Assessment of isolation strategies to remove caseins for high-quality milk-derived extracellular vesicles.

Increasing studies have highlighted the significance of milk-derived extracellular vesicles (MEVs) in mother-newborn integration, as well as their application as novel drug delivery systems and diagnostic biomarkers. However, conventional ultracentrifugation (UC) often results in the co-precipitation of casein micelles in MEV pellets. In this study, we compared methods with different principles to screen the optimal pretreatment in caseins removal, and found that isoelectric precipitation by hydrochloric acid (HA) could most effectively remove caseins in porcine milk. We further characterized MEV populations isolated by UC and HA/UC from diverse aspects, including particle methodology via nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM), RNA and protein contents, and purity analysis. Importantly, the proliferative and anti-inflammatory effects of MEVs were evaluated in vitro, showing the superiority of MEVs via HA/UC in functionality compared with UC. Our results suggest that HA pretreatment before ultracentrifugation could effectively remove caseins and other protein complexes, leading to MEVs via HA/UC with higher purity and more significant effects in vitro. This study provides valuable insights for the advancement of MEVs isolation techniques across different species and accurate function analysis of MEVs.

Pituitary-derived small extracellular vesicles promote liver repair by its cargo miR-143-3p.

The small Extracellular vesicles (sEV) has been recognized to be significant for intercellular communication due to their ability to transfer important cellular cargoes like miRNAs through circulation. The pituitary gland has not been clearly known about the role of its secreted sEV under normal physiological conditions. And Liver disease is a global public health burden. The present study is the first to investigate the effect of pituitary sEV on the liver. Sequencing and qRT-PCR revealed miR-143-3p is one of the richest in the pituitary sEV. MiR-143 Knockout (KO) mice resulted in a remarkable decrease in insulin-like growth factor 1 (IGF-1) levels and a significant increase in insulin-like growth factor binding protein 5 (IGFBP5) levels along with a reduction in liver primary cell growth. More importantly, compared with miR-143-KO-sEV, WT-sEV possesses a more robust capacity to improve miR-143 KO mice liver repair through the Wnt/ß-catenin pathway after an acute injury caused by carbon tetrachloride (CCl4). Our results indicate that pituitary-derived sEV promotes hepatocyte proliferation and liver repair by its cargo miR-143-3p and provides new insight into the regulation mechanism of the pituitary-liver axis, and open a new window for endocrine regulation by using sEV.

Early growth response factor 3 regulates coronary atherosclerosis through the NF-κB signaling pathway and VEGF expression.

AIM: The present study was conducted to measure the expression of early growth response factor 3 (Egr3), inflammatory cytokines (IL-1ß, IL-6), vascular endothelial growth factor (VEGF) and NF-κB in patients with coronary artery disease (CAD) to investigate the relationships of these molecules and Egr3 gene expression. METHODS: We recruited 132 CAD patients and 63 healthy individuals. The expression levels of Egr3, VEGF, p50 and p65 were measured by reverse transcription quantitative polymerase chain reaction and the levels of Egr3, IL-1ß and IL-6 in patients serum and in human coronary artery endothelial cells (HCAECs) were measured by enzyme-linked immunosorbent assay (ELISAs) in CAD patients. HCAECs were treated with ox-LDL to establish an in vitro atherosclerosis model. An oil red O staining assay was used to assess the lipid droplet formation. A colloidal external lumen formed by Matrigel was used to test the migration of HCAECs. The expression of Egr3, VEGF and NF-κB was determined by Western blotting. RESULTS: The levels of serum Egr3 and IL-6 in the severe stenosis group were greater than those in the mild stenosis group and controls (p < 0.05). The level of serum IL-1ß in the severe stenosis group was greater than that in the control group (p < 0.05). Moreover, Egr3 expression was positively associated with IL-6 levels (r= 0.55, p < 0.001), IL-1ß levels (r=0.21, p=0.004) and the Gensini score (r=0.20, p=0.02). We also found that Egr3 expression was significantly greater in CAD patients than that in controls. And its expression was highest in the mild patients. The expression of VEGF, P50 and P65 was also greater in CAD patients. In the in vitro experiment, we found that the inhibition of Egr3 expression significantly reduced the expression levels of p50, p65, IL-6 and CRP. Moreover, the inhibition of Egr3 expression significantly reduced the lipid droplet formation and decreased capability of lumen formation. CONCLUSIONS: In the pathogenesis of atherosclerosis, Egr3 gene expression may induce the expression of inflammatory factors and lipid droplet formation and lumen formation, which could promote the atherosclerosis development.

Organic Trace Elements Improve the Eggshell Quality via Eggshell Formation Regulation during the Late Phase of the Laying Cycle.

The quality of eggshells is critical to the egg production industry. The addition of trace elements has been shown to be involved in eggshell formation. Organic trace elements have been found to have higher biological availability than inorganic trace elements. However, the effects of organic trace elements additive doses on eggshell quality during the laying period of commercial laying hens required further investigation. This experiment aims to explore the potential mechanisms of different doses of organic trace elements replacing inorganic elements to remodel the eggshell quality of egg-laying hens during the laying period. A total of 360 healthy hens (Lohmann Pink, 45-week-old) were randomly divided into four treatments, with six replications per treatment and 15 birds per replication. The dietary treatments included a basal diet supplemented with inorganic iron, copper, zinc and manganese at commercial levels (CON), a basal diet supplemented with organic iron, copper, zinc and manganese at 20% commercial levels (LOT), a basal diet supplemented with organic iron, copper, zinc and manganese at 30% commercial levels (MOT), and a basal diet supplemented with organic iron, copper, zinc and manganese at 40% commercial levels (HOT). The trial lasted for 8 weeks. The results of the experiment showed that the replacement of organic trace elements did not significantly affect the production performance of laying hens (p > 0.05). Compared with inorganic trace elements, the MOT and HOT groups improved the structure of the eggshells, enhanced the hardness and thickness of the eggshells, increased the Haugh unit of the eggs, reduced the proportion of the mammillary layer in the eggshell, and increased the proportion of the palisade layer (p < 0.05). In addition, the MOT and HOT groups also increased the enzyme activity related to carbonate transport in the blood, the expression of uterine shell gland-related genes (CA2, OC116, and OCX32), and the calcium and phosphorus content in the eggshells (p < 0.05). We also found that the MOT group effectively reduced element discharge in the feces and enhanced the transportation of iron (p < 0.05). In conclusion, dietary supplementation with 30-40% organic micronutrients were able to improve eggshell quality in aged laying hens by modulating the activity of serum carbonate transport-related enzymes and the expression of eggshell deposition-related genes.

Effects of emulsifiers on lipid metabolism and performance of yellow-feathered broilers.

BACKGROUND: Reducing production costs while producing high-quality livestock and poultry products is an ongoing concern in the livestock industry. The addition of oil to livestock and poultry diets can enhance feed palatability and improve growth performance. Emulsifiers can be used as potential feed supplements to improve dietary energy utilization and maintain the efficient productivity of broilers. Therefore, further investigation is warranted to evaluate whether dietary emulsifier supplementation can improve the efficiency of fat utilization in the diet of yellow-feathered broilers. In the present study, the effects of adding emulsifier to the diet on lipid metabolism and the performance of yellow-feathered broilers were tested. A total of 240 yellow-feasted broilers (21-day-old) were randomly divided into 4 groups (6 replicates per group, 10 broilers per replicate, half male and half female within each replicate). The groups were as follows: the control group (fed with basal diet), the group fed with basal diet supplemented with 500 mg/kg emulsifier, the group fed with a reduced oil diet (reduced by 1%) supplemented with 500 mg/kg emulsifier, and the group fed with a reduced oil diet supplemented with 500 mg/kg emulsifier. The trial lasted for 42 days, during which the average daily feed intake, average daily gain, and feed-to-gain ratio were measured. Additionally, the expression levels of lipid metabolism-related genes in the liver, abdominal fat and each intestinal segment were assessed. RESULTS: The results showed that compared with the basal diet group, (1) The average daily gain of the basal diet + 500 mg/kg emulsifier group significantly increased (P < 0.05), and the half-even-chamber rate was significantly increased (P < 0.05); (2) The mRNA expression levels of Cd36, Dgat2, Apob, Fatp4, Fabp2, and Mttp in the small intestine were significantly increased (P < 0.05). (3) Furthermore, liver TG content significantly decreased (P < 0.05), and the mRNA expression level of Fasn in liver was significantly decreased (P < 0.05), while the expression of Apob, Lpl, Cpt-1, and Pparα significantly increased (P < 0.05). (4) The mRNA expression levels of Lpl and Fatp4 in adipose tissue were significantly increased (P < 0.05), while the expression of Atgl was significantly decreased (P < 0.05). (5) Compared with the reduced oil diet group, the half-evading rate and abdominal fat rate of broilers in the reduced oil diet + 500 mg/kg emulsifier group were significantly increased (P < 0.05), and the serum level of LDL-C increased significantly (P < 0.05)0.6) The mRNA expression levels of Cd36, Fatp4, Dgat2, Apob, and Mttp in the small intestine were significantly increased (P < 0.05). 7) The mRNA expression levels of Fasn and Acc were significantly decreased in the liver (P < 0.05), while the mRNA expression levels of Lpin1, Dgat2, Apob, Lpl, Cpt-1, and Pparα were significantly increased (P < 0.05). CONCLUSIONS: These results suggest that dietary emulsifier can enhance the fat utilization efficiency of broilers by increasing the small intestinal fatty acid uptake capacity, inhibiting hepatic fatty acid synthesis and promoting hepatic TG synthesis and transport capacity. This study provides valuable insights for the potential use of emulsifier supplementation to improve the performance of broiler chickens.

Oxidation Behavior of Pre-Strained Polycrystalline Ni3Al-Based Superalloy.

The harsh service environment of aeroengine hot-end components requires superalloys possessing excellent antioxidant properties. This study investigated the effect of pre-strain on the oxidation behavior of polycrystalline Ni3Al-based superalloys. The growth behaviors of oxidation products were analyzed by scanning electron microscope, transmission electron microscope, X-ray Photoelectron Spectroscopy and Raman spectroscopy. The results indicated that the 5% pre-strained alloys exhibited lower mass gain, shallower oxidation depth and more compact oxide film structures compared to the original alloy. This is mainly attributed to the formation of rapid diffusion paths for Al atoms diffusing to the surface under 5% pre-strain, which promotes the faster formation of protective Al2O3 film while continuing to increase the pre-strain to 25% results in less protective transient oxidation behavior being aggravated due to the increase in dislocation density within the alloy, which prevents the timely formation of the protective Al2O3 film, resulting in uneven oxidation behavior on the alloy.

Intelligent cholinergic white matter pathways algorithm based on U-net reflects cognitive impairment in patients with silent cerebrovascular disease.

BACKGROUND AND OBJECTIVE: The injury of the cholinergic white matter pathway underlies cognition decline in patients with silent cerebrovascular disease (SCD) with white matter hyperintensities (WMH) of vascular origin. However, the evaluation of the cholinergic white matter pathway is complex with poor consistency. We established an intelligent algorithm to evaluate WMH in the cholinergic pathway. METHODS: Patients with SCD with WMH of vascular origin were enrolled. The Cholinergic Pathways Hyperintensities Scale (CHIPS) was used to measure cholinergic white matter pathway impairment. The intelligent algorithm used a deep learning model based on convolutional neural networks to achieve WMH segmentation and CHIPS scoring. The diagnostic value of the intelligent algorithm for moderate-to-severe cholinergic pathway injury was calculated. The correlation between the WMH in the cholinergic pathway and cognitive function was analysed. RESULTS: A number of 464 patients with SCD were enrolled in internal training and test set. The algorithm was validated using data from an external cohort comprising 100 patients with SCD. The sensitivity, specificity and area under the curve of the intelligent algorithm to assess moderate and severe cholinergic white matter pathway injury were 91.7%, 87.3%, 0.903 (95% CI 0.861 to 0.952) and 86.5%, 81.3%, 0.868 (95% CI 0.819 to 0.921) for the internal test set and external validation set. for the. The general cognitive function, execution function and attention showed significant differences among the three groups of different CHIPS score (all p<0.05). DISCUSSION: We have established the first intelligent algorithm to evaluate the cholinergic white matter pathway with good accuracy compared with the gold standard. It helps more easily assess the cognitive function in patients with SCD.

The Effects of Optimal Dietary Vitamin D3 on Growth and Carcass Performance, Tibia Traits, Meat Quality, and Intestinal Morphology of Chinese Yellow-Feathered Broiler Chickens.

This study aimed to assess the effects of different dietary vitamin D3 (VD3) levels on growth and carcass performance, tibia traits, meat quality, and intestinal morphology of yellow-feathered broilers. One-day-old broilers (n = 1440) were assigned into four treatment groups with six replicates per group, and each replicate contained 60 chicks. Dietary VD3 significantly improved the growth performance and carcass traits of broilers, and only low-dose VD3 supplementation decreased the abdominal fat percentage. High-dose VD3 supplementation improved intestinal morphology in the finisher stage, while the b* value of breast muscle meat color decreased markedly under VD3 supplementation (p < 0.05). Serum Ca and P levels and the tibia composition correlated positively with dietary VD3 supplementation at the early growth stage. The weight, length, and ash contents of the tibia increased linearly with increasing dietary VD3, with maximum values achieved in the high-dose group at all three stages. Intestinal 16S rRNA sequencing and liver transcriptome analysis showed that dietary VD3 might represent an effective treatment in poultry production by regulating lipid and immune-related metabolism in the gut-liver axis, which promotes the metabolism through the absorption of calcium and phosphorus in the intestine and improves their protective humoral immunity and reduces infection mortality. Dietary VD3 positively affected the growth-immunity and bone development of broilers during the early stage, suggesting strategies to optimize poultry feeding.

Extracellular Vesicles: A Crucial Player in the Intestinal Microenvironment and Beyond.

The intestinal ecological environment plays a crucial role in nutrient absorption and overall well-being. In recent years, research has focused on the effects of extracellular vesicles (EVs) in both physiological and pathological conditions of the intestine. The intestine does not only consume EVs from exogenous foods, but also those from other endogenous tissues and cells, and even from the gut microbiota. The alteration of conditions in the intestine and the intestinal microbiota subsequently gives rise to changes in other organs and systems, including the central nervous system (CNS), namely the microbiome-gut-brain axis, which also exhibits a significant involvement of EVs. This review first gives an overview of the generation and isolation techniques of EVs, and then mainly focuses on elucidating the functions of EVs derived from various origins on the intestine and the intestinal microenvironment, as well as the impacts of an altered intestinal microenvironment on other physiological systems. Lastly, we discuss the role of microbial and cellular EVs in the microbiome-gut-brain axis. This review enhances the understanding of the specific roles of EVs in the gut microenvironment and the central nervous system, thereby promoting more effective treatment strategies for certain associated diseases.

Integrative analysis with machine learning identifies diagnostic and prognostic signatures in neuroblastoma based on differentially DNA methylated enhancers between INSS stage 4 and 4S neuroblastoma.

INTRODUCTION: Accumulating evidence demonstrates that aberrant methylation of enhancers is crucial in gene expression profiles across several cancers. However, the latent effect of differently expressed enhancers between INSS stage 4S and 4 neuroblastoma (NB) remains elusive. METHODS: We utilized the transcriptome and methylation data of stage 4S and 4 NB patients to perform Enhancer Linking by Methylation/Expression Relationships (ELMER) analysis, discovering a differently expressed motif within 67 enhancers between stage 4S and 4 NB. Harnessing the 67 motif genes, we established the INSS stage related signature (ISRS) by amalgamating 12 and 10 distinct machine learning (ML) algorithms across 113 and 101 ML combinations to precisely diagnose stage 4 NB among all NB patients and to predict the prognosis of NB patients. Based on risk scores calculated by prognostic ISRS, patients were categorized into high and low-risk groups according to median risk score. We conducted comprehensive comparisons between two risk groups, in terms of clinical applications, immune microenvironment, somatic mutations, immunotherapy, chemotherapy and single-cell analysis. Ultimately, we empirically validated the differential expressions of two ISRS model genes, CAMTA2 and FOXD1, through immunochemistry staining. RESULTS: Through leave-one-out cross-validation, in both feature selection and model construction, we selected the random forest algorithm to diagnose stage 4 NB, and Enet algorithm to develop prognostic ISRS, due to their highest average C-index across five NB cohorts. After validations, the ISRS demonstrated a stable predictive capability, outperforming the previously published NB signatures and several clinic variables. We stratified NB patients into high and low-risk group based on median risk score, which showed the low-risk group with a superior survival outcome, an abundant immune infiltration, a decreased mutation landscape, and an enhanced sensitivity to immunotherapy. Single-cell analysis between two risk groups reveals biologically cellular variations underlying ISRS. Finally, we verified the significantly higher protein levels of CAMTA2 and FOXD1 in stage 4S NB, as well as their protective prognosis value in NB. CONCLUSION: Based on multi-omics data and ML algorithms, we successfully developed the ISRS to enable accurate diagnosis and prognostic stratification in NB, which shed light on molecular mechanisms of spontaneous regression and clinical utilization of ISRS.

Oral delivery of miR-146a-5p overexpression plasmid-loaded Pickering double emulsion modulates intestinal inflammation and the gut microbe.

The function of miRNAs in intestinal inflammatory injury regulation has been studied extensively. However, the targeted delivery of these functional nucleic acid molecules to specific organs through encapsulation carriers and exerting their functional effects remain critical challenges for further research. Here, we constructed miR-146a-5p overexpression plasmid and validated the anti-inflammatory properties in the cell model. Then, the plasmid was encapsulated by the Pickering double emulsion system to investigate the role of Pickering double emulsion system in LPS-induced acute intestinal inflammatory injury. The results showed that the Pickering double emulsion system could effectively protect the integrity of plasmids in the intestinal tract, alleviate intestinal inflammatory injury, and upregulate the relative abundance of Lactobacillus reuteri. Mechanically, in vivo and in vitro experiments have shown that miR-146a-5p inhibits TLR4/NF-κB pathway to alleviate intestinal inflammation. In addition, miR-146a-5p can also regulate intestinal homeostasis by targeting the RNA polymerase sigma factor RpoD and α-galactosidase A, thereby affecting the growth of Lactobacillus reuteri. Above all, this study reveals a potential mechanism for miR-146a-5p to treat intestinal inflammation and provides a new delivery strategy for miRNAs to regulate intestinal homeostasis.

Integrated meta-omics reveals the regulatory landscape involved in lipid metabolism between pig breeds.

BACKGROUND: Domesticated pigs serve as an ideal animal model for biomedical research and also provide the majority of meat for human consumption in China. Porcine intramuscular fat content associates with human health and diseases and is essential in pork quality. The molecular mechanisms controlling lipid metabolism and intramuscular fat accretion across tissues in pigs, and how these changes in response to pig breeds, remain largely unknown. RESULTS: We surveyed the tissue-resident cell types of the porcine jejunum, colon, liver, and longissimus dorsi muscle between Lantang and Landrace breeds by single-cell RNA sequencing. Combining lipidomics and metagenomics approaches, we also characterized gene signatures and determined key discriminating markers of lipid digestibility, absorption, conversion, and deposition across tissues in two pig breeds. In Landrace, lean-meat swine mainly exhibited breed-specific advantages in lipid absorption and oxidation for energy supply in small and large intestinal epitheliums, nascent high-density lipoprotein synthesis for reverse cholesterol transport in enterocytes and hepatocytes, bile acid formation, and secretion for fat emulsification in hepatocytes, as well as intestinal-microbiota gene expression involved in lipid accumulation product. In Lantang, obese-meat swine showed a higher synthesis capacity of chylomicrons responsible for high serum triacylglycerol levels in small intestinal epitheliums, the predominant characteristics of lipid absorption in muscle tissue, and greater intramuscular adipcytogenesis potentials from muscular fibro-adipogenic progenitor subpopulation. CONCLUSIONS: The findings enhanced our understanding of the cellular biology of lipid metabolism and opened new avenues to improve animal production and human diseases. Video Abstract.

Effects of Spinach Extract and Licorice Extract on Growth Performance, Antioxidant Capacity, and Gut Microbiota in Weaned Piglets.

Anemia and weaning stress are important factors affecting piglet growth performance. Spinach extract and licorice extract have been used to improve anemia and antioxidant capacity, respectively. However, whether they have synergistic effects has not been reported. To evaluate the effects of mixed spinach extract and licorice extract on growth performance, serum biochemistry, antioxidant capacity, and gut microbiota in weaned piglets, a total of 160 weaned piglets were randomly allotted to four treatments with four replications of 10 piglets each. The four treatments were as follows: control (CON) group (basal diet), spinach extract (SE) group (basal diet + 1.5 kg/t spinach extract), licorice extract (LE) group (basal diet + 400 g/t licorice extract), and spinach extract and licorice extract (MIX) group (basal diet + 1.5 kg/t spinach extract + 400 g/t licorice extract). The results showed that, compared with the CON group, diets supplemented with spinach extract and licorice extract significantly increased the average daily gain (p < 0.05), while considerably reducing the feed-to-gain ratio (p < 0.05). Moreover, the MIX group exhibited a significant up-regulation of serum total protein, globulin, albumin, glucose, and triglyceride levels in comparison to the CON group (p < 0.05). Meanwhile, both the anemia and antioxidant capacity of piglets were effectively improved. Notably, the MIX group achieved even better results than the individual supplementation in terms of enhancing growth performance, which could potentially be attributed to the increased abundance of the Rikenellaceae_RC9_gut_group. These results demonstrated that the supplementation of diets with spinach extract and licorice extract improves the absorption of nutrients from the diet and antioxidant capacity in weaned piglets.

Synergism of fermented feed and ginseng polysaccharide on growth performance, intestinal development, and immunity of Xuefeng black-bone chickens.

Microbial fermented feed (MF) is considered a valuable strategy to bring advantages to livestock and is widely practiced. Oral supplementation of Ginseng polysaccharide (Gps) eliminated weight loss in chickens following vaccination. This study investigated the effects of the combined use of Gps and MF on growth performance and immune indices in Xuefeng black-bone chickens. A total of 400 Xuefeng black-bone chickens at the age of 1 day were randomly assigned to four groups. Normal feed group (Control group), ginseng polysaccharide (200 mg/kg) group (Gps group), microbially fermented feed (completely replace the normal feed) group (MF group), and microbially fermented feed and add ginseng polysaccharide just before use (MF + Gps group). Each group contained 5 pens per treatment and 20 birds per pen. The body weight and average daily gain in the Gps, MF, and MF + Gps groups increased significantly (P < 0.01), while the feed conversion ratio decreased significantly (P < 0.01). The combined use of MF and Gps showed a synergistic effect. There was no significant difference in villus height (cecal) between the experimental group and the Con group. The crypt depth of the three experimental groups exhibited a significantly lower value compared to the Control group (P < 0.05). The V/C ratio of the Gps group and MF + Gps was significantly increased (P < 0.05), but there was no significant difference in the MF group. Moreover, the diarrhea rate of the Gps and the MF + Gps groups was lower than that of the Con group, while that of the MF + Gps group decreased the mortality rate (P < 0.05). The serum tumor necrosis factor-alpha (TNF-α) and interleukin 6 (IL-6) levels in the MF, Gps, and MF + Gps groups decreased significantly (P < 0.01), the serum immunoglobulin G (IgG) levels increased significantly (P < 0.01), while the combination of MF and Gps had a synergistic effect. The combined use of Gps and MF not only further improved growth performance and immune parameters, but also reduced the diarrhea rate and mortality.

A novel protein encoded by circKANSL1L regulates skeletal myogenesis via the Akt-FoxO3 signaling axis.

Skeletal muscle is one the largest organs of the body and is involved in animal production and human health. Circular RNAs (circRNAs) have been implicated in skeletal myogenesis through largely unknown mechanisms. Herein, we report the phenotypic and metabolomic analysis of porcine longissimus dorsi muscles in Lantang and Landrace piglets, revealing a high-content of slow-oxidative fibers responsible for high-quality meat product in Lantang piglets. Using single-cell transcriptomics, we identified four myogenesis-related cell types, and the Akt-FoxO3 signaling axis was the most significantly enriched pathway in each subpopulation in the different pig breeds, as well as in fast-twitch glycolytic fibers. Using the multi-dimensional bioinformatic tools of circRNAome-seq and Ribo-seq, we identified a novel circRNA, circKANSL1L, with a protein-coding ability in porcine muscles, whose expression level correlated with myoblast proliferation and differentiation in vitro, as well as the transformation between distinct mature myofibers in vivo. The protein product of circKANSL1L could interact with Akt to decrease the phosphorylation level of FoxO3, which subsequently promoted FoxO3 transcriptional activity to regulate skeletal myogenesis. Our results established the existence of a protein encoded by circKANSL1L and demonstrated its potential functions in myogenesis.

The difference of intestinal microbiota composition between Lantang and Landrace newborn piglets.

BACKGROUND: The early development of intestinal microbiota plays a fundamental role in host health and development. To investigate the difference in the intestinal microbial composition between Lantang and Landrace newborn piglets, we amplified and sequenced the V3-V4 region of 16 S rRNA gene in jejunal microbiota of Lantang and landrace newborn. RESULTS: The findings revealed that the dominant phyla in the jejunum of Lantang piglets were Firmicutes, Actinobacteria and Bacteroidetes, while the dominant phyla of Landrace is Proteobacteria and Fusobacteria. Specifically, Corynebacterium_1, Lactobacillus, Rothia, Granulicatella, Corynebacteriales_unclassified, Corynebacterium, Globicatella and Actinomycetales_unclassified were found to be the dominant genera of Lantang group, while Clostridium_sensu_stricto_1, Escherichia-Shigella, Actinobacillus and Bifidobacterium were the dominant genera of Landrace. Based on the functional prediction of bacteria, we found that bacterial communities from Lantang samples had a significantly greater abundance pathways of fatty acid synthesis, protein synthesis, DNA replication, recombination, repair and material transport across membranes, while the carrier protein of pathogenic bacteria was more abundant in Landrace samples. CONCLUSIONS: Overall, there was a tremendous difference in the early intestinal flora composition between Landang and Landrace piglets, which was related to the breed characteristics and may be one of the reasons affecting the growth characteristics. However, more further extensive studies should be included to reveal the underlying relationship between early intestinal flora composition in different breeds and pig growth characteristics.

Human Mesenchymal Stem Cells-Derived Exosome Mimetic Vesicles Regulation of the MAPK Pathway and ROS Levels Inhibits Glucocorticoid-Induced Apoptosis in Osteoblasts.

Background: Long-term extensive use of glucocorticoids will lead to hormonal necrosis of the femoral head, and osteoblasts play an important role in the prevention of osteonecrosis. However, there is no complete cure for necrosis of the femoral head. Mesenchymal stem cell- (MSCs-) derived exosomes are widely used for the repair of various tissue lesions. Therefore, the aim of this study was to investigate the mechanism of dexamethasone- (DEX-) induced osteoblast apoptosis and the therapeutic effect of human umbilical cord MSC- (hucMSC-) derived exosome mimetic vesicles (EMVs) on osteoblast-induced apoptosis by DEX. Methods: The viability and apoptosis of primary MC3T3-E1 cells were determined by the Cell Counting Kit-8 (CCK-8), FITC-Annexin V/PI staining and immunoblot. The intracellular levels of reactive oxygen species (ROS) after DEX treatment were measured by 2', 7' -dichlorodihydrofluorescein diacetate (DCFH-DA) staining. In this study, hucMSC-EMVs and N-acetyl-l-cysteine (NAC) were used as therapeutic measures. The expression of B-cell lymphoma 2-associated X, Bcl 2, HO-1, and nuclear factor erythroid-derived 2-like 2 and MAPK- signaling pathway in osteogenic cell MC3T3-E1 cells treated with Dex was analyzed by the immunoblotting. Results: DEX significantly induced osteoblasts MC3T3-E1 apoptosis and ROS accumulation. MAPK-signaling pathway was activated in MC3T3-E1 after DEX treatment. hucMSC-EMVs intervention significantly downregulated DEX-induced MAPK-signaling pathway activation and ROS accumulation. In addition, hucMSC-EMVs can reduce the apoptosis levels in osteoblast MC3T3-E1 cells induced by DEX. Conclusions: Our study confirmed that hucMSC-EMVs regulates MAPK-signaling pathway and ROS levels to inhibit DEX-induced osteoblast apoptosis.

The Characteristic Function of Blood-Derived Exosomes and Exosomal circRNAs Isolated from Dairy Cattle during the Dry Period and Mid-Lactation.

Exosomes are key mediators of intercellular communication. They are secreted by most cells and contain a cargo of protein-coding genes, long noncoding RNAs (lncRNAs), and circular RNAs (circRNAs), which modulate recipient cell behavior. Herein, we collected blood samples from Holstein cows at days 30 (mid-lactation) and 250 (dry period) of pregnancy. Prolactin, follicle-stimulating hormone, luteinizing hormone, estrogen, and progesterone levels showed an obvious increase during D250. We then extracted exosomes from bovine blood samples and found that their sizes generally ranged from 100 to 200 nm. Further, Western blotting validated that they contained CD9, CD63, and TSG101, but not calnexin. Blood-derived exosomes significantly promoted the proliferation of mammary epithelial cells, particularly from D250. This change was accompanied by increased expression levels of proliferation marker proteins PCNA, cyclin D, and cyclin E, as detected by EdU assay, cell counting kit-8 assay, and flow cytometric cell cycle analysis. Moreover, we treated mammary epithelial cells with blood-derived exosomes that were isolated from the D30 and D250 periods. And RNA-seq of two groups of cells led to the identification of 839 differentially expressed genes that were significantly enriched in KEGG signaling pathways associated with apoptosis, cell cycle and proliferation. In bovine blood-derived exosomes, we found 12,747 protein-coding genes, 31,181 lncRNAs, 9374 transcripts of uncertain coding potential (TUCP) candidates, and 460 circRNAs, and 32 protein-coding genes, 806 lncRNAs, 515 TUCP candidates, and 45 circRNAs that were differentially expressed between the D30 and D250 groups. We selected six highly expressed and four differentially expressed circRNAs to verify their head-to-tail splicing using PCR and Sanger sequencing. To summarize, our findings improve our understanding of the key roles of blood-derived exosomes and the characterization of exosomal circRNAs in mammary gland development.

MiR-143 Targets SYK to Regulate NEFA Uptake and Contribute to Thermogenesis in Male Mice.

Excessive energy intake is the main cause of obesity, and stimulation of brown adipose tissue (BAT) and white adipose tissue (WAT) thermogenesis has emerged as an attractive tool for antiobesity. Although miR-143 has been reported to be associated with BAT thermogenesis, its role remains unclear. Here, we found that miR-143 had highest expression in adipose tissue, especially in BAT. During short-term cold exposure or CL316,243 was injected, miR-143 was markedly downregulated in BAT and subcutaneous WAT (scWAT). Moreover, knockout (KO) of miR-143 increases the body temperature of mice upon cold exposure, which may be due to the increased thermogenesis of BAT and scWAT. More importantly, supplementation of miR-143 in BAT of KO mice can inhibit the increase in body temperature in KO mice. Mechanistically, spleen tyrosine kinase was revealed for the first time as a new target of miR-143, and deletion of miR-143 facilitates fatty acid uptake in BAT. In addition, we found that brown adipocytes can promote fat mobilization of white adipocytes, and miR-143 may participate in this process. Meanwhile, we demonstrate that inactivation of adenylate cyclase 9 (AC9) in BAT inhibits thermogenesis through AC9-PKA-AMPK-CREB-UCP1 signaling pathway. Overall, our results reveal a novel function of miR-143 on thermogenesis, and a new functional link of the BAT and WAT.