Genome-wide association study and annotating candidate gene networks affecting age at first calving in Nellore cattle.

We performed a genome-wide mapping for the age at first calving (AFC) with the goal of annotating candidate genes that regulate fertility in Nellore cattle. Phenotypic data from 762 cows and 777k SNP genotypes from 2,992 bulls and cows were used. Single nucleotide polymorphism (SNP) effects based on the single-step GBLUP methodology were blocked into adjacent windows of 1 Megabase (Mb) to explain the genetic variance. SNP windows explaining more than 0.40% of the AFC genetic variance were identified on chromosomes 2, 8, 9, 14, 16 and 17. From these windows, we identified 123 coding protein genes that were used to build gene networks. From the association study and derived gene networks, putative candidate genes (e.g., PAPPA, PREP, FER1L6, TPR, NMNAT1, ACAD10, PCMTD1, CRH, OPKR1, NPBWR1 and NCOA2) and transcription factors (TF) (STAT1, STAT3, RELA, E2F1 and EGR1) were strongly associated with female fertility (e.g., negative regulation of luteinizing hormone secretion, folliculogenesis and establishment of uterine receptivity). Evidence suggests that AFC inheritance is complex and controlled by multiple loci across the genome. As several windows explaining higher proportion of the genetic variance were identified on chromosome 14, further studies investigating the interaction across haplotypes to better understand the molecular architecture behind AFC in Nellore cattle should be undertaken.

TRIENNIAL GROWTH AND DEVELOPMENT SYMPOSIUM: Dedifferentiated fat cells: Potential and perspectives for their use in clinical and animal science purpose.

An increasing body of evidences has demonstrated the ability of the mature adipocyte to dedifferentiate into a population of proliferative-competent cells known as dedifferentiated fat (DFAT) cells. As early as the 1970s, in vitro studies showed that DFAT cells may be obtained by ceiling culture, which takes advantage of the buoyancy property of lipid-filled cells. It was documented that DFAT cells may acquire a phenotype similar to mesenchymal stem cells and yet may differentiate into multiple cell lineages, such as skeletal and smooth muscle cells, cardiomyocytes, osteoblasts, and adipocytes. Additionally, recent studies showed the ability of isolated mature adipocytes to dedifferentiate in vivo and the capacity of the progeny cells to redifferentiate into mature adipocytes, contributing to the increase of body fatness. These findings shed light on the potential for use of DFAT cells, not only for clinical purposes but also within the animal science field, because increasing intramuscular fat without excessive increase in other fat depots is a challenge in livestock production. Knowledge of the mechanisms underlying the dedifferentiation and redifferentiation of DFAT cells will allow the development of strategies for their use for clinical and animal science purposes. In this review, we highlight several aspects of DFAT cells, their potential for clinical purposes, and their contribution to adipose tissue mass in livestock.

Review: Animal model and the current understanding of molecule dynamics of adipogenesis.

Among several potential animal models that can be used for adipogenic studies, Wagyu cattle is the one that presents unique molecular mechanisms underlying the deposit of substantial amounts of intramuscular fat. As such, this review is focused on current knowledge of such mechanisms related to adipose tissue deposition using Wagyu cattle as model. So abundant is the lipid accumulation in the skeletal muscles of these animals that in many cases, the muscle cross-sectional area appears more white (adipose tissue) than red (muscle fibers). This enhanced marbling accumulation is morphologically similar to that seen in numerous skeletal muscle dysfunctions, disease states and myopathies; this might indicate cross-similar mechanisms between such dysfunctions and fat deposition in Wagyu breed. Animal models can be used not only for a better understanding of fat deposition in livestock, but also as models to an increased comprehension on molecular mechanisms behind human conditions. This revision underlies some of the complex molecular processes of fat deposition in animals.

An inexpensive and worldwide available digital image analysis technique for histological fibrosis quantification in chronic hepatitis C.

Hepatic fibrosis staging is based on semiquantitative scores. Digital imaging analysis (DIA) appears more accurate because fibrosis is quantified in a continuous scale. However, high cost, lack of standardization and worldwide unavailability restrict its use in clinical practice. We developed an inexpensive and widely available DIA technique for fibrosis quantification in hepatitis C, and here, we evaluate its reproducibility and correlation with semiquantitative scores, and determine the fibrosis percentage associated with septal fibrosis and cirrhosis. 282 needle biopsies staged by Ishak and METAVIR scores were included. Images of trichrome-stained sections were captured and processed using Adobe(®) Photoshop(®) CS3 and Adobe(®) Bridge(®) softwares. The percentage of fibrosis (fibrosis index) was determined by the ratio between the fibrosis area and the total sample area, expressed in pixels calculated in an automated way. An excellent correlation between DIA fibrosis index and Ishak and METAVIR scores was observed (Spearman's r = 0.95 and 0.92; P < 0.001, respectively). Excellent intra-observer reproducibility was observed in a randomly chosen subset of 39 biopsies with an intraclass correlation index of 0.99 (95% CI, 0.95-0.99). The best cut-offs associated with septal fibrosis and cirrhosis were 6% (AUROC 0.97, 95% CI, 0.95-0.99) and 27% (AUROC 1.0, 95% CI, 0.99-1), respectively. This new DIA technique had high correlation with semiquantitative scores in hepatitis C. This method is reproducible, inexpensive and available worldwide allowing its use in clinical practice. The incorporation of DIA technique provides a more complete evaluation of fibrosis adding the quantification to architectural patterns.

Eficácia do glyphosate e 2, 4-d no controle de Commelina villosa / Efficacy of glyphosate and 2, 4-D in control of Commelina villosa

O objetivo do presente trabalho foi avaliar a eficácia de glyphosate e 2,4-D, isolados e em mistura, no controle de Commelina villosa. Foram estudadas duas metodologias de avaliação de absorção de herbicidas em oito intervalos de tempo para a lavagem (simulando chuva após a aplicação) e corte (simulando abortamento, como estratégia de defesa) das folhas: 2, 4, 6, 8, 12, 24 e 48 horas após a aplicação dos herbicidas, além de um tratamento sem lavagem ou corte das folhas, em delineamento experimental inteiramente casualizado com quatro repetições, dispostos em um esquema fatorial 3 x 7 + 1 (três herbicidas x sete períodos – horas após a aplicação). Os herbicidas e doses testados foram: glyphosate (1.440 g ha-1), 2,4-D (720 g ha-1) e a mistura glyphosate + 2,4-D (1.080 + 720 g ha-1). A simulação de chuva interferiu de forma negativa no controle das plantas com o herbicida glyphosate. O controle com o herbicida 2,4-D foi influenciado apenas no período de 2 horas. Os períodos de simulação de chuva não influenciaram no controle das plantas com a mistura de glyphosate + 2,4-D. Para o estudo com corte das folhas tratadas, todos os tratamentos independente do período para corte das folhas foram influenciados de forma negativa no controle, sendo que as plantas apresentaram rebrotas quando tratadas com o herbicida 2,4-D isolado.

Efficacy of glyphosate and 2,4-d in the control of Commelina Villosa. The aim of this study was to evaluate the efficacy of glyphosate and 2,4-D alone and in combination, in the control of Commelina villosa. We studied two methodologies for evaluating herbicide absorption in eight time intervals for washing (simulating rainfall after application) and cutting of leaves (simulating abortion as a defense strategy): 2, 4, 6, 8, 12, 24 and 48 hours after herbicide application, and a treatment without washing or cutting the leaves in a completely randomized design with four replications in a 3 x 7 + 1 factorial design (three herbicides x seven periods – hours after application). Herbicides and doses tested were: glyphosate (1,440 g ha-1), 2,4-D (720 g ha-1) and a mixture of glyphosate + 2,4-D (1,080 + 720 g ha-1). The simulation of rain interfered negatively in the plant control with glyphosate. The control with the herbicide 2,4-D was affected only for the period of 2 hours. Periods of rain simulation did not influence the control of plants with a mixture of glyphosate + 2,4-D. For the study with the cutting of treated leaves, all treatments regardless of the period of cutting the leaves were influenced negatively in terms of plant control, the plants showing regrowth when treated with 2,4-D alone.

Glyphosate and imazamox translocation in water hyacinth plants / Translocação dos herbicidas glyphosate e imazamox em plantas de aguapé

Eichhornia crassipes is one of the main weeds found in aquatic environments, being undesirable for many activities. The aim of this study was to evaluate the translocation of glyphosate and imazamox in E. crassipes. Eight intervals were studied for cutting leaves that received herbicides: 2, 4, 6, 8, 12 and 24 hours after application (HAA), and a treatment with no cutting (untreated). The glyphosate dose was 2,160 g a.e. ha-1 (commercial product - Rodeo) + 0.5% v v-1 Aterbane adhesive spreader and imazamox at 290.4 g i.a. ha-1 (commercial product - Clearcast). The treatments were installed in a completely randomized design with four replications. Glyphosate showed a bad control for all the periods of leaf cutting. The imazamox did not provide control within 12 HAA, while from 24 HAA onward the control was effective. There was not a great mobility of the glyphosate molecule in water hyacinth plants, a period above 24 hours being needed for a satisfactory translocation. For imazamox at least 24 hours were needed after herbicide application for the translocation to occur along with subsequent control.

Entre as diversas plantas daninhas encontradas em ambientes aquáticos, a Eichhornia crassipes é uma das principais, sendo espécie indesejável para muitas atividades. O objetivo deste trabalho foi avaliar a translocação de glyphosate e imazamox em plantas de E. crassipes. Foram estudados oito intervalos de tempo para o corte das folhas que receberam os herbicidas: 2, 4, 6, 8, 12 e 24 horas após a aplicação (HAA) dos produtos, além de um tratamento sem corte das plantas (testemunha). Utilizaram-se glyphosate na dose de 2.160 g e.a. ha-1 (produto comercial - Rodeo) + 0,5% v v-1 do espalhante adesivo aterbane e imazamox na dose de 290,4 g i.a. ha-1 (produto comercial - Clearcast). Os tratamentos foram instalados em um delineamento experimental inteiramente casualizado com quatro repetições. O glyphosate apresentou um controle insatisfatório quando realizado os cortes das folhas em todos os períodos avaliados. O herbicida imazamox não proporcionou controle em até 12 HAA, enquanto que a partir de 24 HAA do herbicida o controle foi eficiente. Não houve uma grande mobilidade da molécula de glyphosate nas plantas de aguapé, sendo necessário um período superior a 24 horas para uma translocação satisfatória. Já para o imazamox um tempo de mínimo de 24 horas após a aplicação do herbicida foi suficiente para que ocorresse a translocação e um posterior controle.

Transcript profiling of expressed sequence tags from semimembranosus muscle of commercial and naturalized pig breeds.

In general, genetic differences across different breeds of pig lead to variation in mature body size and slaughter age. The Commercial breeds Duroc and Large White and the local Brazilian breed Piau are ostensibly distinct in terms of growth and muscularity, commercial breeds are much leaner while local breeds grow much slower and are fat type pigs. However, the genetic factors that underlie such distinctions remain unclear. We used expressed sequence tags (ESTs) to characterize and compare transcript profiles in the semimembranosus muscle of these pig breeds. Our aim was to identify differences in breed-related gene expression that might influence growth performance and meat quality. We constructed three non-normalized cDNA libraries from semimembranosus muscle, using two samples from each one, of these three breeds; 6902 high-quality ESTs were obtained. Cluster analysis was performed and these sequences were clustered into 3670 unique sequences; 24.7% of the sequences were categorized as contigs and 75.3% of the sequences were singletons. Based on homology searches against the SwissProt protein database, we were able to assign a putative protein identity to only 1050 unique sequences. Among these, 58.5% were full-length protein sequences and 17.2% were pig-specific sequences. Muscle structural and cytoskeletal proteins, such as actin, and myosin, were the most abundant transcripts (16.7%) followed by those related to mitochondrial function (12.9%), and ribosomal proteins (12.4%). Furthermore, ESTs generated in this study provide a rich source for identification of novel genes and for the comparative analysis of gene expression patterns in divergent pig breeds.

Gene expression in swine granulosa cells and ovarian tissue during the estrous cycle.

The components of the insulin-like growth factor (IGF) system appear to be involved in regulation of ovarian follicular growth and atresia in the pig. We investigated the expression pattern of mRNAs for IGF1 (IGF1), its binding proteins (IGFBP1, IGFBP2, IGFBP3, and IGFBP5), and epidermal growth factor in swine follicle cells and ovarian tissue throughout the estrous cycle using the real-time quantitative PCR technique. The results of gene expression were analyzed using linear regression with gene expression as a dependent variable and days of estrous cycle as an independent variable. Additionally, an analysis was made of the correlation of expression levels with plasma concentration of follicle-stimulating hormone, luteinizing hormone, estradiol-17ß, progesterone, and prolactin. Expression of mRNA of all of these genes was detected in granulosa cells and ovarian tissue. IGFBP3 mRNA showed a quadratic expression pattern (P ≤ 0.001) and was significantly and positively correlated with progesterone (r = 0.81; P ≤ 0.01) but negatively correlated with prolactin (r = -0.596; P ≤ 0.05). Expression of the other genes was unaffected by the stage of the estrous cycle. Real-time quantitative PCR effectively detected all transcripts, including the very low levels of IGFBP1 transcripts, and could be used for studies of follicle dynamics.

Efeito de diferentes herbicidas, doses e volume de calda na dessecação de milheto [Pennisetum Glaucum (L. Leek)] / Effect of different herbicides, doses and spray volume in the chemical management of pearl millet [Pennisetum Glaucum (L. Leek)]

Objetivou-se nesse trabalho avaliar a eficácia de diferentes herbicidas aplicados em pósemergência na de secação do milheto com diferentes doses e volumes de calda de aplicação. Para a dessecação do milheto foram utilizadas doses de 0, 25, 50 e 100% dos herbicidas glyphosate (1.440 g ha-1), em três formulações comerciais (Round up Original, Round up Transorb e Round up WG), paraquat + diuron (500 + 250 g ha-1) e amônio-glufosinato (800 g ha-1). Em todos os tratamentos foram utilizados dois volumes de aplicação (200 e 400 L ha-1). Manteve-se uma testemunha sem aplicação de herbicidas. Os efeitos dos tratamentos foram avaliados visualmente aos 3, 7, 14, 21 e 28 dias após aplicação. Ao final das avaliações foi determinada a massa seca das plantas. O estudo de campo foi instalado no delineamento em blocos casualizados. No final do estudo, aos 28 DAA, todos os tratamentos apresentaram controle total das plantas de milheto, independente da dose e do volume de aplicação, exceto para os tratamentos com paraquat + diuron e amônio-glufosinate na menor dose (25% da dose), nos dois volumes de aplicação, que apresentaram ainda controles considerados bons a muito bom, sendo que para esses dois herbicidas houve efeito do volume de calda, na qual o controle foi superior com o uso de volume menor.

The objective of this work was to evaluate the efficacy of different herbicides applied in post-emergence in the desiccation of pearl millet with different doses and spray volumes. For the desiccation, doses at 0, 25, 50 and 100% of the herbicide glyphosate (1,440 g ha-1) were used in 3 formulations (Original Round up, Round up Transorb and Round up WG), paraquat + diuron (500 + 250 g ha-1) and glufosinateammonium (800 g ha-1). In all treatments, 2 spray volumes (200 and 400 l ha-1) were used, and there was 1 treatment (control) without the application of herbicides. The treatment effects were evaluated visually at 3, 7, 14, 21 and 28 days after application (DAA). At the end of the evaluations, the dry mass of the plants was determined. The field trial was carried out in randomized block design. At the end of the study, at 28 DAA, all treatments presented total control of plant pearl millet, regardless of the dose and spray volume, except the treatments with paraquat + diuron and glufosinate-ammonium at the lower dose (25%), in 2 spray volumes, which nevertheless showed good and very good control, these two herbicides presenting an effect of volume, in which the control was higher with the use of smaller volume.

Candidate gene expression and intramuscular fat content in pigs.

Seventy-two pigs of three genetic groups (Brazilian indigenous breed Piau, Commercial line and Crossbred) of both sexes were slaughtered at four live weights (30, 60, 90 and 120 kg). Intramuscular fat (IMF) content in Longissimus dorsi muscle of each animal was extracted and correlated with candidate gene mRNA expression (ATN1, EEF1A2, FABP3, LDLR, MGP, OBSCN, PDHB, TRDN and RYR1). Within slaughter weight of 120 kg, Piau and Crossbred pigs showed higher IMF content (p < 0.05) than commercial animals, with 2.48, 2.08 and 1.00% respectively. Barrows presented higher values of IMF (p < 0.05) than gilts (1.54 and 1.30% respectively). Gene expression of EEF1A2, FABP3, LDLR, OBSCN, PDHB, TRDN and RYR1 were correlated with IMF (p < 0.05) using the whole dataset. For Piau data only, expression of FABP3, LDLR, MGP, OBSCN, PDHB, TRDN and RYR1 showed correlation with IMF (p < 0.05). Genes that have important roles in lipid transportation inside the cell (FABP3) and tissues (LDLR) showed correlation with IMF of, respectively, 0.68 and 0.63 using the whole data set, and 0.90 and 0.91 using data from Piau animals. The highly positive correlation of the LDLR and FAPB3 expression with IMF content may confirm that these genes are important for fat deposition in the porcine L. dorsi muscle.

Sclerocorneal limbal stem cell autograft transplantation in dogs

The effects of sclerocorneal limbal stem cell autograft transplantation in dogs with corneal wounds were studied. Eighteen dogs were divided in two groups (GI and GII). The animals of GI (n=12) underwent limbal transplantation 30 days after the destruction of limbal stem cells. The dogs of GII (n=6) only underwent destruction of stem cells (control group). Light microscopy examination of the right eye was performed on days 3, 7, 14, 30, 60, and 120 after limbal transplantation (GI), and on days 33, 37, 44, 60, 90, and 150 after limbal destruction (GII). Results showed a complete destruction of limbal stem cells with loss of corneal transparency. Limbal transplantation prevented conjunctivalization in grafted area. Corneal vascularization and a 360º corneal conjunctivalization were noted in the control dogs (GII). Corneal transparency was restored from day 60th after surgery. Histological examination did not distinguish the transition between the graft and the normal corneal epithelium at anytime. Goblet cells were found in control animals (GII) on 33, 37, 60, and 150 days, whereas a single grafted dog (GI) presented a few goblet cells on day 60th post-transplantation. Limbal autograft transplantation was effective in restoring corneal clarity with no development of ocular complications

Avaliaram-se os efeitos do transplante de células tronco autógenas do limbo esclerocórneo de cães, sobre lesões córneo-limbais. Empregaram-se 18 cães, distribuídos em dois grupos, GI e GII. Nos animais do GI (n=12), foram realizados transplantes de limbo, após 30 dias da destruição das células tronco-límbicas. Nos do GII (n=6), realizou-se apenas a destruição do limbo (controle). Aos 3, 7, 15, 30, 60 e 120 dias do transplante de limbo (GI) e aos 33, 37, 45, 60, 90 e 150 dias da destruição do limbo (GII), os olhos foram coletados por enucleação subconjuntival, para estudos em microscopia de luz. A destruição do limbo resultou em completa excisão das células tronco, com perda da transparência corneal. O transplante do limbo evitou a conjuntivalização na área em que foi realizado. Os animais do grupo-controle manifestaram conjuntivalização em 360º e vascularização corneal. Na anatomopatologia, em nenhum dos períodos foi possível distinguir o enxerto do epitélio corneal normal. As células caliciformes foram observadas nos animais do GII, nos períodos 33, 37, 60, 150 dias. No GI, apenas um cão manifestou células caliciformes de forma discreta, aos 60 dias do transplante. O transplante autógeno foi eficiente em possibilitar a melhoria da transparência córnea, sem intercorrências oculares

Excision of sclerocorneal limbus in dogs and resulting clinical events: study of an experimental model

As conseqüências da completa remoção do epitélio do limbo esclerocorneano foram avaliadas, bem como o modelo utilizado para tal, em que se empregou o n-heptanol e a ceratectomia superficial. O n-heptanol e a ceratectomia associados possibilitaram a completa excisão das células-tronco. Da destruição dessas células decorrem alterações tectônicas significativas factíveis para estudos dos transplantes de córnea.

The consequences of complete removal of the sclerocorneal limbus epithelium were evaluated, as well as the model for such, in which n-heptanol and superficial keratectomy were used. The association of n-heptanol and keratectomy allowed a complete excision of stem cells. Moreover, the destruction of these cells led to significant tectonic alterations that could be employed as a feasible model for clinical experimentation of corneal transplantation.

Effects of low-intensity pulsed ultrasound on wound healing in corneas of dogs following keratoplasty.

The effects of low-intensity pulsed ultrasound on wound healing were evaluated at the graft-cornea transition in dogs following lamellar keratoplasty using tunica vaginalis preserved in 98% glycerin. Twenty-one dogs were subdivided into three groups of seven animals. The first group (W/US) received daily treatment of low-intensity pulsed ultrasound (20 mW/cm2) for 15 min for the first 10 days post surgery. The second group (N/US) was submitted to the same procedure but with the ultrasound apparatus turned off. The third group, the control (CO), underwent the surgical procedure only. The animals were clinically evaluated during the initial (1-15 days), intermediate (16-30 days) and late (31-120 days) postoperative period. The corneas were evaluated by light microscopy at 1, 3, 7, 15, 30, 60 and 120 days after surgery. Clinically, there were no differences which would promote an advantage to any of the treatments. Light microscopy, however, revealed more extensive vascularization and more advanced wound healing in the W/US group, as well as a tendency towards early graft incorporation. Based on the present results, low-intensity pulsed ultrasound shows advantages, especially in situations where trophic support is a mandatory condition, facilitating better graft incorporation and rapid recovery of stromal organization.