Verification of Key Target Molecules for Intramuscular Fat Deposition and Screening of SNP Sites in Sheep from Small-Tail Han Sheep Breed and Its Cross with Suffolk.

Intramuscular fat (IMF) is vital for meat tenderness and juiciness. This study aims to explore the IMF deposition mechanism and the related molecular markers in sheep. Two populations, Small-tail Han Sheep (STH) and STH × Suffolk (SFK) F1 (SFK × STH), were used as the research object. Histological staining techniques compared the differences in the longissimus dorsi muscle among populations. A combination of transcriptome sequencing and biological information analysis screened and identified IMF-related target genes. Further, sequencing technology was employed to detect SNP loci of target genes to evaluate their potential as genetic markers. Histological staining revealed that the muscle fiber gap in the SFK × STH F1 was larger and the IMF content was higher. Transcriptome analysis revealed that PIK3R1 and PPARA were candidate genes. Histological experiments revealed that the expressions of PIK3R1 mRNA and PPARA mRNA were lower in SFK × STH F1 compared with the STH. Meanwhile, PIK3R1 and PPARA proteins were located in intramuscular adipocytes and co-located with the lipid metabolism marker molecule (FASN). SNP locus analysis revealed a mutation site in exon 7 of the PIK3R1 gene, which served as a potential genetic marker for IMF deposition. This study's findings will provide a new direction for meat quality breeding in sheep.

Dimethyl fumarate restores Ca2+ dyshomeostasis through activation of the SIRT1 signal to treat nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is characterized by an excessive lipid accumulation in the liver, with a global prevalence of approximately 25 %. While early-stage steatosis is reversible and can be intervened upon, it has the potential to progress to some serious complications, including cirrhosis and even liver cancer. Dimethyl fumarate (DMF), a derivative of fumaric acid shows promise in intervening in certain diseases. However, the precise effect and underlying mechanism of DMF on hepatic steatosis remain unclear. In this study, we demonstrated that DMF mitigates hepatic steatosis in mice subjected to high-fat/high-cholesterol (HFHC) diets. Meanwhile, our in vivo and in vitro results showed that DMF relieves lipid accumulation, oxidative stress, and endoplasmic reticulum (ER) stress. Mechanically, our findings revealed that the effect of DMF on reducing lipid accumulation is linked to the restoration of Ca2+ homeostasis. Furthermore, we found that activation of the SIRT1 signal by DMF plays an important role in correcting the mishandling of the Ca2+ signal, and knockdown of SIRT1 expression reverses the beneficial role of DMF PA-incubated AML12 cells. In conclusion, our results suggested DMF's amelioration of hepatic steatosis is related to the activation of SIRT1-mediated Ca2+ signaling.

Alleviative effect of melatonin against the nephrotoxicity induced by cadmium exposure through regulating renal oxidative stress, inflammatory reaction, and fibrosis in a mouse model.

Chronic cadmium (Cd) exposure causes severe adverse health effects on the human body, especially the kidney tissue. Studies have demonstrated oxidative stress to be involved in renal pathological variations after exposure to Cd, but few effective treatments are available for the disease yet. Therefore, the present study was carried out to investigate the potential therapeutic intervention and its underlying molecular mechanisms of melatonin (MT), a natural antioxidant with multiple biological activities, against renal injury caused by Cd exposure in mice. C57BL/6 male mice (eight-week-old) were intragastrically administered with CdCl2, MT, or both for 30 days. Biochemical analysis showed that MT intervention significantly improved the SOD, GSH, and CAT activities while markedly decreasing the kidney MDA content of the mice exposed to Cd. Histological examination indicated that Cd exposure resulted in the atrophy of the renal glomerular, the degeneration and dilation of tubules, and the accumulation of fibrocytes. By contrast, MT administration effectively ameliorated the histological outcome of the injured kidney tissue. Moreover, administrating MT significantly inhibited proinflammatory cytokines TNF-α and iNOS expression in Cd-treated mice. Further, MT treatment markedly suppressed the expressions of renal fibrosis-related factors TGF-ß1, α-SMA, and collagen Ⅰ in the injured renal tissue and the accumulation and development of renal fibrosis. In addition, the administration of MT significantly reduced the expression of caspase-3 and cell apoptotic death in the kidney tissue of Cd-exposed mice. In all, the data showed that MT has a compelling therapeutic potential in alleviating the pathological variations of renal injury caused by Cd exposure.

A High-Precision Quartz Resonant Ultra-High Pressure Sensor with Integrated Pressure Conversion Structure.

A quartz resonant pressure sensor is proposed for high-precision measurement of ultra-high pressure. The resonant unit realizes a push-pull differential layout, which restrains the common-mode interference factor, and the resonator is only subject to axial force. The pressure conversion unit is made in an integrated manner, avoiding output drift problems caused by residual stress and small gaps during assembly, welding, and other processes in sensor preparation. Theoretical and simulation analysis was conducted on the overall design scheme of the sensor in this paper, verifying the feasibility. Sensor prototypes were created and performance experiments were conducted. The experimental results show that the sensitivity of the ultra-high pressure sensor is 46.32 Hz/MPa at room temperature within the pressure range of 120 MPa, and the comprehensive accuracy is 0.0266%. The comprehensive accuracy of the sensor is better than 0.0288% FS in the full temperature range environment. This proves that the sensor scheme is suitable for high-precision and high-stability detection of ultra-high pressure, providing new solutions in special pressure measurement fields such as deep-sea and oil exploration.

Integrated multi-omics reveals the beneficial role of chlorogenic acid in improving the growth performance and immune function of immunologically stressed broilers.

Intensive production can cause immunological stress in commercial broilers. Chlorogenic acid (CGA) regulates the intestinal microbiota, barrier function, and immune function in chickens. As complex interrelations regulate the dynamic interplay between gut microbiota, the host, and diverse health outcomes, the aim of this study was to elucidate the immunoregulatory mechanisms of CGA using multi-omics approaches. A total of 240 one-day-old male broilers were assigned to a 2 × 2 factorial design with 2 CGA levels (0 or 500 mg/kg) either with or without dexamethasone (DEX) injection for a 21-day experimental period. Therefore, there were 4 dietary treatments: control, DEX, CGA, and DEX + CGA, with 6 replicates per treatment. CGA supplementation improved (P < 0.05) growth performance, jejunal morphology, jejunal barrier function, and immune function in DEX-treated broilers. Moreover, in DEX + CGA-treated broilers, the increase in gut microbiome diversity (P < 0.05) was consistent with a change in taxonomic composition, especially in the Clostridiales vadin BB60_group. Additionally, the levels of short-chain fatty acids increased remarkably (P < 0.01) after CGA supplementation. This was consistent with the Kyoto Encyclopedia of Genes and Genomes analysis results that the "pyruvate fermentation to butanoate" pathway was more enriched (P < 0.01) in the DEX + CGA group than in the DEX group. Proteomics revealed that CGA treatment increased the expression of several health-promoting proteins, thymosin beta (TMSB4X) and legumain (LGMN), which were verified by multiple reaction monitoring. Metabolomics revealed that CGA treatment increased the expression of health-promoting metabolites (2,4-dihydroxy benzoic acid and homogentisic acid). Proteomic and metabolic analyses showed that CGA treatment regulated the peroxisome proliferator-activated receptor (PPAR) and mitogen-activated protein kinase (MAPK) pathways. Western blotting results support these findings. Pearson's correlation analyses showed correlations (P < 0.01) between altered immune function, jejunal barrier function, different microbiota, proteins, and metabolites parameters. Overall, our data indicate that CGA treatment increased growth performance and improved the immunological functions of DEX-treated broilers by regulating gut microbiota and the PPAR and MAPK pathways. The results offer novel insights into a CGA-mediated improvement in immune function and intestinal health.

Gas/Liquid Chromatography-Mass Spectrometry Analysis of Key Functional Substances Regulating Poll Gland Secretion in Male Camels during Seasonal Estrus.

Increased poll gland secretion is a major characteristic and indicator of estrus in male Bactrian camels; however, research on these poll glands and their secretion is extremely rare. In this study, we determine the chemical composition of poll gland secretions and identify the key functional substances that regulate seasonal estrus in male camels. A GC/LC-MS dual platform was used to analyze ventral hair (control) and neck mane samples containing poll gland secretions from male Bactrian camels during estrus. Multidimensional and single-dimensional analyses were used to screen differentially expressed metabolites (DEMs) between groups. Functional prediction of enriched metabolites was performed using a Human Metabolome Database comparison and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis, which were then compared with a behavioral analysis of male Bactrian camels in estrus. A total of 1172 DEMs and 34 differential metabolic pathways were identified. One metabolite group was found to relate to steroid synthesis and metabolism, and another metabolite group was associated with neural metabolism. Therefore, we speculate that steroids and neurochemicals jointly regulate estrous behavior in male Bactrian camels, thus providing theoretical insights into the development and function of poll glands in Bactrian camels.

Genome-wide single-nucleotide polymorphism data and mitochondrial hypervariable region 1 nucleotide sequence reveal the origin of the Akhal-Teke horse.

OBJECTIVE: The study investigated the origin of the Akhal-Teke horse using genome-wide single-nucleotide polymorphism (SNP) data and mitochondrial hypervariable region 1 (HVR-1) nucleotide sequences. METHODS: Genome-wide SNP data from 22 breeds (481 horses) and mitochondrial HVR-1 sequences from 24 breeds (544 sequences) worldwide to examine the origin of the Akhal- Teke horse. The data were analyzed using principal component analysis, linkage disequilibrium analysis, neighbor-joining dendrograms, and ancestry inference to determine the population relationships, ancestral source, genetic structure, and relationships with other varieties. RESULTS: A close genetic relationship between the Akhal-Teke horse and horses from the Middle East was found. Analysis of mitochondrial HVR-1 sequences showed that there were no shared haplotypes between the Akhal-Teke and Tarpan horses, and the mitochondrial data indicated that the Akhal-Teke horse has not historically expanded its group. Ancestral inference suggested that Arabian and Caspian horses were the likely ancestors of the Akhal- Teke horse. CONCLUSION: The Akhal-Teke horse originated in the Middle East.

Localization of FGF21 Protein and Lipid Metabolism-Related Genes in Camels.

With the ability to survive under drought and chronic hunger, camels display a unique regulation characteristic of lipid metabolism. Fibroblast growth factor (FGF) 21 is a peptide hormone that regulates metabolic pathways, especially lipid metabolism, which was considered as a promising therapeutic target for metabolic diseases. To understand the FGF21 expression pattern and its potential relationship with lipid metabolism in camels, this study investigated the distribution and expression of FGF21, receptor FGFR1, and two lipid metabolism markers, leptin and hormone-sensitive lipase (HSL), using an immunohistochemistry (IHC) assay. The results showed that FGF21 was widely expressed in camel central nerve tissue and peripheral organs but absent in lung and gametogenic tissue, including the testis, epididymis, and ovary. In striated muscle, FGF21 is only present at the fiber junction. FGFR1 is expressed in almost all tissues and cells, indicating that all tissues are responsive to FGF21 and other FGF-mediated signals. Leptin and HSL are mainly located in metabolic and energy-consuming organs. In the CNS, leptin and HSL showed a similar expression pattern with FGFR1. In addition, leptin expression is extremely high in the bronchial epithelium, which may be due to its role in the immune responses of respiratory mucosa, in addition to fat stores and energy balance. This study found that FGF21 showed active expression in the nervous system of camels, which may be related to the adaptability of camels to arid environments and the specific regulation of lipid metabolism. This study showed a special FGF21-mediated fat conversion pattern in camels and provides a reference for developing a potential therapeutic method for fat metabolism disease.

Highly efficient neutralizer-free l-malic acid production using engineered Saccharomyces cerevisiae.

In industrial bioproduction of organic acids, numerous neutralizers are required which substantially increases production costs and burdens the environment. To address this challenge, a Saccharomyces cerevisiae mutant (named TAMC) with a low pH tolerance (pH 2.3) was isolated by adaptive laboratory evolution. Taking the synthesis of l-malic acid as an example, the malate dehydrogenase 3 without signal peptide (MDHΔSKL) and pyruvate carboxylase 2 (PYC2) were overexpressed in cytoplasmic synthesis pathway, and the l-malic acid titer increased 5.6-fold. Subsequently, the malic acid transporter SpMae1 was designed, and the extracellular l-malic acid titer was increased from 7.3 to 73.6 g/L. Furthermore, by optimizing the synthesis of the precursor pyruvate, the titer reached 81.8 g/L. Finally, without any neutralizer, the titer in the 3-L bioreactor reached 232.9 g/L, the highest l-malic acid titer reported to date. Herein, the engineered l-malic acid overproducer paves the way for the large-scale green production of l-malic acid.

An oligosaccharide marker for rapid authentication of edible bird's nest.

Edible bird's nest (EBN) is a popular and expensive food material. The limited supply and great demand result in the use of adulterants. The authenticity concern is raised due to the lack of appropriate quality markers. Herein, this study aims to provide a specific oligosaccharide marker for rapid EBN authentication. Comparing the benzocaine (ABEE)-labeled saccharide profiles of multiple batches of EBN and adulterants indicates seven unique EBN oligosaccharides. The most abundant one, named BNM001, was selected as a marker and characterized to be Neu5Ac (2-3) Gal by MS and NMR spectra. This new oligosaccharide marker enables a rapid authentication of EBN within 10 min. ABEE labelling of this marker further upgraded the accuracy and sensitivity of the LC-qTOF-MS quantitative analysis. The relative marker content was associated with the quality of EBN products. These results suggest a specific and efficient quality marker for rapid authentication of EBN and related products.

Polygenic prediction of human longevity on the supposition of pervasive pleiotropy.

The highly polygenic nature of human longevity renders cross-trait pleiotropy an indispensable feature of its genetic architecture. Leveraging the genetic correlation between the aging-related traits (ARTs), we sought to model the additive variance in lifespan as a function of cumulative liability from pleiotropic segregating variants. We tracked allele frequency changes as a function of viability across different age bins and prioritized 34 variants with an immediate implication on lipid metabolism, body mass index (BMI), and cognitive performance, among other traits, revealed by PheWAS analysis in the UK Biobank. Given the highly complex and non-linear interactions between the genetic determinants of longevity, we reasoned that a composite polygenic score would approximate a substantial portion of the variance in lifespan and developed the integrated longevity genetic scores (iLGSs) for distinguishing exceptional survival. We showed that coefficients derived from our ensemble model could potentially reveal an interesting pattern of genomic pleiotropy specific to lifespan. We assessed the predictive performance of our model for distinguishing the enrichment of exceptional longevity among long-lived individuals in two replication cohorts and showed that the median lifespan in the highest decile of our composite prognostic index is up to 4.8 years longer. Finally, using the proteomic correlates of iLGS, we identified protein markers associated with exceptional longevity irrespective of chronological age and prioritized drugs with repurposing potentials for gerotherapeutics. Together, our approach demonstrates a promising framework for polygenic modeling of additive liability conferred by ARTs in defining exceptional longevity and assisting the identification of individuals at higher risk of mortality for targeted lifestyle modifications earlier in life. Furthermore, the proteomic signature associated with iLGS highlights the functional pathway upstream of the PI3K-Akt that can be effectively targeted to slow down aging and extend lifespan.

HMOX1 Promotes Ferroptosis in Mammary Epithelial Cells via FTH1 and Is Involved in the Development of Clinical Mastitis in Dairy Cows.

Ferroptosis is associated with inflammatory diseases as a lethal iron-dependent lipid peroxidation; its role in the development of clinical mastitis (CM) in dairy cows is not well understood. The aim of this study was to identify differentially expressed proteins (DEPs) associated with iron homeostasis and apoptosis, and to investigate further their roles in dairy cows with CM. The results suggested that ferroptosis occurs in the mammary glands of Holstein cows with CM. Using data-independent acquisition proteomics, 302 DEPs included in 11 GO terms related to iron homeostasis and apoptosis were identified. In particular, heme oxygenase-1 (HMOX1) was identified and involved in nine pathways. In addition, ferritin heavy chain 1 (FTH1) was identified and involved in the ferroptosis pathway. HMOX1 and FTH1 were located primarily in mammary epithelial cells (MECs), and displayed significantly up-regulated expression patterns compared to the control group (healthy cows). The expression levels of HMOX1 and FTH1 were up-regulated in a dose-dependent manner in LPS induced MAC-T cells with increased iron accumulation. The expression levels of HMOX1 and FTH1 and iron accumulation levels in the MAC-T cells were significantly up-regulated by using LPS, but were lower than the levels seen with Erastin (ERA). Finally, we deduced the mechanism of ferroptosis in the MECs of Holstein cows with CM. These results provide new insights for the prevention and treatment of ferroptosis-mediated clinical mastitis in dairy animals.

CTH/H2S Regulates LPS-Induced Inflammation through IL-8 Signaling in MAC-T Cells.

Hydrogen sulfide (H2S), as an endogenous gaseous signaling molecule, plays an important role in the inflammatory process. Our previous study found that Cystathionine-γ-lyase (CTH) and H2S are correlated with the occurrence and development of Clinical Mastitis (CM) in Holstein cows. However, the functions and regulatory mechanisms of CTH/H2S are still unknown. In this study, the inflammatory mammary cell model based on the MAC-T cell line was established by Lipopolysaccharide (LPS)-induced manner to further explore the function and regulatory mechanism of CTH/H2S in cows with CM. In the inflammatory MAC-T cell, the CTH expression and H2S production were both repressed in an LPS-dose dependent manner, which demonstrated that CTH/H2S is related to the progression of inflammation. The inhibition of CTH/H2S using a selective CTH inhibitor, ß-cyano-l-Alanine (BCA), promoted LPS-induced inflammation response and the expression of inflammatory cytokines. However, this was reversed by the H2S donor NaHS, demonstrating that H2S can protect cells from inflammatory damage. Intriguingly, interleukin-8 (IL-8) showed an inverse expression pattern correlated with the H2S-mediated cell protection effect during the inflammation process, and the inhibition test using a selective IL-8 receptor antagonist, SB225002, showed that IL-8 signaling plays a critical role in mediating endogenous H2S synthesis, and CTH/H2S exerts its anti-inflammation via IL-8-mediated signaling. This study provided support for the prevention and treatment of CM and the development of a novel anti-inflammatory strategy.

Protocol for gene annotation, prediction, and validation of genomic gene expansion.

Although gene expansion plays an important role in evolution, its identification remains a challenge due to potential errors in genome assembly and annotation. Here, we describe a detailed step-by-step protocol for gene annotation, prediction of genomic gene expansion, and its computational and experimental validation. Finally, we also detail steps to discover functionality of each copy of replicated genes. For complete details on the use and execution of this protocol, please refer to Zhang et al. (2021).

Toll-like Receptor 2 Is Associated with the Immune Response, Apoptosis, and Angiogenesis in the Mammary Glands of Dairy Cows with Clinical Mastitis.

Toll-like receptor 2 (TLR2) plays a crucial role in bacterial recognition and the host immune response during infection. However, its function and downstream biological processes (BPs) in the mammary glands (MGs) of Holstein cows with clinical mastitis (CM) are not fully understood. This study aimed to comprehensively identify the BPs and differentially expressed proteins (DEPs) associated with the bacterial response and TLR2 using data-independent acquisition (DIA) proteomic data. A possible mechanism for the action of TLR2 was proposed, and the results suggested that the expression levels of TLR2 and caspase 8 (CASP8) were positively correlated with the apoptosis of MGs. The expression patterns of TLR2 and TEK receptor tyrosine kinase 2 (Tie2) were negatively correlated with angiogenesis. These results indicated that TLR2 might promote apoptosis in mammary epithelial cells (MECs) and vascular endothelial cells (VECs) via upregulation of CASP8 expression, and inhibition of angiogenesis in VECs via downregulation of Tie2 expression in dairy cows with CM. In conclusion, TLR2 is associated with inflammation, apoptosis, and angiogenesis in the MGs of dairy cows with bacteria-induced mastitis. These results contribute to a deeper understanding of the pathogenic mechanisms and provide the knowledge needed for developing the prevention and treatment of dairy mastitis.

Effects of lipoteichoic and arachidonic acids on the immune-regulatory mechanism of bovine mammary epithelial cells using multi-omics analysis.

Staphylococcus aureus is one of the most important pathogens causing mastitis in dairy cows. It mainly utilizes the properties of its pathogenic factor, lipoteichoic acid (LTA), to elicit a host-cell inflammatory response and evade the host-cell immune response. Arachidonic acid (AA) has a regulatory role in the inflammatory response, cell metabolism, and apoptosis. The study aimed to establish a cell model by determining the optimal concentration of LTA and AA for cell induction using the Cell Counting Kit-8 assay and the quantitative polymerase chain reaction of interleukin (IL)-1ß, IL-2, and IL-6. MAC-T cells were planted in 36 10-cm2 culture dishes at a density of 1 × 107 cells per dish. They were treated with LTA for 24 h to constitute the LTA group and with AA for 12 h to constitute the AA group. The cells were pretreated with LTA for 24 h followed by treatment with AA for 12 h to constitute the LTA + AA group. Using proteomic, transcriptomic, and metabolomic analyses, this study determined that LTA can regulate the expression of Actin Related protein 2/3 complex (ARPC)3, ARPC4, Charged Multivesicular Body Protein 3, protein kinase cGMP-dependent, NF-κB Inhibitor Alpha,and other genes to affect cellular metabolism, immune regulation and promote apoptosis. In contrast, AA was observed to regulate the expression of genes such as ARPC3, ARPC4, Charged Multivesicular Body Protein 3, Laminin Gamma 1, Insulin Receptor, Filamin B, and Casein Kinase 1 Epsilon to inhibit cellular apoptosis and promote immune regulation, which provides a theoretical basis for future studies.

M cells of mouse and human Peyer's patches mediate the lymphatic absorption of an Astragalus hyperbranched heteroglycan.

The gut cell wall is considered an impenetrable barrier to orally administrated polysaccharides. We recently reported a selective lymphatic route for Radix Astragali polysaccharide RAP to enter Peyer's patches (PPs) to trigger immune responses. However, how RAP enters PPs is unclear. Herein, we screened the intestinal epithelial cells of mice and found that the follicle-associated epithelium cells were specifically bound with FITC-RAP. Further studies in vitro and in vivo revealed that RAP was efficiently transported by microfold (M) cells. We also confirmed that M cell-transported RAP directly contacted dendritic cells. More importantly, for the first time, we verified this interesting M cell-mediated transcytosis of RAP in the human distal ileum. Mechanistically, we identified M cells to be the transporter cells that independently deliver RAP into the lymphatic system to trigger immune responses. This interesting transcytosis mechanism might apply to many other immunomodulatory polysaccharides orally dosed to human body.

Central administration of human opiorphin alleviates dextran sodium sulfate-induced colitis in mice through activation of the endogenous opioid system.

The opioid system plays a crucial role in maintaining gastrointestinal homeostasis. Endogenous opioid peptide enkephalins have anti-inflammatory effect and participate in the treatment of inflammatory bowel diseases (IBDs). Here, we investigated the effect of natural enkephalinase inhibitor human opiorphin (HO) on dextran sodium sulfate (DSS)-induced colitis in mice. Our results showed that central administration of HO attenuated DSS-induced colitis, as indicated by the reduction of disease activity index (DAI) scores, macroscopic scores, histological scores, and the myeloperoxidase (MPO) activity. Moreover, HO alleviated DSS-induced inflammation by decreasing inflammatory cytokines TNF-α, IL-6, and IL-1ß, and increasing anti-inflammatory cytokine IL-10 in both serum and colon tissues in DSS-treated mice. The potential anti-inflammatory effect of HO at a dose of 40 µg/kg was observed as evidenced by a decrease in nuclear factor κB (NF-κB) p65, toll-like receptor-4 (TLR-4), iNOS, and COX-2. HO also improved intestinal barrier function by enhancing the expression of tight junction proteins. Furthermore, HO treatment significantly inhibited activities of neutral endopeptidase (NEP) and aminopeptidase N (APN), elevated serum enkephalins concentrations, and increased expressions of mu and delta opioid receptors. In addition, pretreatment with opioid receptor antagonist naloxone hydrochloride (NH) compromised the protective effect of HO and aggravated colitis symptoms, as indicated by inhibited anti-inflammatory effects, disrupted intestinal barrier function, and decreased opioid receptor activity. In conclusion, these data indicate that HO protects against DSS-induced colitis by inhibiting TLR4/NF-κB pathway activation and improving intestinal barrier function through activation of the endogenous opioid system. Therefore, targeting the opioid system with peptidase inhibitors intervention would be a novel strategy in the therapy of IBD.

DIA proteomics identified the potential targets associated with angiogenesis in the mammary glands of dairy cows with hemorrhagic mastitis.

Hemorrhagic mastitis (HM) in dairy cows caused great economic losses in the dairy industry due to decreased milk production and increased costs associated with cattle management and treatment. However, the pathological and molecular mechanisms of HM are not well-understood. The present study aimed to investigate differentially expressed proteins (DEPs) associated with HM according to data-independent acquisition (DIA) proteomics. Compared to the mammary glands of healthylactating Holstein cows (Control, C group), the pathology of the HM group displayed massive alveolar infiltration of hemocytes and neutrophils, and the blood vessels, including arteriole, venules and capillaries were incomplete and damaged, with a loss of endothelial cells. DIA proteomics results showed that a total of 3,739 DEPs and 819 biological process terms were screened in the HM group. We focused on the blood, permeability of blood vessel, vascular and angiogenesis of mammary glands, and a total of 99 candidate DEPs, including 60 up- and 39 down-regulated DEPs, were obtained from the Gene Ontology (GO) and Pathway enrichment analyses. Phenotype prediction and function analysis of the DEPs revealed that three DEPs, particularly Caveolin-1(CAV1), were participated in the regulation of angiogenesis. Immunohistochemical and immunofluorescence staining showed that the CAV1 protein was present mainly in the mammary epithelial cells, vascular endothelial cells and vascular smooth muscle cells. The expression level of CAV1 mRNA and protein in the HM group was significantly down-regulated. The results will be helpful to the further understanding of the pathological and molecular mechanisms of HM in dairy cows.

Recent advances in qualitative and quantitative analysis of polysaccharides in natural medicines: A critical review.

Polysaccharides from natural medicines, being safe and effective natural mixtures, show great potential to be developed into botanical drugs. However, there is yet one polysaccharide-based case that has fulfilled the Botanical Guidance definition of a botanical drug product. One of the reasons is the analytical methods commonly used for qualitative and quantitative analysis of polysaccharides fall far behind the quality control criteria of botanical drugs. Here we systemically reviewed the recent advances in analytical methods. A critical evaluation of the strength and weaknesses of these methods was provided, together with possible solutions to the difficulties. Mass spectrometry with or without robust chromatographic separation was increasingly employed. And scientists have made significant progress in simplifying polysaccharide quantification by depolymerizing it into oligosaccharides. This oligosaccharides-based strategy is promising for qualitative and quantitative analysis of polysaccharides. And continuous efforts are still needed to develop a standardized quality control method that is specific, accurate, repeatable, and applicable for analyzing individual components in natural medicine formulas.