230 and 86.3% for SMIT1 and HMIT, respectively, in the DB12 group. The levels of the main inositol transporters, SMIT1 and HMIT, were greatly reduced in the SN but not in the DRG. SMIT-1 was selectively reduced in the sciatic nerve during experimental STZ-induced diabetes.

Essential oil of Pterodon polygalaeflorus Benth attenuates nociception in mice.

Essential oils (EO) are volatile liquids responsible for the aroma of plants. Pterodon polygalaeflorus seeds have received widespread use in folk medicine for the treatment of inflammatory diseases. For this reason and because Pterodon polygalaeflorus seeds have great EO content, which is frequently pharmacologically active, the present study aimed to evaluate the antinociceptive effect of EO from Pterodon polygalaeflorus (EOPPgfl) and its acute toxic effects. The EEOPPgfl sample, which was extracted by steam distillation of the seeds, had a yield of 2.4% of the seeds weight and had, as major constituents, beta-elemene (48.19%), trans-caryophyllene (19.51%), and epi-bicyclosesquiphellandrene (12.24%). The EOPPgfl sample showed mild acute toxicity and its calculated median lethal dose (LD50) was 3.38 g/kg. EOPPgfl (20-60 mg/kg) showed antinociceptive activity as evidenced by several tests and inhibited writhing induced by acetic acid. The maximum effect was obtained with the 30 mg/kg dose and at 60 min after its administration. EOPPgfl also decreased formalin-induced nociception, as verified by the inhibition of the first and second phase of the formalin test. At 30 mg/kg, EOPPgfl also decreased thermally stimulated nociception. Nociception may be related to inflammatory and antiedematogenic activity and at doses ranging 10-100 mg/kg, EOPPgfl blocked dextran- and carrageenan-induced edema. The results demonstrated that EOPPgfl presented, at doses approximately 100 times smaller than LD50, an antinociceptive effect that probably was due to anti-inflammatory activities.

In vivo and in vitro strategies to support caprine preantral follicle development after ovarian tissue vitrification.

The aim of the present study was to compare fresh and vitrified goat ovarian tissue after autotransplantation and in vitro culture. Adult goats were completely ovariectomised and each ovarian pair was sliced and distributed among six different treatment groups: fresh control, fresh transplant, fresh culture, vitrified control, vitrified transplant and vitrified culture. Follicular morphology, development, growth, density, revascularisation and hormone production were evaluated in all groups. Three antral follicles (two in the fresh transplant and one in the vitrified transplant groups) were observed on the surface of the graft 90 days after transplantation. The percentage of morphologically normal follicles was similar in the fresh control, fresh transplant and vitrified transplant groups. The percentage of developing (transition, primary and secondary) follicles was higher after in vitro culture of fresh or vitrified tissue. Transplantation resulted in a lower follicle density. Serum oestradiol concentrations remained constant during the entire transplantation period. In contrast, progesterone production decreased significantly. Expression of CD31 mRNA was lower in fresh culture. In conclusion, restoration of goat ovarian function can be successfully achieved following transplantation of both fresh and vitrified goat ovarian tissue. However, transplantation induced higher follicle loss than in vitro culture.

Essential oil of Pterodon polygalaeflorus Benth attenuates nociception in mice

Essential oils (EO) are volatile liquids responsible for the aroma of plants. Pterodon polygalaeflorus seeds have received widespread use in folk medicine for the treatment of inflammatory diseases. For this reason and because Pterodon polygalaeflorus seeds have great EO content, which is frequently pharmacologically active, the present study aimed to evaluate the antinociceptive effect of EO from Pterodon polygalaeflorus (EOPPgfl) and its acute toxic effects. The EEOPPgfl sample, which was extracted by steam distillation of the seeds, had a yield of 2.4% of the seeds weight and had, as major constituents, beta-elemene (48.19%), trans-caryophyllene (19.51%), and epi-bicyclosesquiphellandrene (12.24%). The EOPPgfl sample showed mild acute toxicity and its calculated median lethal dose (LD50) was 3.38 g/kg. EOPPgfl (20-60 mg/kg) showed antinociceptive activity as evidenced by several tests and inhibited writhing induced by acetic acid. The maximum effect was obtained with the 30 mg/kg dose and at 60 min after its administration. EOPPgfl also decreased formalin-induced nociception, as verified by the inhibition of the first and second phase of the formalin test. At 30 mg/kg, EOPPgfl also decreased thermally stimulated nociception. Nociception may be related to inflammatory and antiedematogenic activity and at doses ranging 10-100 mg/kg, EOPPgfl blocked dextran- and carrageenan-induced edema. The results demonstrated that EOPPgfl presented, at doses approximately 100 times smaller than LD50, an antinociceptive effect that probably was due to anti-inflammatory activities.

Effects of Lippia sidoides essential oil, thymol, p-cymene, myrcene and caryophyllene on rat sciatic nerve excitability.

Lippia sidoides Cham is a typical herb species of Northeast Brazil with widespread use in folk medicine. The major constituents of the essential oil of L. sidoides (EOLs) are thymol, p-cymene, myrcene, and caryophyllene. Several studies have shown that the EOLs and its constituents have pharmacological effects, including antibacterial, anti-inflammatory, antioxidant and neuroprotective activity. Therefore, this work aimed to investigate the effects of the EOLs and their main constituents on rat sciatic nerve excitability. The sciatic nerves of adult Wistar rats were dissected and mounted in a moist chamber. Nerves were stimulated by square wave pulses, with an amplitude of 40 V, duration of 100 µs to 0.2 Hz. Both EOLs and thymol inhibited compound action potential (CAP) in a concentration-dependent manner. Half maximal inhibitory concentration for CAP peak-to-peak amplitude blockade were 67.85 and 40 µg/mL for EOLs and thymol, respectively. CAP peak-to-peak amplitude was significantly reduced by concentrations ≥60 µg/mL for EOLs and ≥30 µg/mL for thymol. EOLs and thymol in the concentration of 60 µg/mL significantly increased chronaxie and rheobase. The conduction velocities of 1st and 2nd CAP components were also concentration-dependently reduced by EOLs and thymol in the range of 30-100 µg/mL. Differently from EOLs and thymol, p-cymene, myrcene and caryophyllene did not reduce CAP in the higher concentrations of 10 mM. These data demonstrated that EOLs and thymol inhibited neuronal excitability and were promising agents for the development of new drugs for therapeutic use.

Refining insulin concentrations in culture medium containing growth factors BMP15 and GDF9: An in vitro study of the effects on follicle development of goats.

The aim of the present study was to investigate the influence of two insulin concentrations (10ng/mL and 10µg/mL) combined or in the absence of BMP15 and/or GDF9, on the in vitro survival and development of preantral follicles of goat ovarian tissue. Ovarian slices from the same goat ovary pair were randomly assigned to a non-cultured control treatment or to be in vitro cultured for 1 or 7days in α-MEM containing 10ng/mL (Low) or 10µg/mL (High) of insulin in the absence or presence of BMP15 and/or GDF9. With the low insulin treatment, there was a greater percentage of normal follicles than with the high insulin treatment. The addition of BMP15 alone or in association with GDF9 to the medium containing low insulin resulted in a lesser percentage of normal follicles (P<0.05). The addition of BMP15 and GDF9 separately or in combination with the high insulin concentration enhanced the percentage of normal follicles. On day 7 of culture, the use of medium containing low insulin alone or high insulin supplemented with BMP15 and BMP15+GDF9 resulted in a greater percentage of secondary follicles than the non-cultured control, although follicles cultured with low insulin were smaller than those from the control group and had greater rates of oxidative stress. In conclusion, in the presence of physiological concentrations of insulin (10ng/mL), medium supplementation with GDF9 and BMP15 alone or in combination is unnecessary for normal follicle development in vitro.

Expectations and limitations of ovarian tissue transplantation.

Constant progress in the diagnosis and treatment of cancer disease has increased the number and prognosis of cancer survivors. However, the toxic effects of chemotherapy and radiotherapy on ovarian function have resulted in premature ovarian failure. Patients are, therefore, still expecting methods to be developed to preserve their fertility successfully. Several potential options are available to preserve fertility in patients who face premature ovarian failure, including immature or mature oocyte and embryo cryopreservation. However, for children or prepubertal women needing immediate chemotherapy, cryopreservation of ovarian tissue is the only alternative. The ultimate aim of this strategy is to implant ovarian tissue into the pelvic cavity (orthotopic site) or in a heterotopic site once oncological treatment is completed and the patient is disease free. Transplantation of ovarian tissue with sufficiently large numbers of follicles could potentially restore endocrine function and allow multiple cycles for conception. However, the success of ovarian tissue transplantation still has multiple challenges, such as the low number of follicles in the graft that may affect their longevity as well as the survival of the tissue during ex vivo processing and subsequent transplantation. Therefore, this review aims to summarize the achievements of ovary grafting and the potential techniques that have been developed to improve ovarian graft survival.

Equine ovarian tissue viability after cryopreservation and in vitro culture.

Ovarian tissue cryopreservation allows the preservation of the female fertility potential for an undetermined period. The objectives of this study were to compare the efficiency of cryoprotective agents (CPAs; dimethyl sulfoxide, DMSO; ethylene glycol, EG; and propylene glycol, PROH) using slow-freezing and vitrification methods, and evaluate the viability of cryopreserved equine ovarian tissue after 7 days of culture. Fresh and cryopreserved ovarian fragments were evaluated for preantral follicle morphology, stromal cell density, EGFR, Ki-67, Bax, and Bcl-2 protein expression, and DNA fragmentation. Vitrification with EG had the highest rate of morphologically normal preantral follicles, while DMSO had the lowest (76.1 ± 6.1% and 40.9 ± 14.8%, respectively; P < 0.05). In slow-freezing, despite that DMSO had the highest percentage of morphologically normal follicles (77.7 ± 5.8%), no difference among the CPAs was observed. Fluorescence intensity of EGFR and Ki-67 was greater when vitrification with EG was used. Regardless of the cryopreservation treatment, DMSO had the highest (P < 0.05) Bax/Bcl-2 ratio; however, DNA fragmentation was similar (P > 0.05) among treatments after thawing. After in vitro culture, the percentage of normal follicles was similar (P > 0.05) between slow-freezing and vitrification methods; however, vitrification had greater (P < 0.05) stromal cell density than slow-freezing. In summary, equine ovarian tissue was successfully cryopreserved, increasing the viability of the cells in the ovarian tissue after thawing when using DMSO and EG for slow-freezing and vitrification methods, respectively. Therefore, these results are relevant for fertility preservation programs.