The Role of SHP2 in Advancing COPD: Insights into Oxidative Stress, Endoplasmic Reticulum Stress, and Pyroptosis.

Background: Chronic Obstructive Pulmonary Disease (COPD) is characterized by airflow limitation and inflammation resulting from genetic and environmental factors, notably cigarette smoke. Pyroptosis, a cell death process, is implicated in COPD, but its mechanisms are unclear. SHP2, a phosphatase, modulates inflammatory pathways, suggesting a role in COPD pathogenesis and potential therapeutic avenues. Objective: This study investigates the mechanism by which SHP2 regulates cell pyroptosis in bronchial epithelial cells in COPD patients. Methods: In this prospective study, we employed in vivo and in vitro models to investigate the mechanisms underlying COPD progression. Hematoxylin and eosin (H&E) staining were utilized to assess the morphological changes characteristic of COPD. Electron microscopy enabled precise quantification of pyroptotic bodies to highlight cellular changes associated with COPD pathogenesis. Immunofluorescence analysis facilitated the measurement of protein fluorescence intensity, allowing for the assessment of inflammatory responses within bronchial epithelial cells. Additionally, Western blot analysis was conducted to evaluate the expression levels of key pathway proteins involved in COPD progression. Results: In the COPD model, lesions worsened in SHP2-KD mice compared to SHP2-NC. Western blot results showed increased p22, p47, p-IRE1α, XBP1, STING, p-TBK1, NLRP3, Caspase1, and IL-1ß expression levels in both in vivo and in vitro models. Transmission electron microscopy revealed more pyroptotic bodies in SHP2-KD+CSE than in SHP2-NC+CSE. Immunofluorescence demonstrated significantly higher NLRP3 and GSDMD fluorescence intensities in SHP2-KD+CSE versus SHP2-NC+CSE. Additionally, Western blot analysis indicated increased expression of Bax, Caspase3, Caspase8, and Caspase9 proteins in the in vitro model. No differences were observed between SHP2 NC and SHP2-KD groups without CSE stimulation in immunofluorescence, electron microscopy, and Western blot findings in the cellular model. Conclusions: SHP2 promotes COPD progression by inducing oxidative stress, endoplasmic reticulum stress, and pyroptosis.

Formation, characterization, and antigenicity of lecithin-ß-conglycinin complexes.

Lipid binding has been proposed to represent a functional property of many allergenic proteins. This study investigated the formation, characterization, and antigenicity of lecithin-ß-conglycinin complexes. The results indicate that lecithin was combined with ß-conglycinin via static quenching and primarily driven by hydrogen bonds and van der Waals forces. In addition, heat treatment reduced the antigenicity of complexes, as evidenced by changes in molecular weight and secondary and tertiary structures. It revealed that large aggregates developed and more hydrophobic regions were exposed for complexes after heat treatment, as well as a decrease in the ß-sheet contents and an increase in the ß-turn and random coil contents. Furthermore, the average particle size of the complexes increased with increased temperature treatment, and the morphology of the complexes exhibited an amorphous polymer. These findings shedlight on the interaction between lecithin and ß-conglycinin and help us understand the role of lecithin in allergic reactions.

Linoleic acid enrichment of cheese by okara flour and Geotrichum candidum overexpressing Δ12 fatty acid desaturase.

BACKGROUND: Mold-ripened cheeses have low levels of unsaturated fatty acids (UFAs). Geotrichum candidum is an adjunct culture for the development of Geotrichum-ripened cheese but has a low ability to produce high levels of UFAs. Δ12 fatty acid desaturase (FADS12) is a pivotal enzyme that converts oleic acid (OA) to linoleic acid (LA) and plays a vital role in UFA biosynthesis. By investigating FADS12 catalytic activity from various species with OA substrates, we found that FADS12 from Mucor circinelloides (McFADS12) had the highest catalytic activity for OA. RESULTS: In the current study, a plasmid harboring McFADS12 was constructed and overexpressed in G. candidum. Our results showed that LA production increased to 31.1 ± 1.4% in engineered G. candidum - three times higher than that in wild-type G. candidum. To enhance LA production, an exogenous substrate (OA) was supplemented, and the yield of LA was increased to 154 ± 6 mg L-1 in engineered G. candidum. Engineered G. candidum was used as an adjunct culture for Geotrichum-ripened cheese production. The LA level reached 74.3 ± 5.4 g kg-1 cheese, whereas the level of saturated fatty acids (SFAs) decreased by 9.9 ± 0.5%. In addition, the soybean byproduct (okara) was introduced into the engineered G. candidum growth and the level of LA increased to 126 ± 4 g kg-1 cheese and the percentage of UFAs:SFAs increased from 0.8:1 to 1.3:1. CONCLUSION: This study offers a suitable technology for converting SFAs to UFAs in Geotrichum-ripened cheeses and provides a novel trend for converting soybean waste into a value-added product. © 2022 Society of Chemical Industry.

Anti-cancer Potential of Polysaccharide Extracted From Polygonatum sibiricum on HepG2 Cells via Cell Cycle Arrest and Apoptosis.

Polygonatum sibiricum is one of the most widely used traditional Chinese medicine in China. Polygonatum sibiricum polysaccharide (PSP) is the main functional component of Polygonatum sibiricum. In this study, a water-soluble polysaccharide (PSP-1) was first isolated from Polygonatum sibiricum with a molecular weight of 38.65 kDa. Structural analysis was performed via methylation and FT-IR spectroscopy analyses, which in combination with NMR spectroscopy, revealed that PSP-1 has a → 4-α-D-Glcp-1 → backbone with the substitution at O-6 with the ß-D-Glcp-1 → residues. Furthermore, PSP-1 exhibited potent and concentration-dependent anticancer effects, inducing HepG2 cell apoptosis and arresting the cell cycle at the G1 phase. Moreover, PSP-1 also decreased the mitochondrial membrane potential, damaged the nucleus of HepG2 cells, and increased the activity of caspase-9 and-3 in the intrinsic apoptotic pathways to induce HepG2 cell apoptosis. To conclude, PSP-1 might be a good candidate for the treatment of liver cancer, and this work provides important information for understanding the relationship between structure and antitumor activity of PSP-1, which is relevant for the treatment of hepatocellular carcinoma in clinic.

Role of the dietary components in food allergy: A comprehensive review.

Currently, the increasing incidence of food allergy is considered a major public health and food safety concern. Importantly, food-induced anaphylaxis is an acute, life-threatening, systemic reaction with varied clinical presentations and severity that results from the release of mediators from mast cells and basophils. Many factors are blamed for the increasing incidence of food allergy, including hygiene, microbiota (composition and diversity), inopportune complementary foods (a high-fat diet), and increasing processed food consumption. Studies have shown that different food components, including lipids, sugars, polyphenols, and vitamins, can modify the immunostimulating properties of allergenic proteins and change their bioavailability. Understanding the role of the food components in allergy might improve diagnosis, treatment, and prevention of food allergy. This review considers the role of the dietary components, including lipids, sugars, polyphenols, and vitamins, in the development of food allergy as well as results of mechanistic investigations in in vivo and in vitro models.

A multi center real world observation to evaluate the efficacy of cryotherapy versus local hyperthermia for the treatment of plane warts.

Most plane warts are recalcitrant to treatment. Both cryotherapy and local hyperthermia have been applied to treat plane warts. However, no direct comparative study on their respective efficacy and safety has ever been performed. To assess the efficacy and safety of local hyperthermia at 43 ± 1°C versus liquid nitrogen cryotherapy for plane warts. Sequential patients with plane warts entered the study, either receiving cryotherapy or local hyperthermia therapy at the discretion of the patients and the recommendations of consultants. Cryotherapy with liquid nitrogen was delivered in two sessions 2 weeks apart, while local hyperthermia was delivered on three consecutive days, plus two similar treatments 10 ± 3 days later. The temperature over the treated skin surface was set at 43 ± 1°C for 30 min in each session. The primary outcome was the clearance rates of the lesions 6 months after treatment. Among the 194 participants enrolled, 183 were included in the analysis at 6 months. Local hyperthermia and cryotherapy achieved clearance rates of 35.56% (48/135) and 31.25% (15/48), respectively (p = 0.724); recurrence rates of 16.67% (8/48) and 53.33% (8/15) (p = 0.01); and adverse events rates of 20.74% (28/135) and 83.33% (40/48), respectively (p < 0.001). Cryotherapy had a higher pain score (p < 0.001) and a longer healing time (p < 0.001). Local hyperthermia at 43°C and cryotherapy had similar efficacy for plane warts. Local hyperthermia had a safer profile than cryotherapy but it required more treatment visits during a treatment course. More patients preferred local hyperthermia due to its treatment friendly nature.

LAB Fermentation Improves Production of Bioactive Compounds and Antioxidant Activity of Withania somnifera Extract and Its Metabolic Signatures as Revealed by LC-MS/MS.

In this study we investigated the effect of lactic acid bacteria (LAB) fermentation on the ingredients and anti-oxidant activity of Withania somnifera extract. Four strains of LAB could proliferate normally in medium containing W. somnifera extract after the pH reached 3.1~3.5. LAB fermentation increased the content of alcohols and ketones, endowing the extract with the characteristic aroma of fermentation. Compared to the control, the DPPH and ABTS free radical scavenging rates in the fermented samples were significantly improved, ranging from 48.5% to 59.6% and 1.2% to 6.4%. The content of total phenols was significantly increased by 36.1% during the fermentation of mixed bacteria. Moreover, the original composition spectrum of the extract was significantly changed while the differentially accumulated metabolites (DAMs) were closely related to bile secretion, tryptophan metabolism and purine metabolism. Therefore, LAB fermentation can be used as a promising way to improve the flavor and bioactivity of the extracts of W. somnifera, making the ferments more attractive for use as functional food.

Local Hyperthermia Versus Cryotherapy for Treatment of Plantar Warts: A Prospective Multi-centre Non-randomized Concurrent Controlled Clinical Trial.

Cryotherapy is one of the most common treatments for warts; however, pain during treatment and relatively high recurrence rates limit its use. Local hyperthermia has also been used successfully in the treatment of plantar warts. The aim of this study was to compare the clinical effectiveness of local hyperthermia vs cryotherapy for the treatment of plantar warts. This multi- centre, open, 2-arm, non-randomized concurrent controlled trial included 1,027 patients, who received either cryotherapy or local hyperthermia treatment. Three months after treatment, local hyperthermia and cryotherapy achieved complete clearance rates of 50.9% and 54.3%, respectively. Recurrence rates were 0.8% and 12%, respectively. Pain scores during local hyperthermia were significantly lower than for cryotherapy. Both local hyperthermia and cryotherapy demonstrated similar efficacy for clearance of plantar warts; while local hyperthermia had a lower recurrence rate and lower pain sensation during treatment.

Sendeng-4 Suppressed Melanoma Growth by Induction of Autophagy and Apoptosis.

Sendeng-4 is a traditional Chinese medicine that has been successfully applied to anti-inflammatory diseases in clinical practice. Monomers within Sendeng-4 showed promising antitumor activity against lung cancer, colon cancer, and cutaneous cancer. However, potency of Sendeng-4 in melanoma has not been explored. This study aims to explore the potential application of Sendeng-4 in melanoma treatment. In the present study, we systemically investigate the possibility of Sendeng-4 for treatment of melanoma cancer in vitro by proliferation assay, colony formation, flow cell cytometry, RNA-seq, western blot, and fluorescence-based assay. Our data demonstrated that Sendeng-4 suppresses the proliferation and colony formation capacity of melanoma cells and induces cell cycle block at G2/M phase and eventually cell death. Mechanistically, transcriptome sequencing demonstrates that the PI3K-AKT pathway was significantly inactivated upon Sendeng-4 exposure, which was confirmed by western blot showing decreased phosphorylation of AKT. In addition, decreased BCL-2 expression and increased BAX expression were observed, suggesting programmed cell death via apoptosis. Moreover, LC3-II production as well as autophagosomes formation was observed as demonstrated by western blot and immunofluorescence, indicating elevated autophagy network by Sendeng-4 stimulation. Collectively, we concluded that Sendeng-4 might be used as an anticancer drug for melanoma.

Structural, rheological properties and antioxidant activities of polysaccharides from mulberry fruits (Murus alba L.) based on different extraction techniques with superfine grinding pretreatment.

The structural characteristics and biological activity of polysaccharides were influenced by different extraction methods. In this study, polysaccharides from mulberry fruits (Murus alba L., which were pre-treated with superfine grinding process) (MFP) were exacted using hot-water extraction (HWE), enzyme-assisted hot water extraction (EAHE), ultrasonic-assisted hot water extraction (UAHE), and high-speed shear homogenization-assisted hot water extraction (HSEHE). The extraction yield, structure, rheological properties and antioxidant activities of MFPs were investigated. MFP extracted using the HSEHE method have the highest extraction yields than other extraction methods. The smaller particle size of mulberry powder was found to improve the extraction yields. The MFPs were obtained by the combination between different extraction methods and superfine grinding pretreatment (through 100 mesh sieve) (MFP-HWE100, MFP-EAHE100, MFP-UAHE100, MFP-HSEHE100) showed the same levels of monosaccharide compositions and glycosyl linkages, However, these methods can produce MFP with different monosaccharide proportions, branching degree, different molecular weight, particle size and microstructure. MFP-HSEHE100 achieved the lowest molecular weight and particle size, which exhibited better thixotropy and antioxidant activities than other MFPs. This study identified that HSEHE was the most suitable extraction method for MFP.

Preparation and Characterization of Natural Quercetin-Based Mongolia Medicine Sendeng-4 Nanoemulsion (N-QUE-NE) and its Antibacterial Activity.

INTRODUCTION: Quercetin is the main active ingredient of Xanthoceras sorbifolia Bunge. Traditional compatibility theory of traditional Chinese medicine has typically reported a synergistic interaction among multiple components, while the synergistic effects of nanoemulsion have not been fully clarified. OBJECTIVE: The paper aims to study the preparation and characterization of quercetin-based Mongolia Medicine Sendeng-4 nanoemulsion (N-QUE-NE) and its antibacterial activity and mechanisms. METHODS: The morphology of the nanoemulsion was observed by Transmission Electron Microscopy (TEM), and the zeta potential, Polydispersity Index (PDI), and particle size distribution were determined by the nanometer particle size analyze. The stability of nanoemulsion was investigated by light test, high-speed centrifugal test and storage experiment at different temperatures. The combined bacteriostatic effect of N-QUE-NE was studied in vitro by the double-dilution method and checkerboard dilution method. RESULTS: The appearance of N-QUE-NE was pale yellow, clear and transparent. The nanoemulsion particles were spherical and uniformly distributed under TEM. The PDI was 0.052, the average particle size was 19.6nm, and the Zeta potential was -0.2mV. When quercetin nanoemulsion (QUENE) was used in combination with tannin nanoemulsion (TAN-NE) and toosendanin nanoemulsion (TOO-NE), it exhibited a synergistic antibacterial effect. However, the combination of QUE-NE and geniposide nanoemulsion (GEN-NE) exhibited an antagonistic effect. It was revealed that the antibacterial effect was in the order of quercetin-tannin-toosendanin nanoemulsion (QUE-TANTOO- NE) > quercetin-tannin nanoemulsion (QUE-TAN-NE) > QUE-NE > quercetin-tannintoosendanin- geniposide nanoemulsion (QUE-TAN-TOO-GEN-NE). CONCLUSION: This study explored the preparation and efficacy of N-QUE-NE, and the results showed that quercetin, tannin and toosendanin had satisfactory synergistic antibacterial effects. The antagonistic effect of quercetin and geniposide in nanoemulsion indicated that it is not beneficial to the antibacterial effect of Sendeng-4, and further research needs to be conducted to clarify its antibacterial effect.

Characterization and comparison of lipids in bovine colostrum and mature milk based on UHPLC-QTOF-MS lipidomics.

Lipids in bovine milk have several biological activities, with implications for human health and the physical functionality of foods. However, alterations in the lipid profile of bovine milk during lactation are not well-studied. This study aimed to identify differences in lipids between bovine colostrum and mature milk, using a lipidomics approach. Using an advanced mass spectrometry-based quantitative lipidomics approach, 335 lipids assigned to 13 subclasses were characterized in bovine colostrum (BC) and mature milk (BM). In total, 63 significantly differential lipids (SDLs) were identified. Among the 63 SDLs, the levels of 21 lipids were significantly lower in BM than in BC, including 5 glycerophosphatidylethanolamines (PEs), 1 glycerophosphatidylglycerol (PG), and 15 triacylglycerols (TGs). The levels of the remaining 42 lipids increased in BM, including 1 cardiolipin (CL), 9 diacylglycerols (DGs), 9 dihexosylceramides (Hex2Cers), 3 hexosylceramides (HexCers), 3 glycerophosphatidic acids (PAs), 2 glycerophosphatidylcholines (PCs), 12 PEs, and 3 TGs. Furthermore, the correlations and related metabolic pathways of these 63 SDLs were analyzed to explore the mechanisms that alter bovine milk lipids during lactation. The seven most relevant pathways identified herein, ranked in accordance with their degree of influence on lactation, were glycerophospholipid metabolism, sphingolipid metabolism, glycerolipid metabolism, glycosylphosphatidylinositol-anchor biosynthesis, linoleic acid metabolism, alpha-linolenic acid metabolism, and arachidonic acid metabolism. Our results provide essential insights into mechanisms underlying alterations in bovine milk lipids during different lactation periods, along with practical information of specific nutrition and quality assessments for the dairy industry.

Quantitative Phosphoproteomics of Milk Fat Globule Membrane in Human Colostrum and Mature Milk: New Insights into Changes in Protein Phosphorylation during Lactation.

Phosphorylation is a widespread posttranslational protein modification and is important in various biological processes. However, milk fat globule membrane (MFGM) phosphoproteins have not been explored systematically in human milk. Here, we used quantitative phosphoproteomics to analyze phosphorylation sites in human MFGM proteins and their differences at different stages of lactation; 305 phosphorylation sites on 170 proteins and 269 phosphorylation sites on 170 proteins were identified in colostrum and mature MFGM, respectively. Among these, 71 phosphorylation sites on 48 proteins were differentially expressed between the different stages of lactation. Osteopontin in human MFGM was the most heavily phosphorylated protein, with a total of 39 identified phosphorylation sites. Our results shed light on phosphorylation sites, composition, and biological functions of MFGM phosphoproteins in human colostrum and mature milk, and provide novel insights into the crucial roles of protein phosphorylation during infant development.

Effects of different probiotic combinations on the components and bioactivity of Spirulina.

Spirulina acts as a good dietary nutritional supplement. However, few research studies have been conducted on its fermentation. Three groups of probiotic combinations, lactic acid bacteria, Bacillus strains, and their mixture, were used to investigate Spirulina fermentation. The results showed that lactic acid bacteria significantly increased the content of amino acids and the ratio of essential amino acids to total amino acids in the fermented Spirulina, compared with the unfermented Spirulina, and this trend was enhanced by the strains' mixture. However, compared to unfermented Spirulina, the amino acid levels were significantly decreased after fermentation with Bacillus strains and so was the total free amino acid and essential amino acid content. Fermentation significantly reduced the contents of the offensive components of Spirulina, with significant differences among the three mixed bacterial treatments. Moreover, Bacillus strain fermentation increased the contents of flavonoids and polyphenols compared to the unfermented Spirulina, and significantly enhanced 1,1-diphenyl-2-trinitrophenylhydrazine free-radical scavenging ability and total antioxidant ability. On the contrary, treatments with lactic acid bacteria and the mixture of lactic acid bacteria and Bacillus strains endowed the fermented supernatants with good antibacterial ability. The results showed that probiotic fermentation has a good effect on Spirulina and can serve as a new procedure for developing new Spirulina-containing food items.

Revisit gut microbiota and its impact on human health and disease.

Trillions of microbes have evolved with and continue to live on human beings. With the rapid advances in tools and technology in recent years, new knowledge and insight in cross-talk between the microbes and their hosts have gained. It is the aim of this work to critically review and summarize recent literature reports on the role of microbiota and mechanisms involved in the progress and development of major human diseases, which include obesity, hypertension, cardiovascular disease, diabetes, cancer, Inflammatory Bowel Disease (IBD), gout, depression and arthritis, as well as infant health and longevity.

Quantitative analysis of amino acids in human and bovine colostrum milk samples through iTRAQ labeling.

BACKGROUND: The types and quantity of proteins vary widely between bovine and human milk, with corresponding differences in free and hydrolytic amino acids. In this study, the free and hydrolytic amino acids of bovine and human colostrum were for the first time qualitatively and quantitatively determined using isobaric tags for relative and absolute quantification technology combined with liquid chromatography tandem mass spectrometry detection. RESULTS: Total free amino acid content was 0.32 g L-1 and 0.63 g L-1 in bovine and human colostrum respectively, with free amino acid content in human colostrum twice that of bovine colostrum. However, total hydrolytic amino acid content was 4.2 g L-1 and 2.2 g L-1 in bovine and human colostrum respectively. We found that the hydrolytic amino acid content in bovine colostrum was higher than that in human colostrum; however, the amount of free amino acids and the overall amino acid content in human colostrum were respectively substantially higher and more varied than in bovine colostrum. CONCLUSION: Our findings revealed differences between bovine and human colostrum, with these data providing the basis for further research into amino acid metabolomics and infant formula. © 2018 Society of Chemical Industry.

Production of eicosapentaenoic acid by application of a delta-6 desaturase with the highest ALA catalytic activity in algae.

Dunaliella salina is a unicellular green alga with a high α-linolenic acid (ALA) level, but a low eicosapentaenoic acid (EPA) level. In a previous analysis of the catalytic activity of delta 6 fatty acid desaturase (FADS6) from various species, FADS6 from Thalassiosira pseudonana (TpFADS6), a marine diatom, showed the highest catalytic activity for ALA. In this study, to enhance EPA production in D. salina, FADS6 from D. salina (DsFADS6) was identified, and substrate specificities for DsFADS6 and TpFADS6 were characterized. Furthermore, a plasmid harboring the TpFADS6 gene was constructed and overexpressed in D. salina. Our results revealed that EPA production reached 21.3 ± 1.5 mg/L in D. salina transformants. To further increase EPA production, myoinositol (MI) was used as a growth-promoting agent; it increased the dry cell weight of D. salina transformants, and EPA production reached 91.3 ± 11.6 mg/L. The combination of 12% CO2 aeration with glucose/KNO3 in the medium improved EPA production to 192.9 ± 25.7 mg/L in the Ds-TpFADS6 transformant. We confirmed that the increase in ALA was optimal at 8 °C; the EPA percentage reached 41.12 ± 4.78%. The EPA yield was further increased to 554.3 ± 95.6 mg/L by supplementation with 4 g/L perilla seed meal (PeSM), 500 mg/L MI, and 12% CO2 aeration with glucose/KNO3 at varying temperatures. EPA production and the percentage of EPA in D. salina were 343.8-fold and 25-fold higher than those in wild-type D. salina, respectively. IMPORTANCE: FADS6 from Thalassiosira pseudonana, which demonstrates high catalytic activity toward α-linolenic acid, was used to enhance EPA production by Dunaliella salina. Transformation of FADS6 from Thalassiosira pseudonana into Dunaliella salina with myoinositol, CO2, low temperatures, and perilla seed meal supplementation substantially increased EPA production in Dunaliella salina to 554.3 ± 95.6 mg/L. Accordingly, D. salina could be a potential alternative source of EPA and is suitable for its large-scale production.

Quantitative N-glycoproteomics of milk fat globule membrane in human colostrum and mature milk reveals changes in protein glycosylation during lactation.

Milk fat globule membrane (MFGM) proteins have recently gained increasing attention, due to their significant biological function. However, the glycosylation of proteins in human MFGM during lactation has not been studied in detail. In this study, through mass spectroscopy-based N-glycoproteomics, we analyzed protein glycosylation of human MFGM. A total of 912 N-glycosylation sites on 506 N-glycoproteins were identified in human colostrum and mature milk MFGM. Among them, 220 N-glycoproteins with 304 N-glycosylation sites were differentially expressed in colostrum and mature milk MFGM. Gene Ontology (GO) analysis revealed various biological processes, cellular components, and molecular functions of the differentially expressed N-glycoproteins. Specifically, these glycoproteins were involved in biological processes such as single-organism processes, biological regulation, regulation of biological processes, response to stimulus and localization; were cellular components in organelles, membranes, and the extracellular region; and had different molecular functions such as protein binding, receptor activity, and hydrolase activity. KEGG pathway analysis suggested that the majority of the differentially expressed N-glycoproteins were associated with phagosome, cell adhesion molecule and some disease-related pathways. Our results provide an in-depth understanding of the quantitative changes in N-glycosylation of proteins in human colostrum and mature MFGM, and extend our knowledge of the N-glycoproteome and of the distribution of N-glycosylation sites in human MFGM during lactation, providing insight into the biological functions of the highlighted glycoproteins.

Comparative Analysis of Whey N-Glycoproteins in Human Colostrum and Mature Milk Using Quantitative Glycoproteomics.

Glycosylation is a ubiquitous post-translational protein modification that plays a substantial role in various processes. However, whey glycoproteins in human milk have not been completely profiled. Herein, we used quantitative glycoproteomics to quantify whey N-glycosylation sites and their alteration in human milk during lactation; 110 N-glycosylation sites on 63 proteins and 91 N-glycosylation sites on 53 proteins were quantified in colostrum and mature milk whey, respectively. Among these, 68 glycosylation sites on 38 proteins were differentially expressed in human colostrum and mature milk whey. These differentially expressed N-glycoproteins were highly enriched in "localization", "extracellular region part", and "modified amino acid binding" according to gene ontology annotation and mainly involved in complement and coagulation cascades pathway. These results shed light on the glycosylation sites, composition and biological functions of whey N-glycoproteins in human colostrum and mature milk, and provide substantial insight into the role of protein glycosylation during infant development.

Comparative proteomic analysis of milk-derived exosomes in human and bovine colostrum and mature milk samples by iTRAQ-coupled LC-MS/MS.

Exosomes are membranous vesicles found in biological fluids with important functions. However, milk-derived exosome proteins from humans and bovines have not been studied in detail. The advanced iTRAQ proteomic approach was used to analyze milk-derived exosomes in human and bovine colostrum and mature milk samples. A total of 920 milk exosome proteins were identified and quantified. Among these, 575 differentially expressed exosome proteins (P<0.05) were found. Multivariate analysis, gene ontology (GO) annotation and the KEGG pathway were used to interpret the identified proteins. The major biological processes involved were: response to stimulus (22%), localization (16%), establishment of localization (14%), and cellular component organization (14%). Cellular components engaged in intracellular (31%) and intracellular part (31%). The most prevalent molecular function mainly touched upon binding (52%). Milk exosome proteins participated in several KEGG pathways containing ribosome, regulation of actin cytoskeleton, glycolysis/gluconeogenesis, leukocyte transendothelial migration, aminoacyl-tRNA biosynthesis, pentose phosphate pathway, galactose metabolism and fatty acid biosynthesis. These results provide important information on human and bovine milk exosomes, and increase knowledge on the proteomes of these exosomes across different lactation stages, which could provide potential directions for newborn milk powder, biological markers and functional foods.