ABSTRACT
Cerebral metabolic dysfunction is a critical pathological hallmark observed in the aftermath of traumatic brain injury (TBI), as extensively documented in clinical investigations and experimental models. An in-depth understanding of the bioenergetic disturbances that occur following TBI promises to reveal novel therapeutic targets, paving the way for the timely development of interventions to improve patient outcomes. The 13C isotope tracing technique represents a robust methodological advance, harnessing biochemical quantification to delineate the metabolic trajectories of isotopically labeled substrates. This nuanced approach enables real-time mapping of metabolic fluxes, providing a window into the cellular energetic state and elucidating the perturbations in key metabolic circuits. By applying this sophisticated tool, researchers can dissect the complexities of bioenergetic networks within the central nervous system, offering insights into the metabolic derangements specific to TBI pathology. Embraced by both animal studies and clinical research, 13C isotope tracing has bolstered our understanding of TBI-induced metabolic dysregulation. This review synthesizes current applications of isotope tracing and its transformative potential in evaluating and addressing the metabolic sequelae of TBI.
Subject(s)
Brain Injuries, Traumatic , Animals , Humans , Brain Injuries, Traumatic/metabolism , Energy Metabolism , IsotopesABSTRACT
Adherent-invasive Escherichia coli plays an important role in the pathogenesis of inflammatory bowel disease. Blocking the adhesion of E. coli to intestinal epithelial cells appears to be useful for attenuating inflammatory bowel disease. Lycopene has been reported to have anti-inflammatory and antimicrobial activities. The aim of this study was to test the intervention effect of lycopene on colitis in mice and to investigate the possible mechanism through which lycopene affects the adhesion of E. coli to intestinal epithelial cells. Lycopene (12 mg/kg BW) attenuated dextran sulfate sodium (DSS)-induced colitis, decreased the proportion of E. coli, and activated the NLR family pyrin domain containing 12 and inactivated nuclear factor kappa B pathways. Furthermore, lycopene inhibited the adhesion of E. coli O157:H7 to Caco-2 cells by blocking the interaction between E. coli O157:H7 and integrin ß1. Lycopene ameliorated DSS-induced colitis by improving epithelial barrier functions and inhibiting E. coli adhesion. Overall, these results show that lycopene may be a promising component for the prevention and treatment of colitis.
Subject(s)
Colitis , Inflammatory Bowel Diseases , Humans , Animals , Mice , Lycopene/pharmacology , Escherichia coli , Caco-2 Cells , Intestinal Mucosa/metabolism , Colitis/chemically induced , Colitis/drug therapy , Colitis/metabolism , Dextran Sulfate/adverse effects , Mice, Inbred C57BL , Disease Models, Animal , Colon/metabolismABSTRACT
In China, 45% of adolescents with obesity develop fatty liver disease, a condition that increases the long-term risk of developing cirrhosis and liver cancer. Although the factors triggering nonalcoholic fatty liver disease (NAFLD) vary in children, the composition of intestinal microflora has been found to play an increasingly important role. However, evidence is limited on the prevalence of nonalcoholic fatty liver (NAFL) and nonalcoholic steatohepatitis (NASH) in Chinese children. Therefore, this study aimed to evaluate the fecal microbiome of Chinese children with NAFLD and further analyze the potential of flora in regulating NAFLD-related symptoms and metabolic functions. Specifically, the study applied a 16S rRNA and metagenomic sequencing to the fecal samples of pediatric patients with NAFLD, NASH, and NAFL, as well as healthy controls, to explore the correlation among NAFLD-related indexes, metabolic pathways, and gut flora. The findings showed that some fecal microbiota had a negative correlation with body mass index, and various NAFLD-related bacteria, including Lachnoclostridium, Escherichia-Shigella, and Faecalibacterium prausnitzii, were detected. Consequently, the study concluded that the variation in gut microbiota might be more important in improving NAFLD/NASH compared with single species, providing a microbiota diagnostic profile of NAFLD/NASH.IMPORTANCEThis study aims to characterize the gut microbiota in Chinese children with nonalcoholic fatty liver disease (NAFLD) through 16S rRNA and metagenomic sequencing. The results highlight the association between fecal microbiota and NAFLD in Chinese children, demonstrating distinct characteristics compared to adults and children from other countries. Based on the sequencing data from our cohort's fecal samples, we propose a microbiota model with a high area under the curve for distinguishing between NAFLD and healthy individuals. Furthermore, our follow-up study reveals that changes in the relative abundance of microbial biomarkers in this model are consistent with variations in patients' body mass index. These findings suggest the potential utility of the microbiota model and microbial biomarkers for diagnosing and treating NAFLD in children.
Subject(s)
Gastrointestinal Microbiome , Non-alcoholic Fatty Liver Disease , Adult , Adolescent , Humans , Child , RNA, Ribosomal, 16S , Follow-Up Studies , Biomarkers/metabolism , Liver/metabolismABSTRACT
Probiotics have been increasingly investigated for their role in alleviating symptoms of ulcerative colitis (UC), but the specific mechanism involved remains unclear. We investigated the alleviating effect of Bifidobacterium animalis A6 (BAA6) in UC through a mouse dextran sulfate sodium (DSS) model. When treated with a high dose of BAA6 (1 × 1010 cfu/ml), it was found that colitis symptoms were significantly alleviated, and mucosal damages experienced obvious relief. Moreover, a high dose of BAA6 effectively upregulated free fatty acid receptors 2 and 3 (FFAR2 and FFAR3) expression and butyric acid metabolism specifically. Furthermore, the supplement of BAA6 significantly suppressed pro-inflammatory cytokines levels (interleukin-13) and the expression of pore-forming protein claudin-2. The upstream regulatory genes of claudin-2, such as STAT6, GATA4, Cdx2, were also significantly inhibited by BAA6. Collectively, this study concludes that BAA6 attenuated DSS-induced colitis by increasing the levels of intestinal butyric acid, activating the butyric acid-FFAR pathway, suppressing excessive proinflammatory response, and protecting the function of the colon epithelial barrier.
ABSTRACT
Cerebral ischemia activates neural progenitors that participate in brain remodeling following acute injury. Sphingosine-1-phosphate receptor (S1PR) signaling governs cell proliferation and mobilization, yet its potential impact on neural progenitors and stroke recovery remains poorly understood. The goal of this study was to investigate the impact of S1PR modulation on post-stroke neurogenesis and functional recovery, using a S1PR modulator BAF312. Mice were subjected to 60 min middle cerebral artery occlusion (MCAO) and received BAF312 starting from day 3 after MCAO until the end of experiment. BAF312 facilitated motor function recovery in MCAO mice until day 14 after surgery. Flow cytometry analysis revealed that BAF312 treatment led to an increase of type A cells in subventricular zone (SVZ), but not other progenitor cell subsets in MCAO mice. We found an increase of BrdU incorporation in SVZ DCX+ cells from MCAO mice receiving BAF312 and augmented proliferation of DCX+ cells in cultured neurospheres isolated from SVZ tissues. Notably, a S1PR1 antagonist W146 abolished BAF312-induced increase of SVZ type A cells from MCAO mice and proliferation of DCX+ cells in cultured neurospheres. Additionally, the benefit of BAF312 to improve neurogenesis and stroke recovery remains in Rag2-/- mice lacking of T and B cells. Our results demonstrate that S1PR modulation improves neurogenesis and functional recovery following brain ischemia.
Subject(s)
Brain Ischemia , Stroke , Mice , Animals , Recovery of Function , Sphingosine-1-Phosphate Receptors , Neurogenesis/physiology , Stroke/drug therapy , Infarction, Middle Cerebral ArteryABSTRACT
Akkermansia muciniphila is well known for the amelioration of inflammatory responses and restoration of intestinal barrier function. The beneficial effect of A. muciniphila occurred through contacting Toll-like receptor 2 (TLR2) on intestinal epithelial cells by wall components. In this case, the downstream mechanism of pasteurized A. muciniphila stimulating TLR2 for ameliorated intestinal barrier function is worth investigating. In this study, we evaluated the effect of live and pasteurized A. muciniphila on protecting the barrier dysfunction of Caco-2 intestinal epithelial cells induced by lipopolysaccharide (LPS). We discovered that both live and pasteurized A. muciniphila could attenuate an inflammatory response and improve intestinal barrier integrity in Caco-2 monolayers. We demonstrated that A. muciniphila enhances AMP-activated protein kinase (AMPK) activation and inhibits Nuclear Factor-Kappa B (NF-κB) activation through the stimulation of TLR2. Overall, we provided a specific mechanism for the probiotic effect of A. muciniphila on the intestinal barrier function of Caco-2 cells.
Subject(s)
NF-kappa B , Toll-Like Receptor 2 , AMP-Activated Protein Kinases/metabolism , Akkermansia , Caco-2 Cells , Humans , Intestinal Mucosa/metabolism , Lipopolysaccharides/pharmacology , NF-kappa B/metabolism , Toll-Like Receptor 2/metabolismABSTRACT
Mimops orientalis Kraepelin, 1903 is a monotypic species of Mimopidae endemic to China. The species is known only from a single specimen, the holotype. Little is known about its biology, habitat associations, or phylogenetic relationships. It was rediscovered on Qinling Mountain in Shaanxi and Henan provinces, China, 117 years after its last record. Detailed descriptions and colour photographs of living specimens are provided along with its ecology, updated conservation notes, and data on sexual dimorphism. A genetic analysis (COI, 16S rRNA, and 28S rRNA) was conducted to assess the phylogenetic relationships among Mimopidae, Cryptopidae, Scolopendridae, Scolopocryptopidae, and Plutoniumidae. The results support classifying Mimopidae as a valid family.
ABSTRACT
High circulating branched-chain amino acid (BCAA) levels can be diagnosis indicators for obesity. Luffa cylindrica (luffa) is one of vegetables against obesity. However, whether the anti-obesity of luffa is associated with BCAA metabolism and gut microbiota remains unknown. Here, we used conventionally raised diet-induced obese (DIO) mice to prove dietary luffa could reduce higher circulating BCAA levels and upregulate the tissue-specific expressions of BCAA-catabolizing enzymes. Meanwhile, dietary luffa selectively decreased the relative abundances of g_Enterortabdus, g_Eubacterium_xylanophilum_group and g_Butyricicoccus that exhibited significantly positive correlations with BCAA levels, BMI and HOMA-IR. Bacterial functionality prediction indicated dietary luffa potentially inhibited bacterial BCAA biosynthesis for reducing BCAAs supplementation. More importantly, dietary luffa had no impacts on BCAA catabolism in germ-free-mimic DIO mice. Thus, dietary luffa improved BCAA dysfunction via gut microbiota to attenuate obesity. This study offers a novel insight into dietary intervention against obesity from the aspect of gut microbiota-amino acid metabolism.
Subject(s)
Amino Acids, Branched-Chain/metabolism , Anti-Obesity Agents/pharmacology , Gastrointestinal Microbiome/physiology , Luffa , Obesity/diet therapy , Animals , Diet, High-Fat/adverse effects , Dietary Supplements , Insulin Resistance , Male , Mice, Inbred C57BL , Mice, Obese , Obesity/etiologyABSTRACT
Luffa cylindrica is a nutrient-dense vegetable with medical properties and can alleviate metabolic diseases. Numerous evidences demonstrated gut microbiota impacted the progress of nonalcoholic fatty liver disease (NAFLD). This study was to investigate the underlying mechanism of L. cylindrica supplementation against NALFD via gut microbiota from hepatic transcriptional and metabolic analysis. In diet-induced obese mice, we observed L. cylindrica supplementation (2 g/kg body weight) effectively alleviated high-fat diet-induced obese symptoms such as body weight, fat deposition, and insulin resistance. Notably, L. cylindrica supplementation significantly relieved hepatic steatosis and inflammation infiltration to decrease hepatic toxicity. RNA-sequencing analysis showed that 130 hepatic genes in total significantly altered responding to L. cylindrica supplementation. And signaling pathway analysis revealed that L. cylindrica supplementation down-regulated the transcriptional expressions of CD36 and Rxrg to inhibit hepatic lipid synthesis. Moreover, L. cylindrica supplementation increased the transcriptional expressions of Ass1, Cps1, Cth, Got1, Tat, and Gls2 to enhance amino acid levels (Gly, Ala, Pro, Val, Ile, Asn, Met, and Phe) and improve hepatic abnormal gluconeogenesis. Furthermore, in antibiotic-treated obese mice, L. cylindrica supplementation did not change these gene expressions along with the hepatic levels of lipid and amino acids. Taken together, L. cylindrica supplementation could effectively suppress hepatic steatosis in diet-induced obese mice through inhibiting lipid synthesis and enhancing amino acid levels in liver, which depended on gut microbiota. Thus, L. cylindrica might be one promising dietary supplementation targeting at gut microbiota to reduce NAFLD risk.
Subject(s)
Amino Acids/metabolism , Gastrointestinal Microbiome/drug effects , Luffa/chemistry , Non-alcoholic Fatty Liver Disease/drug therapy , Plant Extracts/pharmacology , Animals , CD36 Antigens/metabolism , Diet, High-Fat , Dietary Supplements , Gene Expression , Gene Expression Profiling , Insulin Resistance , Lipogenesis/drug effects , Liver/metabolism , Male , Metabolic Diseases/metabolism , Mice , Mice, Inbred C57BL , Mice, Obese , Non-alcoholic Fatty Liver Disease/genetics , Non-alcoholic Fatty Liver Disease/metabolism , Plant Extracts/administration & dosage , Vegetables/chemistry , Vegetables/metabolismABSTRACT
The gut microbiota, identified as the target for vegetables, can affect the development of obesity and associated metabolic syndromes. As a medicinal and edible plant, Luffa cylindrica (L.) Roem (LC) has abundant nutrients that can effectively reduce obesity risk. However, the interaction between the prevention effects of LC against obesity and the modulating gut microbiota of LC remain obscure. This study demonstrated LC supplementation improved high-fat diet (HFD)-induced gut microbiota dysbiosis and significantly enhanced short-chain fatty acid (SCFA)-producing bacteria (e.g., Blautia) along with SCFA content accumulation in the gut. Meanwhile, LC supplementation substantially restored gut barrier damage in long-term HFD treatment. Moreover, LC supplementation improved HFD-induced overweight, hyperlipidemia, insulin resistance, and chronic inflammation. Gene expression profiles showed that LC displayed an important impact on hepatic lipid transport and lipid synthesis (sterol regulatory element binding transcriptional factor 1c-peroxisome proliferator-activated receptor γ signaling pathway). More importantly, an antibiotic treatment experiment demonstrated that the beneficial effects of LC in reducing obesity risk largely depended on the gut microbiota, especially SCFA-producing bacteria (e.g., Blautia). Therefore, LC supplementation improved gut microbiota dysbiosis via enhancing SCFA-producing bacteria (e.g., Blautia), maintained gut barrier integrity, and alleviated the development of obesity. Overall, LC would provide a potential dietary intervention strategy against obesity and enteral homeostasis dysbiosis through modulating the gut microbiota.-Zhang, L., Shi, M., Ji, J., Hu, X., Chen, F. Gut microbiota determines the prevention effects of Luffa cylindrica (L.) Roem supplementation against obesity and associated metabolic disorders induced by high-fat diet.
Subject(s)
Diet, High-Fat/adverse effects , Dietary Supplements , Gastrointestinal Microbiome/drug effects , Inflammation/prevention & control , Luffa/chemistry , Metabolic Diseases/prevention & control , Obesity/complications , Animals , Inflammation/etiology , Inflammation/pathology , Male , Metabolic Diseases/etiology , Metabolic Diseases/pathology , Mice , Mice, Inbred C57BLABSTRACT
AIM: To achieve enhanced anticancer efficacy by combined siPlk1 and curcumin (cur) therapy using α-lactalbumin (α-lac) nanocarrier delivery. MATERIALS & METHODS: α-Lac was partially hydrolyzed into amphiphilic peptides, and then self-assembled into nanospheres (NS). Cur was loaded into their hydrophobic core during the self-assembly process. siPlk1-SH was cross-linked with the endogenous cysteines on the NS. CRGDK peptide was conjugated on NS to target integrins overexpressed in HeLa cells. RESULTS & CONCLUSION: The Cur and siPlk1 coloaded NS formulations possessed an enhanced tumor targeting and antitumor properties. Drugs were responsively released from disulfide bonds cross-linked RGD-NS/Cur/siPlk1 corresponding to the high intracellular glutathione concentrations of cancer cells. Both in vitro cell viability experiments and in vivo antitumor evaluations demonstrated that the codelivered nanosphere platform exhibited excellent tumor targeting and synergistic antitumor efficacy.
Subject(s)
Curcumin/chemistry , Lactalbumin/chemistry , Nanospheres/chemistry , Animals , Antineoplastic Agents/chemistry , Drug Compounding , Drug Delivery Systems/methods , Female , Flow Cytometry , HeLa Cells , Humans , Lysosomes/chemistry , Mice , Mice, Inbred BALB C , Micelles , RNA, Messenger/genetics , Rats , Real-Time Polymerase Chain Reaction , Xenograft Model Antitumor AssaysABSTRACT
In this study, pH-responsive microspheres loaded with multiple antioxidants were developed for intestine-specific delivery and exhibited synergistic activity. They consist of chitosan (CS)-coated microspheres made of TEMPO-oxidized Konjac glucomannan (OKGM) polymers, of which the carboxyl (COO-) groups are cross-linked via ferric ions (Fe3+), allowing the hydrophobic (ß-carotene) and hydrophilic (anthocyanins) antioxidants to be simultaneously incorporated. CLSM images showed successful co-encapsulation of ß-carotene and anthocyanins. The in vitro release kinetics of co-loaded CS-OKGM microspheres in simulated GI fluids indicated that the microspheres retain the dual antioxidants in an acidic gastric environment and release them at intestinal pH. Free radical scavenging experiments demonstrated that multiple antioxidants loaded into OKGM system had synergistic activity with enhanced stability against heat. The multi-functional CS-OKGM microspheres showed great potential for multiple antioxidants and intestine-specific delivery with enhanced stability.
Subject(s)
Antioxidants/administration & dosage , Cyclic N-Oxides , Drug Carriers/chemistry , Microspheres , Polysaccharides/chemistry , Anthocyanins/administration & dosage , HEK293 Cells , Humans , Intestines , beta Carotene/administration & dosageABSTRACT
TEMPO-oxidized Konjac glucomannan (OKGM) was developed as new material for preparing vegetarian hard capsules. OKGM of different degrees of oxidation: DO30%, DO50%, and DO80% were prepared to select optimum DO for capsule formation. FT-IR results proved that the primary alcohol groups on KGM were oxidized into carboxyl groups. XRD analysis suggested that TEMPO-oxidation decreased the crystallinity of KGM. DO80% was considered as the optimum candidate for capsule preparation owing to its superior solubility, transparency and reduced viscosity. The hydrophilicity of OKGM films, measured by contact angle measurement, increased with increasing DO. The elongation at break and tensile strength of the OKGM films enhanced with increasing DO. In vitro drug dissolution profile of OKGM capsules showed that the shell rupture time of DO80% capsule is about 5-10 min, and 80% of the drugs were released within 30-45 min. Thus DO80% OKGM was qualified to be used for gastric soluble hard capsules.
Subject(s)
Mannans/chemistry , Piperidines/chemistry , Amorphophallus , Capsules , Drug Liberation , Erythromycin/analogs & derivatives , Erythromycin/chemistry , Mannans/chemical synthesis , Oxidation-Reduction , Spectroscopy, Fourier Transform Infrared , Tensile Strength , Viscosity , beta-Mannosidase/chemistryABSTRACT
The epicardium contributes both multi-lineage descendants and paracrine factors to the heart during cardiogenesis and cardiac repair, underscoring its potential for cardiac regenerative medicine. Yet little is known about the cellular and molecular mechanisms that regulate human epicardial development and regeneration. Here, we show that the temporal modulation of canonical Wnt signaling is sufficient for epicardial induction from 6 different human pluripotent stem cell (hPSC) lines, including a WT1-2A-eGFP knock-in reporter line, under chemically-defined, xeno-free conditions. We also show that treatment with transforming growth factor beta (TGF-ß)-signalling inhibitors permitted long-term expansion of the hPSC-derived epicardial cells, resulting in a more than 25 population doublings of WT1+ cells in homogenous monolayers. The hPSC-derived epicardial cells were similar to primary epicardial cells both in vitro and in vivo, as determined by morphological and functional assays, including RNA-seq. Our findings have implications for the understanding of self-renewal mechanisms of the epicardium and for epicardial regeneration using cellular or small-molecule therapies.
ABSTRACT
An intestine-specific delivery system for hydrophobic bioactives with improved stability was developed. It consists of oxidized potato starch polymers, where the carboxyl groups were physically cross-linked via ferric ions. The model hydrophobic ingredients (ß-carotene) were incorporated inside the starch microspheres via a double-emulsion method. Confocal laser scanning microscopy images showed that ß-carotene were distributed homogeneously in the inner oil phase of the starch microspheres. The negative value of the ζ-potential of microspheres increased with increasing pH and decreasing ionic strength. In vitro release experiments showed that the microspheres were stable at acidic stomach conditions (pH < 2), whereas at neutral intestinal conditions (pH 7.0), they rupture to release the loaded ß-carotene. The 1,1-diphenyl-2-picrylhydrazyl radical, 2,2-diphenyl-1-(2,4,6-trinitriphenyl), scavenging activity results suggested that microsphere-encapsulated ß-carotene had an improved activity after thermal treatment at 80 °C. The storage stability of encapsulated ß-carotene at room temperature was also enhanced. The starch microspheres showed potential as intestine-specific carriers with an enhanced stability.
Subject(s)
Drug Carriers/chemistry , Drug Delivery Systems/methods , Intestines/drug effects , Starch/chemistry , beta Carotene/chemistry , Drug Delivery Systems/instrumentation , Drug Stability , Drug Storage , Humans , Hydrophobic and Hydrophilic Interactions , Microspheres , Osmolar Concentration , Oxidation-Reduction , Particle Size , Plant Extracts/chemistry , Solanum tuberosum/chemistry , beta Carotene/pharmacologyABSTRACT
Human pluripotent stem cell (hPSC)-derived endothelial cells and their progenitors are important for vascular research and therapeutic revascularization. Here, we report a completely defined endothelial progenitor differentiation platform that uses a minimalistic medium consisting of Dulbecco's modified eagle medium and ascorbic acid, lacking of albumin and growth factors. Following hPSC treatment with a GSK-3ß inhibitor and culture in this medium, this protocol generates more than 30% multipotent CD34+ CD31+ endothelial progenitors that can be purified to >95% CD34+ cells via magnetic activated cell sorting (MACS). These CD34+ progenitors are capable of differentiating into endothelial cells in serum-free inductive media. These hPSC-derived endothelial cells express key endothelial markers including CD31, VE-cadherin, and von Willebrand factor (vWF), exhibit endothelial-specific phenotypes and functions including tube formation and acetylated low-density lipoprotein (Ac-LDL) uptake. This fully defined platform should facilitate production of proliferative, xeno-free endothelial progenitor cells for both research and clinical applications.
Subject(s)
Albumins/chemistry , Cell Differentiation , Endothelial Progenitor Cells/cytology , Pluripotent Stem Cells/cytology , Antigens, CD/metabolism , Cell Differentiation/drug effects , Culture Media/pharmacology , Endothelial Progenitor Cells/drug effects , Humans , Multipotent Stem Cells/cytology , Multipotent Stem Cells/drug effectsABSTRACT
A biodegradable microgel system based on glycerol-1,3-diglycidyl ether (GDGE) cross-linked TEMPO-oxidized potato starch polymers was developed for controlled uptake and release of proteins. A series of microgels were prepared with a wide range of charge density and cross-link density. We found both swelling capacity (SWw) and lysozyme uptake at saturation (Γsat) increased with increasing degree of oxidation (DO) and decreasing cross-link density. Microgel of DO100% with a low cross-link density (RGDGE/polymer (w/w) of 0.025) was selected to be the optimum gel type for lysozyme absorption; Γsat increased with increasing pH and decreasing ionic strength. It suggests that the binding strength was the strongest at high pH and low ionic strength, which was recognized as the optimum absorption conditions. The lysozyme release was promoted at low pH and high ionic strength, which were considered to be the most suitable conditions for triggering protein release. These results may provide useful information for the controlled uptake and release of proteins by oxidized starch microgels.
Subject(s)
Epoxy Compounds/chemistry , Gels/chemistry , Glyceryl Ethers/chemistry , Muramidase/chemistry , Starch/chemistry , Cross-Linking Reagents/chemistry , Oxidation-ReductionABSTRACT
A Pickering (o/w) emulsion was formed and stabilized by whey protein isolate nanoparticles (WPI NPs). Those WPI NPs were prepared by thermal cross-linking of denatured WPI proteins within w/o emulsion droplets at 80°C for 15 min. During heating of w/o emulsions containing 10% (w/v) WPI proteins in the water phase, the emulsions displayed turbid-transparent-turbid phase transitions, which is ascribed to the change in the size of the protein-containing water droplets caused by thermal cross-linking between denatured protein molecules. The transparent stage indicated the formation of WPI NPs. WPI NPs of different sizes were obtained by varying the mixing speed. WPI NPs of 200-500 nm were selected to prepare o/w Pickering emulsions because of their good stability against coalescence. By Confocal Laser Scanning Microscopy, it was observed that WPI NPs were closely packed and distributed at the surface of the emulsion droplets. By measuring water contact angles of WPI NPs films, it was found that under most conditions WPI NPs present good partial wetting properties, but that at the isoelectric point (pI) and high ionic strength the particles become more hydrophobic, resulting in less stable Pickering emulsion. Thus, at pH above and below the pI of WPI NPs and low to moderate ionic strengths (1-10 mM), and with a WPI NPs concentration of 2% (w/v), a stable Pickering emulsion can be obtained. The results may provide useful information for applications of WPI NPs in environmentally friendly and food grade applications, notably in food, pharmaceutical and cosmetic products.