Mechanism of Iron Ion Homeostasis in Intestinal Immunity and Gut Microbiota Remodeling.

Iron is a vital trace element that plays an important role in humans and other organisms. It plays an active role in the growth, development, and reproduction of bacteria, such as Bifidobacteria. Iron deficiency or excess can negatively affect bacterial hosts. Studies have reported a major role of iron in the human intestine, which is necessary for maintaining body homeostasis and intestinal barrier function. Organisms can maintain their normal activities and regulate some cancer cells in the body by regulating iron excretion and iron-dependent ferroptosis. In addition, iron can modify the interaction between hosts and microorganisms by altering their growth and virulence or by affecting the immune system of the host. Lactic acid bacteria such as Lactobacillus acidophilus (L. acidophilus), Lactobacillus rhamnosus (L. rhamnosus), and Lactobacillus casei (L. casei) were reported to increase trace elements, protect the host intestinal barrier, mitigate intestinal inflammation, and regulate immune function. This review article focuses on the two aspects of the iron and gut and generally summarizes the mechanistic role of iron ions in intestinal immunity and the remodeling of gut microbiota.

Remodeling the hepatic fibrotic microenvironment with emerging nanotherapeutics: a comprehensive review.

Liver fibrosis could be the last hope for treating liver cancer and remodeling of the hepatic microenvironment has emerged as a strategy to promote the ablation of liver fibrosis. In recent years, especially with the rapid development of nanomedicine, hepatic microenvironment therapy has been widely researched in studies concerning liver cancer and fibrosis. In this comprehensive review, we summarized recent advances in nano therapy-based remodeling of the hepatic microenvironment. Firstly, we discussed novel strategies for regulatory immune suppression caused by capillarization of liver sinusoidal endothelial cells (LSECs) and macrophage polarization. Furthermore, metabolic reprogramming and extracellular matrix (ECM) deposition are caused by the activation of hepatic stellate cells (HSCs). In addition, recent advances in ROS, hypoxia, and impaired vascular remodeling in the hepatic fibrotic microenvironment due to ECM deposition have also been summarized. Finally, emerging nanotherapeutic approaches based on correlated signals were discussed in this review. We have proposed novel strategies such as engineered nanotherapeutics targeting antigen-presenting cells (APCs) or direct targeting T cells in liver fibrotic immunotherapy to be used in preventing liver fibrosis. In summary, this comprehensive review illustrated the opportunities in drug targeting and nanomedicine, and the current challenges to be addressed.

Potential therapeutic effects of milk-derived exosomes on intestinal diseases.

Exosomes are extracellular vesicles with the diameter of 30 ~ 150 nm, and are widely involved in intercellular communication, disease diagnosis and drug delivery carriers for targeted disease therapy. Therapeutic application of exosomes as drug carriers is limited due to the lack of sources and methods for obtaining adequate exosomes. Milk contains abundant exosomes, several studies have shown that milk-derived exosomes play crucial roles in preventing and treating intestinal diseases. In this review, we summarized the biogenesis, secretion and structure, current novel methods used for the extraction and identification of exosomes, as well as discussed the role of milk-derived exosomes in treating intestinal diseases, such as inflammatory bowel disease, necrotizing enterocolitis, colorectal cancer, and intestinal ischemia and reperfusion injury by regulating intestinal immune homeostasis, restoring gut microbiota composition and improving intestinal structure and integrity, alleviating conditions such as oxidative stress, cell apoptosis and inflammation, and reducing mitochondrial reactive oxygen species (ROS) and lysosome accumulation in both humans and animals. In addition, we discussed future prospects for the standardization of milk exosome production platform to obtain higher concentration and purity, and complete exosomes derived from milk. Several in vivo clinical studies are needed to establish milk-derived exosomes as an effective and efficient drug delivery system, and promote its application in the treatment of various diseases in both humans and animals.

Circadian miR-449c-5p regulates uterine Ca2+ transport during eggshell calcification in chickens.

BACKGROUND: miRNAs regulate circadian patterns by modulating the biological clocks of animals. In our previous study, we found that the clock gene exhibited a cosine expression pattern in the fallopian tube of chicken uterus. Clock-controlled miRNAs are present in mammals and Drosophila; however, whether there are clock-controlled miRNAs in the chicken uterus and, if so, how they regulate egg-laying rhythms is unclear. In this study, we selected 18 layer hens with similar ovipositional rhythmicity (each of three birds were sacrificed for study per 4 h throughout 24 h); their transcriptomes were scanned to identify the circadian miRNAs and to explore regulatory mechanisms within the uterus of chickens. RESULTS: We identified six circadian miRNAs that are mainly associated with several biological processes including ion trans-membrane transportation, response to calcium ion, and enrichment of calcium signaling pathways. Verification of the experimental results revealed that miR-449c-5p exhibited a cosine expression pattern in the chicken uterus. Ca2+-transporting ATPase 4 (ATP2B4) in the plasma membrane is the predicted target gene of circadian miR-449c-5p and is highly enriched in the calcium signaling pathway. We speculated that clock-controlled miR-449c-5p regulated Ca2+ transportation during eggshell calcification in the chicken uterus by targeting ATP2B4. ATP2B4 mRNA and protein were rhythmically expressed in the chicken uterus, and dual-luciferase reporter gene assays confirmed that ATP2B4 was directly targeted by miR-449c-5p. The expression of miR-449c-5p showed an opposite trend to that of ATP2B4 within a 24 h cycle in the chicken uterus; it inhibited mRNA and protein expression of ATP2B4 in the uterine tubular gland cells. In addition, overexpression of ATP2B4 significantly decreased intracellular Ca2+ concentration (P < 0.05), while knockdown of ATP2B4 accelerated intracellular Ca2+ concentrations. We found similar results after ATP2B4 knockdown by miR-449c-5p. Taken together, these results indicate that ATP2B4 promotes uterine Ca2+ trans-epithelial transport. CONCLUSIONS: Clock-controlled miR-449c-5p regulates Ca2+ transport in the chicken uterus by targeting ATP2B4 during eggshell calcification.

A functional polymorphism of inhibin alpha subunit at miR-181b-1-3p-binding site regulates proliferation and apoptosis of chicken ovarian granular cells.

INHA, the gene encoding the inhibin alpha subunit, was involved in folliculogenesis in mammals, but no study was reported for its working pathway in birds. Here we hypothesize that gene polymorphism in INHA 3'UTR might influence miRNAs binding efficiency and further affect the function of this gene. Thus, we investigated the association between the 3'UTR single-nucleotide polymorphisms (SNPs) in INHA and the laying performance in chickens and further explore their possible molecular cascades in granulosa cells (GC). Five SNPs were detected in Tianfu green-shell layers and g. 22,178,975 G > A was significantly associated with total egg numbers at the age of 300 days (EN, n = 286). Birds carrying the AA genotype laid more EN than those with GG (P < 0.05). The allele transition from G to A in the 3'UTR of INHA gene destroyed a binding site which was targeted by miR-181b-1-3p. The expression abundances of INHA mRNA increased firstly and then decreased with follicle growing, and reached the top in the sixth largest pre-ovulation follicle, whereas miR-181b-1-3p levels in chicken pre-hierarchical follicles had the contrary tendency. Further studies indicated that high levels of miR-181b-1-3p increased apoptosis and reduced GC proliferation while miR-181b-1-3p inhibitors decreased apoptosis and promoted GC proliferation. Additionally, depression of INHA increased apoptosis and reduced GC proliferation via a caspase-3-dependent mitochondrial pathway. Generally, the mutation in INHA 3'UTR was tightly correlated with egg production in chickens, and blocked a binding site of miR-181b-1-3p. miR-181b-1-3p inhibited GC proliferation and promoted apoptosis by targeting INHA.

Inhibin A regulates follicular development via hormone secretion and granulosa cell behaviors in laying hens.

Inhibin A regulates follicular development, and its expression level is related to physiological activities, such as the recruitment, selection, and predominance during follicular development. Therefore, examining inhibin A and its regulatory effects on the reproductive performance of poultry is crucial. In this study, we measured the mRNA and protein abundances of INHA and INHBA in the chicken reproductive system and determined the hormone secretion and apoptosis of follicular granulosa cells (GCs) after being treated with inhibin A protein, and flow cytometry was performed to analyze GC apoptosis in INHA-specific small RNA interference (siRNA). We detected that INHA and INHBA were mainly expressed in chicken follicles. The highest INHA mRNA abundance was found in the fifth largest preovulatory follicle (F5) (P < 0.05). INHBA mRNA expression in the largest preovulatory follicle (F1) was significantly higher than those in other follicles (P < 0.05). Similar results were found for INHA and INHBA protein expression in those follicles (P < 0.05). Treatment with inhibin A protein increased the activity of GCs in a dose-dependent manner (P < 0.05), which was characterized by decreased gene expression of pro-apoptotic factors Bax and Caspase-3 (P < 0.05) and increased expression of proliferation genes Bcl-2 and PCNA (P < 0.05). Additionally, inhibin A significantly increased the secretion of progesterone and estradiol (P < 0.05). RNAi-mediated knockdown of INHA increased apoptosis in GCs via a Caspase-3-dependent mitochondrial pathway.

Whole transcriptome analysis revealed the regulatory network and related pathways of non-coding RNA regulating ovarian atrophy in broody hens.

Poultry broodiness can cause ovarian atresia, which has a detrimental impact on egg production. Non-coding RNAs (ncRNAs) have become one of the most talked-about topics in life sciences because of the increasing evidence of their novel biological roles in regulatory systems. However, the molecular mechanisms of ncRNAs functions and processes in chicken ovarian development remain largely unknown. Whole-transcriptome RNA sequencing of the ovaries of broodiness and laying chickens was thus performed to identify the ncRNA regulatory mechanisms associated with ovarian atresia in chickens. Subsequent analysis revealed that the ovaries of laying chickens and those with broodiness had 40 differentially expressed MicroRNA (miRNAs) (15 up-regulated and 25 down-regulated), 379 differentially expressed Long Noncoding RNA (lncRNAs) (213 up-regulated and 166 down-regulated), and 129 differentially expressed circular RNA (circRNAs) (63 up-regulated and 66 down-regulated). The competing endogenous RNAs (ceRNA) network analysis further revealed the involvement of ECM-receptor interaction, AGE-RAGE signaling pathway, focal adhesion, cytokine-cytokine receptor interaction, inflammatory mediator regulation of TRP channels, renin secretion, gap junction, insulin secretion, serotonergic synapse, and IL-17 signaling pathways in broodiness. Upon further analysis, it became evident that THBS1 and MYLK are significant candidate genes implicated in the regulation of broodiness. The expression of these genes is linked to miR-155-x, miR-211-z, miR-1682-z, gga-miR-155, and gga-miR-1682, as well as to the competitive binding of novel_circ_014674 and MSTRG.3306.4. The findings of this study reveal the existence of a regulatory link between non-coding RNAs and their competing mRNAs, which provide a better comprehension of the ncRNA function and processes in chicken ovarian development.

Integrative metabolomics and transcriptomics analysis revealed specific genes and metabolites affecting meat quality of chickens under different rearing systems.

Different rearing systems have varying effect on animal welfare and meat quality of poultry. Currently, there are no established standards for the rearing systems of Chinese indigenous chickens. Our study aimed to investigate the effects of different rearing systems on the meat quality, gene profiles, and metabolites of Chinese indigenous chickens (Nanchuan chicken). 10-wk-old Nanchuan chickens (n=360) were randomly divided into 3 groups (cage, net, and free-range groups), with 6 replicates per group (20 chickens per replicate). The experiment lasted for 12 wk. At 154-days-old, 36 healthy chickens (6 males and 6 females per group) were randomly selected, euthanized, and their breast muscles were collected to assess the meat quality parameters and histomorphological characteristics. Additionally, breast muscles from 18 random hens (3 males and 3 females per group) were used for metabolomics and RNA-seq analysis. The results showed that rearing systems significantly affected the meat quality and myofiber characteristics. The meat quality of breast muscles from free-range chickens was superior to that of caged chickens, characterized by more tender meat and smaller myofiber cross-sectional areas. Integrative metabolomics and transcriptomics analysis revealed that the differentially expressed genes of chicken breast muscles were primarily involved in the myofiber differentiation. Mechanically, the improved meat quality of breast muscle in free-range chickens were mainly associated with enhanced skeletal muscle differentiation facilitated by fibromodulin, increased levels of up-regulated Acetyl-L-carnitine and Propionylcarnitine level, and decreased levels of Nonanoic acid and Elaidic acid abundance (Graphical abstract). This provides a comprehensive understanding of the most effective and sustainable breeding, production, and rearing systems for Chinese indigenous chickens. It also contributes to the current knowledge of the molecular mechanisms underlying the effects of rearing systems on growth performance and meat quality of chickens.

Multiomics analyses reveal high yield-related genes in the hypothalamic-pituitary-ovarian/liver axis of chicken.

Egg production, regulated by multiple tissues, is among the most important economic traits in poultry. However, current research only focuses on the hypothalamic-pituitary-ovarian axis, ignoring the most important organ for substance metabolism in the body, the liver. Eggs are rich in lipids, proteins, and other nutrients, which are biosynthesized in the liver. Therefore, here the liver was included in the study of the hypothalamic-pituitary axis. This study used hypothalamus (HH_vs_LH), pituitary (HP_vs_LP), liver (HL_vs_LL), and ovary (HO_vs_LO) tissue samples from high- and low-laying Chengkou mountain chickens (CMC) for epihistological, transcriptome and metabolomic analyses aimed at improving the reproductive performance of CMC. The results showed that the liver of the high-laying group was yellowish, the cell boundary was clear, and the lipid droplets were evenly distributed. The ovaries of the high-laying group had a complete sequence of hierarchical follicles, which were rich in yolk. In contrast, the ovaries of the low-laying group were atrophic, except for a few small yellow follicles, and numerous primordial follicles that remained. The transcriptome sequences yielded 167.11 Gb of clean data, containing 28,715 genes. Furthermore, 285, 822, 787, and 1,183 differentially expressed genes (DEG) were identified in HH_vs_LH, HP_vs_LP, HL_vs_LL and HO_vs_LO and the DEGs significantly enriched 77, 163, 170, 171 pathways, respectively. Metabolome sequencing yielded 21,808 peaks containing 4,006 metabolites. The differential metabolite analysis yielded 343 and 682 significantly different metabolites (SDM) that significantly enriched 136 and 87 pathways in the liver and ovaries, respectively. A combined analysis of the transcriptome and metabolome of the liver and ovaries identified "CYP51A1-4α-carboxy-stigmasta7, 24(24(1))-dien-3ß-ol" and "ACSS1B-estrone 3-sulfate" and other multiple gene-metabolite pairs. The DEGs in the hypothalamus and pituitary mainly enriched signaling transduction. In contrast, the DEGs and SDMs in the liver and ovaries mainly enriched the substance metabolism pathways: "gap junction", "extracellular matrix (ECM)-receptor interaction", "Steroid biosynthesis", and "Steroid hormone biosynthesis". These results suggest that the hypothalamic-pituitary axis may affect egg production mainly by regulating lipid metabolism in the liver and ovaries.

Genome-wide characteristics and potential functions of circular RNAs from the embryo muscle development in Chengkou mountain chicken.

The Chengkou mountain chicken, a native Chinese poultry breed, holds significant importance in the country's poultry sector due to its delectable meat and robust stress tolerance. Muscle growth and development are pivotal characteristics in poultry breeding, with muscle fiber development during the embryonic period crucial for determining inherent muscle growth potential. Extensive evidence indicates that non-coding RNAs (ncRNAs) play a regulatory role in muscle growth and development. Among ncRNAs, circular RNAs (circRNAs), characterized by a closed-loop structure, have been shown to modulate biological processes through the regulation of microRNAs (miRNAs). This study seeks to identify and characterize the spatiotemporal-specific expression of circRNAs during embryonic muscle development in Chengkou mountain chicken, and to construct the potential regulatory network of circRNAs-miRNA-mRNAs. The muscle fibers of HE-stained sections became more distinct, and their boundaries were more defined over time. Subsequent RNA sequencing of 12 samples from four periods generated 9,904 novel circRNAs, including 917 differentially expressed circRNAs. The weighted gene co-expression network analysis (WGCNA)-identified circRNA source genes significantly enriched pathways related to cell fraction, cell growth, and muscle fiber growth regulation. Furthermore, a competitive endogenous RNA (ceRNA) network constructed using combined data of present and previous differentially expressed circRNAs, miRNA, and mRNA revealed that several circRNA transcripts regulate MYH1D, MYH1B, CAPZA1, and PERM1 proteins. These findings provide insight into the potential pathways and mechanisms through which circRNAs regulate embryonic muscle development in poultry, a theoretical support for trait improvement in domestic chickens.

Steatosis in metabolic diseases: A focus on lipolysis and lipophagy.

Fatty acids (FAs), as part of lipids, are involved in cell membrane composition, cellular energy storage, and cell signaling. FAs can also be toxic when their concentrations inside and/or outside the cell exceed physiological levels, which is called "lipotoxicity", and steatosis is a form of lipotoxity. To facilitate the storage of large quantities of FAs in cells, they undergo a process called lipolysis or lipophagy. This review focuses on the effects of lipolytic enzymes including cytoplasmic "neutral" lipolysis, lysosomal "acid" lipolysis, and lipophagy. Moreover, the impact of related lipolytic enzymes on lipid metabolism homeostasis and energy conservation, as well as their role in lipid-related metabolic diseases. In addition, we describe how they affect lipid metabolism homeostasis and energy conservation in lipid-related metabolic diseases with a focus on hepatic steatosis and cancer and the pathogenesis and therapeutic targets of AMPK/SIRTs/FOXOs, PI3K/Akt, PPARs/PGC-1α, MAPK/ERK1/2, TLR4/NF-κB, AMPK/mTOR/TFEB, Wnt/ß-catenin through immune inflammation, oxidative stress and autophagy-related pathways. As well as the current application of lipolytic enzyme inhibitors (especially Monoacylglycerol lipase (MGL) inhibitors) to provide new strategies for future exploration of metabolic programming in metabolic diseases.

High expression of miR-22-3p in chicken hierarchical follicles promotes granulosa cell proliferation, steroidogenesis, and lipid metabolism via PTEN/PI3K/Akt/mTOR signaling pathway.

MicroRNAs (miRNAs) are a class of RNA macromolecules that play regulatory roles in follicle development by inhibiting protein translation through binding to the 3'UTR of its target genes. Granulosa cell (GC) proliferation, steroidogenesis, and lipid metabolism have indispensable effect during folliculogenesis. In this study, we found that miR-22-3p was highly expressed in the hierarchical follicles of the chickens, which indicated that it may be involved in follicle development. The results obtained suggested that miR-22-3p promoted proliferation, hormone secretion (progesterone and estrogen), and the content of lipid droplets (LDs) in the chicken primary GC. The results from the bioinformatics analysis, luciferase reporter assay, qRT-PCR, and Western blotting, confirmed that PTEN was directly targeted to miR-22-3p. Subsequently, it was revealed that PTEN inhibited proliferation, hormone secretion, and the content of LDs in GC. Therefore, this study showed that miR-22-3p could activate PI3K/Akt/mTOR pathway via targeting PTEN. Taken together, the findings from this study indicated that miR-22-3p was highly expressed in the hierarchical follicles of chickens, which promotes GC proliferation, steroidogenesis, and lipid metabolism by repressing PTEN to activate PI3K/AKT/mTOR pathway.

microRNAs-based diagnostic and therapeutic applications in liver fibrosis.

Liver fibrosis is a process of over-extracellular matrix (ECM) aggregation and angiogenesis, which develops into cirrhosis and hepatocellular carcinoma (HCC). With the increasing pressure of liver fibrosis, new therapeutics to cure this disease requires much attention. Exosome-cargoed microRNAs (miRNAs) are emerging approaches in the precision of the liver fibrotic paradigm. In this review, we outlined the different types of hepatic cells derived miRNAs that drive intra-/extra-cellular interactive communication in liver fibrosis with different physiological and pathological processes. Specifically, we highlighted the possible mechanism of liver fibrosis pathogenesis associated with immune response and angiogenesis. In addition, potential clinical biomarkers and different stem cell transplant-derived miRNAs-based therapeutic strategies in liver fibrosis were summarized in this review. miRNAs-based approaches might help researchers devise new candidates for the cell-free treatment of liver fibrosis. This article is categorized under: RNA in Disease and Development > RNA in Disease.

miR-128-3p regulates chicken granulosa cell function via 14-3-3ß/FoxO and PPAR-γ/LPL signaling pathways.

MicroRNAs (miRNAs) are class of 22 nt short RNA sequences which inhibit protein translation through binding to the 3'UTR of its target genes. The continuous ovulatory property of chicken follicle makes it a perfect model for studying granulosa cell (GC) functions. In this study, we found that large number of miRNAs including miR-128-3p, were differentially expressed in the GCs of F1 and F5 follicles of chicken. Subsequently, the results revealed that miR-128-3p inhibited proliferation, the formation of lipid droplets, and hormone secretion in chicken primary GCs through directly targeting YWHAB and PPAR-γ genes. To determine the effects of 14-3-3ß (encoded by YWHAB) protein on GCs functions, we overexpressed or inhibited the expression of YWHAB, and the results showed that YWHAB inhibited the function of FoxO proteins. Collectively, we found that miR-128-3p was highly expressed in the chicken F1 follicles compared to the F5 follicles. In addition, the results indicated that miR-128-3p promoted GC apoptosis through 14-3-3ß/FoxO pathway via repressing YWHAB, and inhibited lipid synthesis by impeding the PPAR-γ/LPL pathway, as well as reduced the secretion of progesterone and estrogen. Taken together, the results showed that miR-128-3p plays a regulatory role in chicken granulosa cell function via 14-3-3ß/FoxO and PPAR-γ/LPL signaling pathways.

Therapeutic application of quercetin in aging-related diseases: SIRT1 as a potential mechanism.

Quercetin, a naturally non-toxic flavonoid within the safe dose range with antioxidant, anti-apoptotic and anti-inflammatory properties, plays an important role in the treatment of aging-related diseases. Sirtuin 1 (SIRT1), a member of NAD+-dependent deacetylase enzyme family, is extensively explored as a potential therapeutic target for attenuating aging-induced disorders. SIRT1 possess beneficial effects against aging-related diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), Depression, Osteoporosis, Myocardial ischemia (M/I) and reperfusion (MI/R), Atherosclerosis (AS), and Diabetes. Previous studies have reported that aging increases tissue susceptibility, whereas, SIRT1 regulates cellular senescence and multiple aging-related cellular processes, including SIRT1/Keap1/Nrf2/HO-1 and SIRTI/PI3K/Akt/GSK-3ß mediated oxidative stress, SIRT1/NF-κB and SIRT1/NLRP3 regulated inflammatory response, SIRT1/PGC1α/eIF2α/ATF4/CHOP and SIRT1/PKD1/CREB controlled phosphorylation, SIRT1-PINK1-Parkin mediated mitochondrial damage, SIRT1/FoxO mediated autophagy, and SIRT1/FoxG1/CREB/BDNF/Trkß-catenin mediated neuroprotective effects. In this review, we summarized the role of SIRT1 in the improvement of the attenuation effect of quercetin on aging-related diseases and the relationship between relevant signaling pathways regulated by SIRT1. Moreover, the functional regulation of quercetin in aging-related markers such as oxidative stress, inflammatory response, mitochondrial function, autophagy and apoptosis through SIRT1 was discussed. Finally, the prospects of an extracellular vesicles (EVs) as quercetin loading and delivery, and SIRT1-mediated EVs as signal carriers for treating aging-related diseases, as well as discussed the ferroptosis alleviation effects of quercetin to protect against aging-related disease via activating SIRT1. Generally, SIRT1 may serve as a promising therapeutic target in the treatment of aging-related diseases via inhibiting oxidative stress, reducing inflammatory responses, and restoring mitochondrial dysfunction.

Altitude-adaption of gut microbiota in Tibetan chicken.

Low oxygen levels and extremely cold weather in high-altitude environments requires more energy intake to maintain body temperature in animals. However, little is known about the characteristics of cecal and ileac microbiota in Tibetan chicken and how the high and low altitude environments affect the gut microbiota communities in Tibetan chicken. In the present study, In the present study, Tibetan chickens (Group HA, 3572 m, 578.5 Pa) and their introduced flatland counterparts (Group LA, 580 m, 894.6 Pa) in the cecum and ileum to identify the possible bacterial species that are helpful for their host in environmental adaption. High-throughput sequencing was used to sequence the V3 to V4 hypervariable regions of the bacterial 16S rRNA gene. By comparing the gut microbial diversity of HA chicken with that of LA, the results indicated that the microbial diversity of the cecum and ileum in group HA was significantly lower (P < 0.05) than those in group LA. The cecum microbiome maintained higher population diversity and richness than the ileum (P < 0.05). Four phyla Firmicutes, Bacterioidetes, Actinobacteria, and Proteobacteria were dominant in two groups. Interestingly, there were significant differences in abundance ratio among the four groups (P < 0.05). The predominant bacteria in HA and LA ileum belong to Proteobacteria and Firmicutes, whereas in cecum, Bacterioidetes and Actinobacteria were predominant in both groups (P < 0.05). Correlation analysis showed that Sporosarcina, Enterococcus, and Lactococcus were strongly related to air pressure, and Peptoclostridium and Ruminococcaceae_UCG-014 are related to altitude and gut microbiota of LA group was influenced by altitude, while HA group affected by air pressure. Meanwhile, the Ruminococcus-torques-group was negatively correlated with the relative abundance of Paenibacillus, and positive correlated with those of other microorganisms. Furthermore, HA has higher abundance of microbiota involved in energy and glycan biosynthesis metabolism pathway, while LA has higher abundance of microbiota involved in membrane transport, signal transduction, and xenobiotics biodegradation and metabolism. Generally, our results suggested that the composition and diversity of gut microbes changed after Tibetan chickens were introduced to the plain. Tibetan chicken may adapt to new environment via reshaping the gut microbiota. Gut microbes may contribute to the host adaption to high altitude environments by increasing host energy and glycan biosynthesis.

Integrated Proteomic and Metabolomic Analyses of Chicken Ovary Revealed the Crucial Role of Lipoprotein Lipase on Lipid Metabolism and Steroidogenesis During Sexual Maturity.

During sexual maturation and ovulatory cycle in chickens, ovaries undergo dynamic morphological and functional changes. The aim of this study was to evaluate the integrated proteome and metabolome analyses of chicken ovaries to characterize the changes in protein and metabolite profiles during sexual maturity. The ovary of Rohman layers before (125 days of age) and after (139 days of age) sexual maturation were collected for proteome and metabolome sequencing. The results showed that a total of 680 differentially expressed proteins (DEPs) and 1,046 differential metabolites (DMs) were identified in the chicken ovary during sexual maturity. Among the DEPs, 595 proteins were up-regulated and 85 were down-regulated, whereas 519 metabolites were up-regulated and 527 were down-regulated. KEGG pathway enrichment analysis showed that DEPs were significantly enriched in glycerolipid metabolism, calcium signaling pathway, folate biosynthesis, fat digestion and absorption, NF-kB signaling pathway, and PPAR signaling pathway. However, DMs were significantly enriched in the metabolism pathways, PPAR signalling pathway, glycerolipid metabolism, ferroptosis, biosynthesis of amino acids, and biosynthesis of unsaturated fatty acids. The results of the integrated analyses of DEPs and DMs revealed that the PPAR signaling pathway and glycerolipid metabolism were the most significantly enriched pathways. Among the identified DEPs, lipoprotein lipase (LPL) was upregulated in sexually mature chicken ovaries and was significantly enriched in the glycerolipid metabolism pathway, which may partially explain the possible reasons for steroidogenesis and lipid reserves responsible for oocyte maturation and ovarian follicle development during sexual maturity in chickens. The results further revealed that LPL silencing decreased the content of lipid droplets (LDs), as well as the mRNA expression of lipid metabolism-related genes including; sterol regulatory element binding protein-1 (SREBP-1) and fatty acid synthase (FASN); and steroidogenesis-related genes such as; cytochrome P450 11A1 (CYP11A1) and steroidogenic acute regulatory (StAR). The present study revealed that upregulation of LPL in the chicken ovary during sexual maturity promotes granulosa cell (GC) lipid metabolism and steroidogenesis. These findings provide a theoretical support for further studies to elucidate the mechanism of lipid metabolism to regulate the function of avian GCs during sexual maturity in chickens.

miR-122-5p regulates proliferation and apoptosis of chicken granulosa cells of hierarchal follicles by targeting MAPK3.

Chicken follicles plays a crucial role in the reproductive performance, especially in laying period. Recently, miR-122-5p has been found to be differentially expressed in the ovaries of rats with polycystic ovary syndrome and normal rats, indicating the potential role of miR-122-5p in the development of granulosa cells (GCs). In present study, we found that miR-122-5p was highly expressed in chicken atrophic ovaries. Herein, we investigated its function on GC proliferation and apoptosis of chicken in vitro. We found that overexpression of miR-122-5p significantly inhibited proliferation and promoted apoptosis of GCs, whereas the opposite effects were detected in miR-122-5p knockdown GCs. Meanwhile, mitogen-activated protein kinase 3 (MAPK3) was confirmed as a new target gene of miR-122-5p by bioinformatics software prediction and the dual-luciferase reporter assay verification. Furthermore, after knockdown of MAPK3, the function of MAPK3 for GC proliferation and apoptosis was opposite to that of miR-122-5p. Collectively, our results indicated that miR-122-5p impeded chicken GC proliferation and promoted apoptosis through the post-transcriptional downregulation of MAPK3.

Dietary supplementation of salidroside alleviates liver lipid metabolism disorder and inflammatory response to promote hepatocyte regeneration via PI3K/AKT/Gsk3-ß pathway.

Fatty liver hemorrhagic syndrome (FLHS) is a chronic hepatic disease which occurs when there is a disorder in lipid metabolism. FLHS is often observed in caged laying hens and characterized by a decrease in egg production and dramatic increase of mortality. Salidroside (SDS) is an herbal drug which has shown numerous pharmacological activities, such as protecting mitochondrial function, attenuating cell apoptosis and inflammation, and promoting antioxidant defense system. We aimed to determine the therapeutic effects of SDS on FLHS in laying hens and investigate the underlying mechanisms through which SDS operates these functions. We constructed oleic acid (OA)-induced fatty liver model in vitro and high-fat diet-induced FLHS of laying hens in vivo. The results indicated that SDS inhibited OA-induced lipid accumulation in chicken primary hepatocytes, increased hepatocyte activity, elevated the mRNA expression of proliferation related genes PCNA, CDK2, and cyclinD1 and increased the protein levels of PCNA and CDK2 (P < 0.05), as well as decreased the cleavage levels of Caspase-9, Caspase-8, and Caspase-3 and apoptosis in hepatocytes (P < 0.05). Moreover, SDS promoted the phosphorylation levels of PDK1, AKT, and Gsk3-ß, while inhibited the PI3K inhibitor (P < 0.05). Additionally, we found that high-fat diet-induced FLHS hens had heavier body weight, liver weight, and abdominal fat weight, and severe steatosis in histology, compared with the control group (Con). However, hens fed with SDS maintained lighter body weight, liver weight, and abdominal fat weight, as well as normal liver without hepatic steatosis. In addition, high-fat diet-induced FLHS hens had high levels of serum total cholesterol (TC), triglyceride (TG), alanine transaminase (ALT), and aspartate aminotransferase (AST) compared to the Con group, however, in the Model+SDS group, the levels of TC, TG, ALT, and AST decreased significantly, whereas the level of superoxide dismutase (SOD) increased significantly (P < 0.05). We also found that SDS significantly decreased the mRNA expression abundance of PPARγ, SCD, and FAS in the liver, as well as increased levels of PPARα and MTTP, and decreased the mRNA expression of TNF-α, IL-1ß, IL-6, and IL-8 in the Model+SDS group (P < 0.05). In summary, this study showed that 0.3 mg/mL SDS attenuated ROS generation, inhibited lipid accumulation and hepatocyte apoptosis, and promoted hepatocyte proliferation by targeting the PI3K/AKT/Gsk3-ß pathway in OA-induced fatty liver model in vitro, and 20 mg/kg SDS alleviated high-fat-diet-induced hepatic steatosis, oxidative stress, and inflammatory response in laying hens in vivo.