Eggshell translucency: its relationship with specific gravity and eggshell color and its influence on broiler egg weight loss, hatchability, and embryonic mortalities.

Eggshell quality is among the most important factors affecting hatchability in broiler breeders, and therefore several methods for its assessment are available in the poultry industry. Among them, eggshell translucency has received special attention in recent years due to its connection with ultrastructural disorganization of the shell layers. However, there is very limited data on the impact of translucency on hatching eggs and on the possible links between this trait and specific gravity (SG) or shell color. Thus, our study investigated associations and interactions between eggshell translucency, SG, and color on incubation parameters of eggs from the same breeding flock (Ross 308AP, 51 wk of age). To this end, light and dark eggs within 5 different SG categories (≥1.065, 1.070, 1.075, 1.080, and ≤1.085) were selected from 15,976 eggs, graded into 3 translucency scores, and later incubated to evaluate egg weight loss, hatchability and embryonic mortalities. In general, translucency scores were evenly distributed within SG categories (χ2 [8, N = 1,138] = 13.67, P = 0.090) and color (χ2 [2, N = 1,138] = 4.93, P = 0.084). No interactions between eggshell translucency and SG or between translucency and color were found for the analyzed variables. An interaction was observed between SG and eggshell color for the variable egg weight loss, where the light-shelled eggs, in most SG categories lost more weight throughout incubation than dark eggs. Eggshell translucency affected egg weight loss, hatchability, and embryonic mortality on 11 to 18 d of incubation, with highly translucent eggs showing the worst results. At the same time, eggs with SG lower than 1.070 displayed the greatest weight loss, lowest hatchability, and highest contamination. We found no influence of eggshell color on weight loss or hatchability, but light-shelled eggs exhibited higher late embryonic mortality. Together, these data suggest that despite its effects on certain hatching parameters, shell translucency bears no relationship to SG or color.

Ex ovo electroporation of early chicken embryos.

The chick embryo is a favored model for developmental studies owing to its accessibility and ease of manipulation. Ex ovo electroporation provides a highly efficient method for screening perturbation phenotypes using a variety of reagents, including CRISPR and morpholinos. Additionally, the chick system lends itself well to rapid medium-throughput enhancer screening. Constructs facilitating tissue-specific protein pull-down can also be transfected using this protocol. Furthermore, bilateral electroporation with control and experimental reagents provides a robust assay for accurately interpreting functional perturbations. For complete details on the use and execution of this protocol, please refer to Williams et al. (2019).

Characterising open chromatin in chick embryos identifies cis-regulatory elements important for paraxial mesoderm formation and axis extension.

Somites arising from paraxial mesoderm are a hallmark of the segmented vertebrate body plan. They form sequentially during axis extension and generate musculoskeletal cell lineages. How paraxial mesoderm becomes regionalised along the axis and how this correlates with dynamic changes of chromatin accessibility and the transcriptome remains unknown. Here, we report a spatiotemporal series of ATAC-seq and RNA-seq along the chick embryonic axis. Footprint analysis shows differential coverage of binding sites for several key transcription factors, including CDX2, LEF1 and members of HOX clusters. Associating accessible chromatin with nearby expressed genes identifies cis-regulatory elements (CRE) for TCF15 and MEOX1. We determine their spatiotemporal activity and evolutionary conservation in Xenopus and human. Epigenome silencing of endogenous CREs disrupts TCF15 and MEOX1 gene expression and recapitulates phenotypic abnormalities of anterior-posterior axis extension. Our integrated approach allows dissection of paraxial mesoderm regulatory circuits in vivo and has implications for investigating gene regulatory networks.

A cell atlas of the chick retina based on single-cell transcriptomics.

Retinal structure and function have been studied in many vertebrate orders, but molecular characterization has been largely confined to mammals. We used single-cell RNA sequencing (scRNA-seq) to generate a cell atlas of the chick retina. We identified 136 cell types plus 14 positional or developmental intermediates distributed among the six classes conserved across vertebrates - photoreceptor, horizontal, bipolar, amacrine, retinal ganglion, and glial cells. To assess morphology of molecularly defined types, we adapted a method for CRISPR-based integration of reporters into selectively expressed genes. For Müller glia, we found that transcriptionally distinct cells were regionally localized along the anterior-posterior, dorsal-ventral, and central-peripheral retinal axes. We also identified immature photoreceptor, horizontal cell, and oligodendrocyte types that persist into late embryonic stages. Finally, we analyzed relationships among chick, mouse, and primate retinal cell classes and types. Our results provide a foundation for anatomical, physiological, evolutionary, and developmental studies of the avian visual system.

The evolutionary relationships of organisms and of genes have long been studied in various ways, including genome sequencing. More recently, the evolutionary relationships among the different types of cells that perform distinct roles in an organism, have become a subject of inquiry. High throughput single-cell RNA sequencing is a technique that allows scientists to determine what genes are switched on in single cells. This technique makes it possible to catalogue the cell types that make up a tissue and generate an atlas of the tissue based on what genes are switched on in each cell. The atlases can then be compared among species. The retina is a light-sensitive tissue that animals with a backbone, called vertebrates, use to see. The basic plan of the retina is very similar in vertebrates: five classes of neurons ­ the cells that make up the nervous system ­ are arranged into three layers. The chicken is a highly visual animal and it has frequently been used to study the development of the retina, from understanding how unspecialized embryonic cells become neurons to examining how circuits of neurons form. The structure and role of the retina have been studied in many vertebrates, but detailed descriptions of this tissue at the molecular level have been largely limited to mammals. To bridge this gap, Yamagata, Yan and Sanes generated the first cell atlas of the chicken retina. Additionally, they developed a gene editing-based technique based on CRISPR technology called eCHIKIN to label different cell types based on genes each type switched on selectively, providing a means of matching their shape and location to their molecular identity. Using these methods, it was possible to subdivide each of the five classes of neurons in the retina into multiple distinct types for a total of 136. The atlas provided a foundation for evolutionary analysis of how retinas evolve to serve the very different visual needs of different species. The chicken cell types could be compared to types previously identified in similar studies of mouse and primate retinas. Comparing the relationships among retinal cells in chickens, mice and primates revealed strong similarities in the overall cell classes represented. However, the results also showed big differences among species in the specific types within each class, and the genes that were switched on within each cell type. These findings may provide a foundation to study the anatomy, physiology, evolution, and development of the avian visual system. Until now, neural development of the chicken retina was being studied without comprehensive knowledge of its cell types or the developmentally important genes they express. The system developed by Yamagata, Yan and Sanes may be used in the future to learn more about vision and to investigate how neural cell types evolve to match the repertoire of each species to its environment.

Neither altered incubation temperature during fetal development nor preferred rearing temperature improves leg bone characteristics of broilers.

The present study evaluated whether broiler femoral and tibiotarsal characteristics (as assessed at slaughter age) could be improved if birds were reared under their preferred temperature and whether continuous high or low incubation temperature during the fetal period improves bone characteristics of broilers reared under heat stress or thermal preference. Broiler breeder eggs were incubated from day 13 until hatching under cold (36 °C), control (37.5 °C), or hot (39 °C) temperatures. Under these conditions, the eggshell temperatures were 37.4 ± 0.1°C, 37.8 ± 0.15°C, and 38.8 ± 0.3°C, respectively. Then, broiler chicks were reared under control, preferred (determined previously in thermal preference test), or high temperatures. At day 42 of age, the broilers were weighed and euthanized, and femora and tibiotarsi collected to measure weight, length, diaphysis perimeter, breaking strength, maximum flexion, rigidity, ash, phosphorus, and calcium. Rearing under the preferred temperature did not affect broiler body weight or femoral and tibiotarsal characteristics (P > 0.05). In contrast, high rearing temperature, decreased the body weight, mineral contents of both bones, femoral breaking strength, and tibiotarsal rigidity (P < 0.05). Regarding incubation temperature effects, egg exposure to cold and hot temperatures during the fetal period minimized or avoided a few effects of high rearing temperature, such as those on femoral and tibiotarsal morphological characteristics, mineral composition, and mechanical properties at slaughter age (P < 0.05), but not all. In conclusion, rearing under the preferred broiler temperature did not improve the bone characteristics, and the negative effects of high rearing temperature on bone development were minimized but not completely prevented by high or low temperature incubation during the fetal period.

Non-Invasive PPG-Based System for Continuous Heart Rate Monitoring of Incubated Avian Embryo.

The chicken embryo is a widely used experimental animal model in many studies, including in the field of developmental biology, of the physiological responses and adaptation to altered environments, and for cancer and neurobiology research. The embryonic heart rate is an important physiological variable used as an index reflecting the embryo's natural activity and is considered one of the most difficult parameters to measure. An acceptable measurement technique of embryonic heart rate should provide a reliable cardiac signal quality while maintaining adequate gas exchange through the eggshell during the incubation and embryonic developmental period. In this paper, we present a detailed design and methodology for a non-invasive photoplethysmography (PPG)-based prototype (Egg-PPG) for real-time and continuous monitoring of embryonic heart rate during incubation. An automatic embryonic cardiac wave detection algorithm, based on normalised spectral entropy, is described. The developed algorithm successfully estimated the embryonic heart rate with 98.7% accuracy. We believe that the system presented in this paper is a promising solution for non-invasive, real-time monitoring of the embryonic cardiac signal. The proposed system can be used in both experimental studies (e.g., developmental embryology and cardiovascular research) and in industrial incubation applications.

Effects of thermal manipulation of eggs on the response of jejunal mucosae to posthatch chronic heat stress in broiler chickens.

In this study, the aim was to investigate effects of chronic heat stress (CHS) on the mRNA levels of proinflammatory cytokines (interleukin [IL]-6, IL-8, IL-1ß, and tumor necrosis factor alpha [TNF-α]), toll-like receptors (TLR2 and TLR4), heat shock proteins (Hsp70, heat shock transcription factor [HSF]-1, and HSF3) and antioxidant enzymes (catalase, glutathione peroxidase, NADPH oxidase, and superoxide-dismutase) in the jejunal mucosae of broiler chickens subjected to thermal manipulation (TM) during embryogenesis. TM was carried out at 39°C and 65% relative humidity (RH) for 18 h daily from embryonic days 10 to 18. Control group was incubated at 37.8°C and 56% RH. CHS was induced by raising the temperature to 35°C for 7 D throughout posthatch days 28 to 35. On post-hatch-day 28 (day zero of CHS) and after 1, 3, 5, and 7 D of CHS, the jejunal mucosae were collected from both groups to evaluate the mRNA levels by real-time reverse transcription-PCR analysis. On day zero of CHS, the mRNA levels of antioxidant enzymes, TLRs, HSF3, IL-1ß, and TNF-α were not significantly different between TM and control groups, while the levels of IL-6, IL-8, and HSF1 were lower and the level of Hsp70 was higher in TM. However, during CHS, the mRNA levels of antioxidant enzymes, IL-1ß, TNF-α, TLR4, and HSF1 were significantly lower in TM than in controls, while the levels of TLR2 and IL-8 were significantly higher in TM than in controls. In addition, TM led to significant increase of mRNA levels of IL-6 and HSF3 after 1 D and Hsp70 after 3 D of CHS and to significant decrease of mRNA levels of IL-6 after 3 and 5 D, HSF3 after 7 D, and Hsp70 after 5 D of CHS. Results of this study suggest that TM led to altered posthatch antioxidant, immunological, and Hsp response to CHS in the jejunal mucosae of broiler chickens, probably indicating that TM may mitigate the adverse effects of CHS.

Chicken embryos can maintain heart rate during hypoxia on day 4 of incubation.

Acute exposure to hypoxic conditions is a frequent natural event during the development of bird eggs. However, little is known about the effect of such exposure on the ability of young embryos in which cardiovascular regulation is not yet developed to maintain a normal heart rate (HR). To address this question, we studied the effect of 10-20 min of exposure to moderate or severe acute hypoxia (10% or 5% O2, respectively) on the HR of day 4 (D4) chicken embryos. In ovo, video recording of the beating embryo heart inside the egg revealed that severe, but not moderate, hypoxia resulted in significant HR changes. The HR response to severe hypoxia consisted of two phases: the first phase, consisting of an initial decrease in HR, was followed by a phase of partial HR recovery. Upon the restoration of normoxia, after an overshoot period of a few minutes, the HR completely recovered to its basal level. In vitro (isolated heart preparation), the first phase of the HR response to severe hypoxia was strengthened (nearly complete heart silencing) compared to that in ovo, and the HR recovery phase was greatly attenuated. Furthermore, neither an overshoot period nor complete HR recovery after hypoxia was observed. Thus, the D4 chicken embryo heart can partially maintain its rhythm during hypoxia in ovo, but not in vitro. Some factors from the egg, such as catecholamines, are likely to be critical for avian embryo responding to hypoxic condition and survival.

Disconnect between the developing eye and craniofacial prominences in the avian embryo.

In the amniote embryo, the upper jaw and nasal cavities form through coordinated outgrowth and fusion of craniofacial prominences. Adjacent to the embryonic prominences are the developing eyes, which abut the maxillary and lateral nasal prominences. The embryos of extant sauropsids (birds and nonavian reptiles) develop particularly large eyes in comparison to mammals, leading researchers to propose that the developing eye may facilitate outgrowth of prominences towards the midline in order to aid prominence fusion. To test this hypothesis, we performed unilateral and bilateral ablation of the developing eyes in chicken embryos, with the aim of evaluating subsequent prominence formation and fusion. Our analyses revealed minor interaction between the developing craniofacial prominences and the eyes, inconsequential to the fusion of the upper beak. At later developmental stages, the skull exhibited only localized effects from missing eyes, while geometric morphometrics revealed minimal effect on overall shape of the upper jaw when it develops without eyes. Our results indicate that the substantial size of the developing eyes in the chicken embryo exert little influence over the fusion of the craniofacial prominences, despite their effect on the size and shape of maxillary prominences and components of the skull.

Label-free proteomic analysis reveals the differentiation between unfertilized and fertilized Beijing-You chicken eggs.

Egg fertilization is a dynamic process, including varieties of biochemical changes. To better understand the molecular mechanisms during the egg embryo development, the objective of this study was to quantify protein expression changes between fertilized and unfertilized Beijing-You chicken eggs using label-free liquid chromatography-tandem mass spectrometry method. The results showed that a total of 1241 proteins were identified from fertilized and unfertilized eggs, 229 proteins were observed difference in fertilized eggs (p < 0.05) compared with that in unfertilized eggs. The expressions of 86 proteins were up-regulated and 48 proteins were down-regulated in fertilized eggs. STRING database analysis and Gene Ontology analysis results showed that these differentially expressed proteins significantly interacted and were involved in lipid transport and inflammatory response biological processes. The mRNA and protein expression levels of most differentially expressed proteins Apolipoprotein B, Fibrinogen alpha chain, Transferrin receptor protein 1, Phospholipid transfer protein and Vimentin were validated by RT-PCR and western blot. These results could provide possible novel insights for the molecular mechanism of egg fertilization.

Smartphone-Based Device for Non-Invasive Heart-Rate Measurement of Chicken Embryos.

Heart rate (HR) is an important parameter in the study of the developmental physiology of chicken embryos and a crucial indicator of dead or live embryo grading in artificial incubation processes. A non-invasive HR measurement technique is required for long-term and routine HR assessment with minimal influence on embryo development. Accordingly, in this study, a non-invasive HR measurement technique of chicken embryos using a smartphone is demonstrated. The detection method of the proposed device is based on the photoplethysmography principle in which a smartphone camera is used for video recording, and the chicken embryonic HR is obtained from the recorded video images using a custom Android application. We used a smartphone to measure the embryonic HR of 60 native chicken eggs and found that it can measure the chicken embryonic HR from day 4 to day 20. The proposed smartphone HR device will be beneficial for scientific research and industrial applications. With internet connectivity, users can utilize their smartphone to measure the HR, display, share, and store the results.

Differential proteomic analysis revealed crucial egg white proteins for hatchability of chickens.

For healthy development, an avian embryo needs the nutritional and functional molecules maternally deposited in avian eggs. Egg white not only provides nutritional components but also exhibits functional properties, such as defenses against microbial invasion. However, the roles of the more detailed messages in embryo development remain unclear. In this study, a tandem mass tag labeling quantitation approach was used to innovatively identify the differential proteins in the egg whites of fresh eggs produced by hens with divergent high/low hatchability and in the egg whites of embryonated eggs with healthy and dead embryos. A total of 378 proteins were quantified in egg white, which is the most complete proteome identified for egg white to date, and up to 102 differential proteins were identified. GO enrichment, pathway, and hierarchical clustering analysis revealed some of the differential proteins that are the main participants in several biological processes, including blood coagulation, intermediate filament, antibacterial activity, and neurodevelopment. A list of 11 putative protein biomarkers, such as keratin (KRT19, KRT12, KRT15, and KRT6A), which is involved in cell architecture, and fibrinogen (fibrinogen alpha chain, fibrinogen beta chain, and fibrinogen gamma chain), which is related to blood coagulation, were ultimately screened. The current study screened egg white proteins that can predict low hatchability and embryonic death and deciphered the role of these proteins in embryonic development, which is meaningful for the comprehensive understanding of embryonic growth.

Migration of chicken egg-white protein ovalbumin-related protein X and its alteration in heparin-binding affinity during embryogenesis of fertilized egg.

Ovalbumin-related protein X (OVAX) is a 50 kDa egg-white protein which has heparin-binding affinity. In this study, migration of OVAX and its heparin-binding performance during embryogenesis of fertilized egg were investigated to explore a possible involvement in chick embryo development. Western blotting of egg yolks at different stages using an anti-OVAX antibody showed that OVAX accumulates in yolk during incubation of fertilized egg. Immunohistochemical analysis of embryo resided in 10-day-incubated eggs showed that OVAX existed in almost all tissues of the embryo. These suggest that OVAX is incorporated from egg white into the embryo through yolk sac. Heparin-sepharose chromatography, isothermal titrating calorimetry using fondaparinux as a ligand, and zeta potential measurement indicated that OVAX retained the heparin-binding affinity (Kd = 0.185 ± 0.037) even after 10 D incubation of fertilized egg, although the affinity was slightly decreased during egg incubation because of acidification of molecular surface charge. In conclusion, although heparin-binding ability of egg-white OVAX slightly decreases during embryogenesis, OVAX incorporated into embryo can retain heparin-binding affinity. Our findings provide a new insight that OVAX participates in the functions of heparin during embryogenesis.

Embryonic Chicken (Gallus gallus domesticus) as a Model of Cardiac Biology and Development.

Cardiovascular disease remains one of the top contributors to morbidity and mortality in the United States. Increasing evidence suggests that many processes, pathways, and programs observed during development and organogenesis are recapitulated in adults in the face of disease. Therefore, a heightened understanding of cardiac development and organogenesis will help increase our understanding of developmental defects and cardiovascular diseases in adults. Chicks have long served as a model system in which to study developmental problems. Detailed descriptions of morphogenesis, low cost, accessibility, ease of manipulation, and the optimization of genetic engineering techniques have made chicks a robust model for studying development and make it a powerful platform for cardiovascular research. This review summarizes the cardiac developmental milestones of embryonic chickens, practical considerations when working with chicken embryos, and techniques available for use in chicks (including tissue chimeras, genetic manipulations, and live imaging). In addition, this article highlights examples that accentuate the utility of the embryonic chicken as model system in which to study cardiac development, particularly epicardial development, and that underscore the importance of how studying development informs our understanding of disease.

Field study on the factors affecting egg weight loss, early embryonic mortality, hatchability, and chick mortality with the use of classification tree technique.

The aim of this research was to study the factors affecting hatchability, percentage of egg weight loss during incubation, early embryonic mortality, and chick mortality in the first seven days of life in the field. This study was performed on 20,817,600 hatching eggs originating from 7 different breeder flocks. The factors responsible for variability of investigated traits were the following: genotype (ROSS 308, ROSS PM3), hen age (25 to 30, 31 to 45, 46 to 50, and 51 to 60 wk), egg storage time (0 to 13 d), and setter and hatcher type (Digital, Airstreamer, Vision, Focus, Biostreamer), which were determined using the classification tree technique. Statistical calculations were performed using R software, version 3.4.3. Moreover, 2-way analysis of variance followed by Duncan's multiple comparison test was performed. The breeder flock age and egg storage time were the most important factors responsible for variability in the percentage of fertile hatchability; however, setter and hatcher type also affected the level of this trait. The highest hatchability was observed for eggs laid by hens aged 31 to 50 wk and stored up to 6 d. Genotype was the most important determinant of the percentage of egg weight loss and that Ross 308 eggs lose less weight when compared to Ross PM3 ones. Egg storage time was the most important factor, which affected early embryonic mortality. The present study has shown that the 2 main factors, i.e., breeder flock age and egg storage time, that affected hatchability have also influenced chick mortality. Moreover, it was found that both hatcher and setter type affected chick mortality in the first 7 d of life. The importance of this research lies in the fact that besides showing the most important factors affecting hatching process, we were also able to suggest how to adjust the management decisions at commercial hatcheries in order to increase production results in different egg sets, which vary in respect of breeder flock age, egg storage time, setter, and hatcher type and genotype.

Effect of thermal manipulation of broilers embryos on the response to heat-induced oxidative stress.

Effects of embryonic thermal manipulation (TM) on mRNA expressional levels and total antioxidant capacity of genes associated with heat-induced oxidative stress (NOX4, GpX2, SOD2, catalase, and AvUCP) in 2 breeds of broiler chicken were investigated. Fertile Cobb and Hubbard eggs (n = 1,200) were divided into 4 treatment groups: Cobb control, Cobb TM, Hubbard control, and Hubbard TM. Control groups were maintained under standard conditions (37.8°C; 56% relative humidity), whereas TM groups were incubated at 39°C and 65% relative humidity for 18 h a day from embryonic days (ED) 10 to 18. On post-hatch day 28, the broilers were subject to acute heat stress (AHS) at 40°C for 7 h. At certain intervals (0, 1, 3, 5, and 7 h), 12 chickens from each of the 4 groups were humanely euthanized, and liver samples were immediately isolated. During AHS, in both breeds, the mRNA expression levels of NOX4, GPx2, SOD2, and catalase in TM chickens were significantly lower than in controls, but AvUCP mRNA expression in the TM group was higher. The total antioxidant capacity and activity of superoxidase dismutase and catalase were significantly lower in the TM than in the control group in both breeds. The results of this study suggest that TM has a long-lasting effect on the acquisition of thermotolerance in 2 broiler chicken breeds as indicated by the reduction of system genes associated with heat-induced oxidative stress.

Effect of vitamin E in ovo feeding to broiler embryos on hatchability, chick quality, oxidative state, and performance.

The objective of the present study was to ascertain the effect of in ovo feeding of vitamin E (VE) on the incubation results, quality, and oxidative state of newborn chicks and on the initial performance results. The design consisted of randomized blocks with treatments at different levels of VE (0.0, 27.5, 38.5, 49.5, and 60.4 IU). On 17.5 d of embryonic development, 780 eggs underwent in ovo injection using a manual needle. VE supplementation of 60.4 IU provided the highest hatching rate (P < 0.05) and shortest hatch window (P < 0.05). Better results regarding chick physical quality were observed in groups supplemented with VE (body weight, length, newborn chick quality score) and higher chick weight/egg weight ratios (P < 0.05). VE inoculation did not have any effect on the chicks' immunological system (P > 0.05). Greater development of the small intestine (intestine weight/yolk free chick weight and higher villi in duodenum) and better feed conversion over all periods studied (1 to 7, 1 to 14, and 1 to 21 d) were observed among chicks that received in ovo VE supplementation (P < 0.05). The total protein concentrations in the liver and striated breast skeletal muscle tissue were highest in chicks that received 60.4 IU of VE (P < 0.05). The highest catalase activity was observed in the livers of newborn chicks supplemented with 60.4 IU of VE (P < 0.05). It was concluded that in ovo VE supplementation improved the chicks' oxidative state, which led to improvements in incubation results, chick quality and performance results.

Effects of pre-hatch thermal manipulation and post-hatch acute heat stress on the mRNA expression of interleukin-6 and genes involved in its induction pathways in 2 broiler chicken breeds.

In response to heat stress, interleukin-6 (IL-6) expression is upregulated in broiler chickens. The main aim of the present study was to evaluate the cumulative effects of thermal manipulation (TM) and subsequent acute heat stress (AHS) on the mRNA expression of IL-6 and genes involved in its induction pathways. The studied genes include IL-6, IL-1ß, TNF-α, TLR2, TLR4, NFκB50, NFκB65, Hsp70, and HSF3 in the spleen and liver tissues. TM was carried out at 39°C for 18 h and 65% relative humidity during days 10 to 18 of embryonic development, while AHS was stimulated by raising the temperature to 40°C for 7 h on post-hatch day 28. During AHS at 0, 1, 3, 5, and 7 h, the spleen and liver were collected from all groups to measure the mRNA expression by relative-quantitation real-time RT-PCR, and the blood was collected to measure plasma IL-6 level. TM significantly reduced Tc during AHS for both breeds from 1 to 7 h time intervals. TM resulted in enhanced basal mRNA expression of IL-6, HSF3, and Hsp70, but decreased the basal mRNA level of TLR4. During heat stress, TM enhanced the expression dynamics of Hsp70, HSF3, IL-6, IL-1ß, TNF-α, TLR2, TLR4, NFκB50, and NFκB65. The results of the current study indicate that TM enhanced the heat tolerance through improving the protective immunological response to heat stress by enhancing the expression of IL-6 and modulating the expression of genes important in its induction pathways.

Sulforaphane Rescues Ethanol-Suppressed Angiogenesis through Oxidative and Endoplasmic Reticulum Stress in Chick Embryos.

Our previous study showed that ethanol exposure inhibited embryonic angiogenesis mainly due to the excessive stimulation of reactive oxygen species (ROS) production. In this study, we investigated whether sulforaphane (SFN), a known dietary bioactive compound, could ameliorate ethanol-suppressed angiogenesis using chick embryo angiogenesis models. Using chick yolk sac membrane (YSM) and chorioallantoic membrane (CAM) models, we demonstrated that administration of low concentrations of SFN (2.5-10 µM) alone increased angiogenesis, but high concentrations of SFN (20-40 µM) inhibited angiogenesis. SFN administration alleviated ethanol-suppressed angiogenesis and angiogenesis-related gene expression in both angiogenesis models. Ethanol exposure caused cell apoptosis in chick CAM, and the cell apoptosis could be remitted by administration of SFN. Subsequently, we demonstrated that the ethanol-induced increase in production of ROS and reduction of antioxidant enzymes' activity were partially rescued by SFN. Similar results were obtained in endoplasmic reticulum (ER) stress determination, indicated by ATF6 and GRP78 expression or thapsigargin-induced ER stress in the presence or absence of SFN. Taken together, our experiments show that SFN administration can ameliorate ethanol-suppressed embryonic angiogenesis, and this is mainly achieved by alleviating excessive ROS production and ER stress. This study suggests that SFN, in appropriate concentrations, could be a potential candidate compound for preventing the negative impact of alcohol on angiogenesis.

Short periods of incubation, egg turning during storage and broiler breeder hens age for early development of embryos, hatching results, chicks quality and juvenile growth.

An effect of modification of storage conditions of the eggs of broiler breeder flocks at the age of 49-, 52- and 70-, 73-wks of life on an early embryonic development, hatching time and synchronization, hatchability rates, chicks quality and broiler growth was investigated. The eggs were divided into 4 experimental groups: COI = eggs storage 5 d, at turning every 12 h; NSP = eggs storage 12 d, at turning every 12 h; SPIDES = were treated with 4 h pre-incubation at 30°C and 50-55% air humidity, delivered at 5 and 10 d over of 12 d of storage, and turning every 12 h; NCOI = eggs storage 12 d, no turning and no pre-incubation. Eggs from older hens were characterized by poorer hatchability and poorer chicks quality. The use of 2 × 4 h pre-incubation in 12 d of eggs storage could have an effect on the initial acceleration of embryonic development in eggs of young hens, contributing to the alignment of embryos development in eggs from young and older hens to 72 h of incubation. Pre-incubation had no effect on the length of incubation period, hatching window, but it increased the hatchability of the set and apparently fertilized eggs and decreased the number of eggs not hatched, and also improved chicks quality. Eggs turning by 90° every 12 h during the storage positively affected the embryonic development, shortening the incubation time and the quality of chicks, but had no effect on hatchability rates and body weight in 42 d of life. Based on the obtained results, it can be concluded that the applied modifications can be effective in counteracting the negative effects of storage of hatching eggs from both young and older birds.