Pigmentation of the aortic and pulmonary valves in C57BL/6J x Balb/cByJ hybrid mice of different coat colours

Neural crest‐derived melanocytes have been recorded in several parts of the mammalian heart but not in the pulmonary valve. We report here the presence of melanin‐containing cells in the leaflets (cusps) of both the aortic and pulmonary valves. A total of 158 C57BL/6J x Balb/cByJ hybrid mice exhibiting four coat colours, namely black, white, agouti and non‐agouti brown, were examined. We sought for any relationship between the presence of melanocytes in the valves and the coat colour of the animals. The pigmentation levels of the leaflets were accomplished using a scale of five pigment intensities. White mice lacked pigment in the heart. In 10.5% of the remaining animals, there were melanocytes in the pulmonary valve leaflets. Thus, this is the first study to report the presence of such cells in the pulmonary valve of mammals. Melanocytes occurred in the leaflets of the aortic valves of 87.2% of mice. The incidence of melanocytes and the pigmentation level of the leaflets did not statistically differ according to the coat colours of the animals. This disagrees with previous observations, indicating that the amount of melanocytes in the heart reflects that of the skin. The incidence and distribution of melanocytes in aortic and pulmonary valves are consistent with the notion that the formation of the arterial valves is mediated by specific subpopulations of neural crest cells. We hypothesize that melanocytes, even not producing melanin, may be more frequent in the heart than previously thought, exerting presumably an immunological function.     

E12 technique: Conventional epoxy resin sheet plastination

Epoxy plastination techniques were developed to obtain thin transparent body slices with high anatomical detail. This is facilitated because the plastinated tissue is transparent and the topography of the anatomical structures well preserved. For this reason, thin epoxy slices are currently used for research purposes in both macroscopic and microscopic studies. The protocol for the conventional epoxy technique (E12) follows the main steps of plastination—specimen preparation, dehydration, impregnation and curing/casting. Preparation begins with selection of the specimen, followed by freezing and slicing. Either fresh or fixed (embalmed) tissue is suitable for epoxy plastination, while slice thickness is kept between 1.5 and 3 mm. Impregnation mixture is made of epoxy E12 resin plus E1 hardener (100 ppw; 28 ppw). This mixture is reactive and temperature sensitive, and for this reason, total impregnation time under vacuum at room laboratory temperature should not last for more than 20–24 hr. Casting of impregnated slices is done in either flat chambers or by the so‐called sandwich method in either fresh mixture or the one used for impregnation. Curing is completed at 40°C to allow a complete polymerization of the epoxy‐mixture. After curing, slices can be photographed, scanned or used for anatomical study under screen negatoscope, magnification glass or fluorescent microscope. Based on epoxy sheet plastination, many anatomical papers have recent observations of and/or clarification of anatomical concepts in different areas of medical expertice.     

In ovo injection of branched‐chain amino acids: Embryonic development,hatchability and hatching quality of turkey poults

In this study, the influence of a branched‐chain amino acid blend (BCAA composed of 3 l ‐leucine:1 l ‐valine:2 l ‐isoleucine) injected into the amniotic fluid was evaluated for embryonic growth, yolk‐sac (YS) utilization and development of gastrointestinal tract (GIT) and skeletal muscles of turkey embryos from day 24 of incubation (24E) to hatching, together with hatchability, poult quality and liver L* (lightness), a* (redness) and b* (yellowness) values at hatch. At day 22 of incubation, embryonated eggs (n = 240) were assigned to three treatments, that is, eggs were not injected (control, NC) or injected with 1.5 ml sterile solution with 0.9% salt (SA) or 0.2% BCAA blend (BCAAb). These solutions were injected manually into the amniotic fluid of the embryonated eggs. To determine weights and lengths (where appropriate) of the studied organs and tissues, four embryonated eggs and poults per treatment were selected at 24E and at hatch. While the BCAAb decreased the YS and embryo weight, hatchability and the liver L* value, it increased the weight and quality of poults and the weights of breast and thigh muscles at hatch. In conclusion, the in ovo feeding of the BCAA blend negatively affected hatchability but positively affected hatching weight and poult quality by improving development of skeletal muscles and by regulating energy metabolism.     

Whole‐genome analysis identifying candidate genes of altitude adaptive ecological thresholds in yak populations

The domestic yak (Bos grunniens) is an iconic symbol of animal husbandry on the Qinghai–Tibet Plateau. Long‐term domestication and natural selection have led to a wide distribution of yak, forming many ecological populations to adapt to the local ecological environment. High altitude is closely related to oxygen density, and it is an important environmental ecological factor for biological survival and livestock production. The aim of the present study was to perform a preliminary analysis to identify the candidate genes of altitude distribution adapted ecological thresholds in yak using next‐generation sequence technology. A total of 15,762,829 SNPs were obtained from 29 yaks with high‐ and low‐altitude distribution by genome‐wide sequencing. According to the results of the selective sweep analysis with FST and ZHp, 21 candidate genes were identified. 14 genes (serine/threonine protein kinase TNNI3K, TEN1, DYM, ITPR1, ZC4H2, KNTC1, ADGRB3, CLYBL, TANGO6, ASCC3, KLHL3, PDE4D, DEPDC1B and AGBL4) were grouped into 32 Gene Ontology terms, and four genes (RPS6KA6, ITPR1, GNAO1 and PDE4D) annotated in 35 pathways, including seven environmental information processing and one environmental adaptation. Therefore, the novel candidate genes found in the current study do not only support new theories about high‐altitude adaptation, but also further explain the molecular mechanisms of altitude adaptation threshold in yaks.     

Follicular dynamics and changes in oestradiol‐17β, progesterone and LH profiles following PGF2α induced oestrus in early and late ovulating Beetal goats

This study compares the factors associated with variable interval to oestrus and ovulation between early versus late ovulating goats following PGF_2α administration. The time of ovulation in Beetal goats (n = 38) was monitored through transrectal ultrasound at every 6 hr following a single dose of PGF_2α (experiment 1). Variations in oestrus and ovulation times were further explored through the changes in follicular dynamics, endocrine profiles and behaviour in another set of goats (n = 13) following single PGF_2α given randomly during the luteal phase (experiment 2). The ovulation time varied between 60 and 96 hr, and 57% of ovulations occurred by 72 hr following PGF_2α (experiment 1). Accordingly, the goats (n = 13) in the second experiment were retrospectively divided either into early and/or late ovulating, that is, ≤72 and/or ≥84 hr following PGF_2α. The onset of oestrus, peak estradiol‐17β concentration and LH surge after PGF_2α was first observed in early than late ovulating goats (p < 0.05). The goats ovulating early had larger follicle and smaller CL in diameter at the time of PGF_2α administration than those ovulating late (5.4 ± 0.2 vs. 4.3 ± 0.2 mm and 10 ± 0.6 vs. 11.8 ± 0.3 mm, respectively; p < 0.05). Likewise, plasma progesterone concentration tended to be lower (p = 0.087) in early than late ovulating goats. In conclusion, the size of dominant follicle and CL at the time of PGF_2a determines the interval to ovulation following a single dose of PGF_2a during the luteal phase.     

Diet is a main source of vitamin D in Finnish pet rabbits (Oryctolagus cuniculus)

During the winter time in Finland, sunlight is inadequate for vitamin D synthesis. Many pet rabbits live as house rabbits with limited outdoor access even during summer and may therefore be dependent on dietary sources of vitamin D. The aims of this study were to report the serum 25‐hydroxyvitamin D concentrations in Finnish pet rabbits and to identify factors that influence vitamin D status. Serum 25‐hydroxyvitamin D concentrations from 140 pet rabbits were determined using a vitamin D enzyme immunoassay (EIA) kit. Eleven rabbits were excluded from the statistical analysis because of unclear dietary data. The remaining 129 rabbits were divided into groups depending on outdoor access during summer (no access n = 26, periodic n = 57, regular n = 46) as well as daily diet: little or no hay and commercial rabbit food ≤1/2 dl (n = 12); a lot of hay and no commercial food daily (n = 23); a lot of hay and commercial food <1 dl (n = 59); a lot of hay and commercial food ≥1 dl (n = 35). The range of serum 25‐hydroxyvitamin D concentration was from 4.5 to 67.5 ng/ml with a mean of 26.1 ng/ml. Statistical general linear model adjusted for weight, age and season indicated that diet was associated with vitamin D concentrations (p = 0.001), but outdoor access during summer was not (p = 0.41). Mean 25‐hydroxyvitamin D concentration was significantly higher in the rabbits receiving a lot of hay and commercial food ≥1 dl (33.9 ± 13.2 ng/ml) than in rabbits in other diet groups (24.0 ± 8.5 ng/ml, 21.7 ± 8.1 ng/ml, and 22.2 ± 18.0 ng/ml, respectively). This investigation showed wide variation in 25‐hydroxyvitamin D concentrations among Finnish pet rabbits. Diet remains a main source since outdoor access seems to be too limited to provide adequate vitamin D synthesis for most of them, and the use of vitamin D supplements is rare.     

Effects of methionine partially replaced by methionyl‐methionine dipeptide on intestinal function in methionine‐deficient pregnant mice

This study was to compare the effects of parenteral supplementation of methionyl‐methionine (Met‐Met) or Met on intestinal barrier function in Met‐deficient pregnant mice. Pregnant mice were randomly divided into three groups. The Control group was provided a diet containing Met and received i.p. injection of saline. The Met group was fed the same diet but without Met and received daily i.p. injection of 35% of the Met contained in the control diet. The Met‐Met group was treated the same as the Met group, except that 25% of the Met injected was replaced with Met‐Met. Met‐Met promoted villus surface area in ileum compared with Met alone. In addition, the mRNA abundance of amino acid and glucose transporters in the small intestine was altered with Met‐Met. Moreover, Met‐Met increased tight junction protein and decreased apoptosis‐related proteins expression in the jejunum and ileum. These results suggest that Met‐Met can promote intestinal function over Met alone in Met‐deficient mice.     

Effect of genistein on IGF‐1 and IGFBP‐1 in young and aged female rat ovary

The objective of this study was to assess the effects of genistein (GEN) on expression of insulin‐like growth factor 1 (IGF‐1) and insulin‐like growth factor binding protein 1 (IGFBP‐1) in young and aged rat ovary. Forty young female Sprague Dawley (SD) rats (200 ± 20 g) and forty aged female SD rats (490 ± 20 g) were selected and according to weight, they were divided into the following five groups with eight animals in each: negative control group (NC), low‐dose group (L), middle‐dose group (M), high‐dose group (H) and positive control group (PC). GEN group received GEN of 15, 30, 60 mg/kg respectively. It lasted 30 days. Concentrations of serum hormones, IGF‐1 and IGFBP‐1 were determined by enzyme‐linked immunosorbent assay (ELISA). Gene and protein expressions of IGF‐1 and IGFBP‐1 were determined by real‐time PCR and Western blot respectively. Compared with NC, GEN significantly increased oestradiol‐17β(E₂) level in aged rat, reduced luteinizing hormone (LH) level in young and aged rat. Serum levels of IGFBP‐1 in young rats were significantly higher in GEN groups (p < 0.05). mRNA and protein expression levels of IGF‐1 and IGFBP‐1 were positively correlated with GEN dose. GEN could significantly reduce the ratio of IGF‐1/IGFBP‐1 of aged rats. Multivariate Cox regression analysis result showed IGF‐1 and IGFBP‐1 levels significantly correlated with GEN dose. We speculate that there is an association between the addition of GEN and expression of IGF‐1 and IGFBP‐1, and the relationship between them is different in young and aged rat.     

Influence of storage time and processing on chemical composition and in vitro ruminal fermentation of olive cake

Olive oil extraction generates olive cake (OC) that could be used in ruminant feeding. However, the chemical composition of OC is affected by multiple factors, being therefore highly variable. The objective of this study was to analyse the influence of storage time and further processing: crude, exhausted (subjected to a second oil extraction) and cyclone (obtained from a cyclone separator) on nutritive value of OC samples. Twelve samples (six crude and six exhausted) were obtained monthly from the same pond from 1 to 6 storage months, and nine samples (three crude, three exhausted and three cyclone) were obtained monthly from a different pond from 6 to 9 months storage. Chemical composition was analysed, and OC samples were fermented in vitro with sheep rumen fluid. Increasing storage time up to 6 months decreased sugars and total soluble polyphenols content but increased fibre content in OC. Dry matter effective degradability (DMED) decreased linearly (p < 0.001) by 35.9 and 45.5% as storage time augmented from 1 to 6 months for crude and exhausted OC, respectively. Crude OC had lower DMED values than exhausted OC (averaged values 0.255 and 0.294 g/g, respectively). Both potential production and rate of gas production were lower (p ≤ 0.018) in crude compared with exhausted OC, which was attributed to the high fat content of crude OC (≥86 g/kg dry matter). For samples stored longer than 6 months, cyclone had greater (p < 0.05) DMED than crude and exhausted OC (averaged values 0.207, 0.164 and 0.164 g/g, respectively). The results indicate that ruminal degradability of OC is reduced with advancing storage time, but only subtle changes were observed during the first two months. Cyclone showed greater degradability than crude and exhausted OC, but differences between crude and exhausted OC became negligible after five storage months.     

Heart rate variability parameters as markers of the adaptation to a sealed environment (a hypoxic normobaric chamber) in the horse

Simulated hypoxic normobaric devices have been used in human beings in order to enhance endurance capacity. These devices are sealed chambers where the athletes are supposed to stay for at least 6–8 hr daily. The current research assesses the changes in time‐domain, spectral and non‐geometrical heart rate variability (HRV) parameters in 6 horses subjected to progressive duration periods inside of a hermetically sealed chamber. It was pursued, firstly to evaluate the intensity of the stress experienced by the animals and secondly to elucidate whether the horses might require an acclimation period before implementation of hypoxic conditions. HRV parameters were monitored for 6 days: day 0 (6‐hr duration; in paddocks; basal conditions), and days 1, 2, 3, 4 and 5 (1, 2, 3, 4 and 6 hr inside the chamber every day respectively). During day 1 and during the first hours of days 2 and 3, compared to day 0, horses presented increased HR and SDHR values and decreased RR interval duration. SD1 values decreased on some hours of days 2 and 3, but differences with day 0 were not found on day 1. Increased SDNN, RMSSD, SD1 and SD2 values were observed on days 4 and 5. These results showed an activation of the sympathetic activity together with an attenuation of the parasympathetic activity during the days 1 to 3. Increased parasympathetic activity was found only during the first hours of days 4 and 5. Spectral parameters experienced minor variations, with increased LF_peak and LF% during some hours of days 4 and 5. In conclusion, at least 3 days are needed to adapt the horse to a sealed environment before starting to subject the animals to hypoxic conditions. When the horses were acclimatized, however, a minor stress was detected with they spent more than 4 hr inside of the chamber.