Huadong Yin

Expression profiles of muscle genes in postnatal skeletal muscle in lines of chickens divergently selected for high and low body weight

Long-term genetic selection for BW has generated high weight select (HWS) and low weight select (LWS) lines of chickens. These lines show an approximate 10-fold difference in BW at selection age (d 56). The objective of this study was to profile the expression of master regulators of early lineage specification (Pax3, Pax7) and myogenic regulatory factors (Myf5, MyoD1, MyoG, and Mrf4) on day of hatch and d 7, 28, and 56 in pectoralis major and gastrocnemius muscles. There was a line × age interaction for expression of all 6 genes in both muscles. In pectoralis major muscle, Pax3, MyoD1, and Mrf4 showed greater expression in LWS than HWS at day of hatch, whereas all 6 genes showed greater expression in HWS than LWS at d 28. In gastrocnemius muscle, Pax3, Myf5, MyoD1, and MyoG showed greater expression in LWS than HWS at day of hatch, whereas Pax7, Myf5, MyoD1, and Mrf4 showed greater expression in HWS than LWS at d 28. At day of hatch there was no difference in fiber number in gastrocnemius muscle between HWS and LWS; however, HWS had greater fiber diameter than LWS. These results indicate that in LWS there is enhanced expression of genes that are necessary for proliferation of progenitor muscle cells and muscle cell differentiation at day of hatch compared with HWS, but by d 28 these genes are expressed greater in HWS than LWS. Thus, long-term selection for growth has altered the pattern of muscle gene expression.

Evolution of primate α and θ defensins revealed by analysis of genomes.

Abstract Defensins are endogenous peptides with cysteine-rich antimicrobial ability that contribute to host defence against bacterial, fungal and viral infections. There are three subfamilies of defensins in primates: α, β and θ-defensins. α-defensins are most present in neutrophils and Paneth cells; β-defensins are involved in protecting the skin and the mucous membranes of the respiratory, genitourinary and gastrointestinal tracts; and θ-defensins are physically distinguished as the only known fully-cyclic peptides of animal origin, which are first isolated from rhesus macaques. All three kinds of defensins have six conserved cysteines, three intramolecular disulfide bonds, a net positive charge, and β-sheet regions. α and θ-defensins are closely related, comparative amino acid sequences showed that the difference between them is that θ-defensins have an additional stop codon limits the initial defensin domain peptides to 12 residues. Humans, chimpanzees and gorillas do not produce θ-defensin peptides due to a premature stop codon present in the signal sequence of all θ-defensin pseudogenes. By using comprehensive computational searches, here we report the discovery of complete repertoires of the α and θ-defensin gene family in ten primate species. Consistent with previous studies, our phylogenetic analyses showed all primate θ-defensins evident formed one distinct clusters evolved from α-defensins. β-defensins are ancestors of both α and θ-defensins. Human has two copies of DEFA1 and DEFT1P, and two extra DEFA3 and DEFA10P genes compared with gorilla. As different primates inhabit in quite different ecological niches, the production of species-specific α and θ-defensins and these highly evolved θ-defensins in old world monkeys would presumably allow them to better respond to the specific microbial challenges that they face.

Myogenic regulatory factor (MRF) expression is affected by exercise in postnatal chicken skeletal muscles.

The MyoD1, MyoG, Myf5, and Mrf4 proteins belong to the family of muscle regulatory factors (MRFs) and play important roles in skeletal muscle hyperplasia and hypertrophy. We hypothesized that exercise would affect MRF mRNA and protein abundance in postnatal chicken skeletal muscle driving molecular changes that could ultimately lead to increased muscle fiber diameter. At day (d) 43, twelve hundred chickens with similar body weight were randomly assigned to cage, pen, and free-range groups. The MRF mRNA abundance was measured in the pectoralis major and thigh muscle at d56, d70, and d84, and the protein levels of MRFs were determined from the thigh muscle at d84. The results showed no significant difference in mRNA of the MRFs among the three groups at d56 (P>0.05). At d84, chicken in the pen and free-range group showed higher MyoD1, MyoG, Myf5, and Mrf4 mRNA abundance compared to the caged chickens (P<0.05). Free-range chickens had higher Mrf4 and MyoG expression than those in penned ones (P<0.05). Protein abundances of all four factors were lowest in the caged group, and Mrf4 and MyoG protein quantities were greatest in free-range chickens (P<0.05), but Myf5 and MyoD1 protein abundance did not differ between penned and caged groups. The results suggested that exercise up-regulated MRF expression in the postnatal skeletal muscles, which led to an increase in muscle fiber diameter, and eventually affected the meat quality of the skeletal muscles in adult chickens.

Effect of Monochromatic Light on Expression of Estrogen Receptor (ER) and Progesterone Receptor (PR) in Ovarian Follicles of Chicken

Artificial illumination is widely used in modern poultry houses and different wavelengths of light affect poultry production and behaviour. In this study, we measure mRNA and protein abundance of estrogen receptors (ERs) and progesterone receptors (PRs) in order to investigate the effect of monochromatic light on egg production traits and gonadal hormone function in chicken ovarian follicles. Five hundred and fifty-two 19-wk-old laying hens were exposed to three monochromatic lights: red (RL; 660 nm), green (GL; 560 nm), blue (BL; 480 nm) and control cool white (400–760 nm) light with an LED (light-emitting diode). There were 4 identical light-controlled rooms (n = 138) each containing 3 replicate pens (46 birds per pen). Water was supplied ad libitum and daily rations were determined according to the nutrient suggestions for poultry. Results showed that under BL conditions there was an increase in the total number of eggs at 300 days of age and egg-laying rate during the peak laying period. The BL and GL extended the duration of the peak laying period. Plasma melatonin was lowest in birds reared under BL. Plasma estradiol was elevated in the GL-exposed laying hens, and GL and BL increased progesterone at 28 wk of age. In the granulosa layers of the fifth largest preovulatory follicle (F5), the third largest preovulatory follicle (F3) and the largest preovulatory follicle (F1), ERα mRNA was increased by BL and GL. Treatment with BL increased ERβ mRNA in granulosa layers of F5, F3 and F1, while GL increased ERβ mRNA in F5 and F3. There was a corresponding increase in abundance of the proteins in the granulosa layers of F5, with an increase in PR-B, generated via an alternative splice site, relative to PR-A. Treatment with BL also increased expression of PR mRNA in all of the granulosa layers of follicles, while treatment with GL increased expression of PR mRNA in granulosa layers of SYF(small yellow follicle), F5 and F1. These results indicate that blue and green monochromatic lights promote egg production traits via stimulating gonadal hormone secretion and up-regulating expression of ERs and PRs. Changes in PR-B protein suggest that this form of the progesterone receptor is predominant for progesterone action in the granulosa layers of preovulatory follicles in chickens during light stimulation.

Circadian clock genes are rhythmically expressed in specific segments of the hen oviduct

In animals, core clock genes are expressed in many peripheral tissues throughout the body that contribute to tissue specific temporal regulation including those that comprise the reproductive system. The chicken ovulatory cycle seems to provide an example of a system in which circadian and interval timing mechanisms operate during ovulation-oviposition. However, little is known about the possible role of circadian regulation during egg formation and laying. To this end, we determined the rhythmic expression of several known canonical clock genes and clock controlled genes in the 4 segments of the chicken oviduct (infundibulum, magnum, isthmus, and uterus) taken from the same biological state (laying sequence and oviposition time) using real time RT-PCR. Except for Cry1, the other genes we analyzed were expressed in all 4 segments of the oviduct. Intriguingly, in a daily light-dark cycle, Bmal1, Clock, Per2, Per3, Cry2, and Rev-erbβ have highly significant rhythmic expression in the infundibulum and uterus but not in the magnum and isthmus. These results show that there is spatial specificity in the localization of clock cells in the hen reproductive tract and that peripheral clocks might have a direct role in the infundibulum and uterus where yolk is captured and the eggshell is formed, respectively.

Induction of mitochondria-mediated apoptosis and PI3K/Akt/ mTOR-mediated autophagy by aflatoxin B 2 in hepatocytes of broilers

Aflatoxins have been shown to induce hepatotoxicity in animal models, but the effects of aflatoxin B2 (AFB2) on broiler hepatocytes is unclear. This study aimed to investigate the effects of AFB2 on apoptosis and autophagy to provide an experimental basis for understanding the mechanism of aflatoxin-induced hepatotoxicity. One hundred-twenty Cobb500 broilers were allocated to four groups and exposed to 0 mg/kg, 0.2 mg/kg, 0.4 mg/kg, and 0.8 mg/kg of AFB2 per day for 21 d. AFB2 exerted potent proapoptotic and proautophagic effects on hepatocytes, with increased numbers of apoptotic and autophagic hepatocytes. Poly ADP-ribose polymerase (PARP) was cleaved and caspase-3 was activated in experimental groups, showing that the apoptosis of hepatocytes was triggered by AFB2. Increased levels of the autophagy factors Beclin-1 and LC3-II/LC3-I, as well as down-regulation of p62, a marker of autophagic flux, provided additional evidence for AFB2-triggered autophagy. AFB2 induced mitochondria-mediated apoptosis via the production of reactive oxygen species (ROS) and promotion of the translocation of Bax and cytochrome c (cyt c) between mitochondria and the cytosol, triggering the formation of apoptosomes. AFB2 also inhibited the phosphoinositide 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway by activating PI3K, Akt, and mTOR and inhibiting their phosphorylation, contributing to the proautophagic activity of AFB2. These findings provide new insights into the mechanisms involved in AFB2-induced hepatotoxicity in broilers.

Housing system influences abundance of Pax3 and Pax7 in postnatal chicken skeletal muscles

Paired box (Pax) proteins 3 and 7 are associated with activation of muscle satellite cells and play a major role in hyperplastic and hypertrophic growth in postnatal skeletal muscle fibers. The objective of this study was to evaluate the effect of housing system on abundance of Pax3 and Pax7 in postnatal chicken skeletal muscles. At 42 d, 1,200 chickens with similar BW were randomly assigned to cage, pen, and free-range group. The mRNA abundance was measured in pectoralis major and thigh muscle at d 56, 70, and 84, and the protein expression was quantified at d 84. Increases in mRNA abundance of PAX3 and PAX7 with age were less pronounced in caged system chickens than in pen and free-range chickens from d 56 to 84, and free-range chickens showed a more pronounced increase in gene expression with age compared with penned chickens. At d 84, quantities of PAX3 and PAX7 mRNA and protein were highest in both pectoralis major and thigh muscle of chickens raised in the free-range group, lowest in penned chickens, and intermediate in caged chickens (P < 0.05). These data indicate that housing system may influence muscle fiber muscle accretion by coordinating the expression of Pax3 and Pax7 in adult chicken skeletal muscles.

Characterization of the Expression Profile and Genetic Polymorphism of the Cellular Retinol-Binding Protein (CRBP IV) Gene in Erlang Mountainous Chickens

In this study, we cloned the coding sequence of chicken CRBP IV, quantified the mRNA expression in Erlang Mountainous Chickens, and investigated a polymorphism in this gene and its association with egg production traits among 349 individuals. The cloned fragment contained a 384 bp open reading frame, which encoded a predicted protein of 127 amino acids and was highly conserved among species. Expression of CRBP IV mRNA was detected in all eight tissues (small intestine, heart, liver, kidney, oviduct, ovary, pituitary, and hypothalamus) at different ages (12, 24, 32 and 45 w). High expression was found in small intestine, pituitary, kidney and liver, whereas it was low in the heart (p < 0.05). The CRBP IV mRNA levels changed with age in the various tissues, and were highly expressed in all tissues at 32 w, except for the heart. We identified one nucleotide substitution (c. 826T>C) in the second exon, which caused an amino acid change (p. S49L). Genotypes (TT, TC and CC) had significant effects on the age at first egg (AFE), total eggs for 300 days (TE300) and highest continuous laying days (HCLD). The CC genotype would be genetically advantageous to improve egg production traits due to earlier AFE, more TE300, and longer HCLD.

Ontogenic expression pattern and genetic polymorphisms of the retinol-binding protein 4 (RBP4) gene in Erlang mountainous chickens.

Retinol-binding protein 4 (RBP4) is the only circulatory transport protein for vitamin A. Based on the essential role of vitamin A in chicken reproduction, we measured RBP4 mRNA abundance in Erlang mountainous chickens. We also identified and analyzed the gene polymorphism and its effect on reproduction traits among 349 chickens. The expression of RBP4 mRNA showed specific developmental changes and striking differences among tissues. The mRNA abundance was greatest (P<0.05) in the liver, intermediate in the ovary, kidney, small intestine, oviduct and heart, and lowest in the hypothalamus and pituitary, as compared to all other tissues (P<0.05). We detected one single nucleotide polymorphism (g.19942455C>G) in intron 2 of the RBP4 gene. Three genotypes (CC, CG and GG) were identified, with a significant effect of genotype on the age at first egg (AFE), first egg weight (FEW), total eggs at 300 days (TE300), highest continuous laying days (HCLD) and average laying interval (ALI). The GG genotype, where chickens display earlier AFE, more TE300, longer HCLD and shorter ALI, would be genetically advantageous and its selection may improve reproduction traits. These results suggested that the RBP4 gene might play an important role in reproduction traits in chickens.

Genetic evidence from mitochondrial DNA corroborates the origin of Tibetan chickens

Chicken is the most common poultry species and is important to human societies. Tibetan chicken (Gallus gallus domesticus) is a breed endemic to China that is distributed mainly on the Qinghai-Tibet Plateau. However, its origin has not been well characterized. In the present study, we sequenced partial mitochondrial DNA (mtDNA) control region of 239 and 283 samples from Tibetan and Sichuan indigenous chickens, respectively. Incorporating 1091 published sequences, we constructed the matrilineal genealogy of Tibetan chickens to further document their domestication history. We found that the genetic structure of the mtDNA haplotypes of Tibetan chickens are dominated by seven major haplogroups (A-G). In addition, phylogenetic and network analyses showed that Tibetan chickens are not distinguishable from the indigenous chickens in surrounding areas. Furthermore, some clades of Tibetan chickens may have originated from game fowls. In summary, our results collectively indicated that Tibetan chickens may have diverged from indigenous chickens in the adjacent regions and hybridized with various chickens.