Early life cow's milk allergy prevention: PLGA Nanoparticles for the Oral Delivery of β-Lactoglobulin Derived Peptide and CpG Oligodeoxynucleotides

The prevalence and lack of good solutions for cow's milk allergy (CMA) in young children are leading to public concern and increasing socioeconomic problems. This requires new preventive approaches. In this thesis, we investigated the potential of an oral formulation – based on poly(lactic acid-co-glycolic acid) (PLGA) nanoparticles (NP) – of selected peptides from whey proteins and CpG-Oligodeoxynucleotides for CMA prevention, using both in vitro and in vivo studies. In vitro, we showed, through Förster Resonance Energy Transfer (FRET) and imaging in living cells, that PLGA NP protected the encapsulated peptides for up to 6 hours in simulated gastric fluid and extended their residence time in isolated mouse dendritic cells to 96 hours. This demonstrates the hypothesis that PLGA nanoparticles improved the stability of the encapsulated peptide cargo in the gastrointestinal lumen and retention in intestinal DC upon internalization. In vivo, we showed that oral pretreatment with two PLGA-NP-encapsulated peptides, prior to allergic sensitization to whey, protected mice from acute allergic skin reactions upon intradermal exposure to whey. In addition, we have shown that this protection is dose-related and may be associated with whey-specific systemic silencing of ex vivo whey-stimulated TNF-α release by splenocytes. Intriguingly, in vivo, we showed the superior tolerogenic effect of orally administered PLGA nanoparticles co-loaded with T-cell epitopes containing peptide and a CpG adjuvant as compared to the separately encapsulated peptide and CpG counterpart. The combination of peptides and CpG prevented rise in specific IgE antibody levels in serum and whey-induced acute allergic skin reactions, and showed a beneficial immunomodulatory effect on immune cells. Finally, allergy-associated immunomodulatory effects of empty, peptide or CpG-containing PLGA nanoparticles were evaluated in vitro using dendritic cells derived from human monocytes. The immature moDCs primed with CpG-ODN loaded PLGA NP also effectively increased the ratios of Treg/Th2 and Th1/Th2. This finding supports the hypothesis that the co-encapsulated CpG-ODN facilitates the inhibition of Th2 immunity also in vivo. In conclusion, our findings substantiate the translational potential of PLGA nanoparticles in which T-cell epitopes containing peptides are encapsulated for future development of effective and safe strategies for early life CMA prevention (i.e., supplementing hydrolyzed formula milk for infants at risk). In addition, besides BLG-peptides also CpG-ODN motives, which may include bacterial DNA of selected bacterial strains, can be co-loaded in the PLGA NP to increase the efficacy of the formulations in oral tolerance induction. This strategy may be further developed as a modality to treat cow’s milk allergy for example as adjunct treatment for allergen specific oral immunotherapy

PAHs in the North Atlantic Ocean and the Arctic Ocean: Spatial Distribution and Water Mass Transport

In the Arctic Ocean, it is still unclear what role oceanic transport plays in the fate of semivolatile organic compounds. The strong-stratified Arctic Ocean undergoes complex inputs and outputs of polycyclic aromatic hydrocarbons (PAHs) from the neighboring oceans and continents. To better understand PAHs’ transport processes and their contribution to high-latitude oceans, surface seawater, and water column, samples were collected from the North Atlantic Ocean and the Arctic Ocean in 2012. The spatial distribution of dissolved PAHs (∑9PAH) in surface seawater showed an “Arctic Shelf \u3e Atlantic Ocean \u3e Arctic Basin” pattern, with a range of 0.3–10.2 ng L−1. Positive matrix factorization modeling results suggested that vehicle emissions and biomass combustion were the major PAHs sources in the surface seawater. According to principal component analysis, PAHs in different water masses showed unique profiles indicating their different origins. Carried by the Norwegian Atlantic Current (0–800 m) and East Greenland Current (0–300 m), PAH individuals’ net transport mass fluxes ranged from −4.4 ± 1.7 to 53 ± 39 tons year−1 to the Arctic Ocean. We suggested the limited contribution of ocean currents on PAHs’ delivery to the Arctic Ocean, but their role in modulating PAHs’ air–sea interactions and other biogeochemical processes needs further studies

Inhibition of cow’s milk allergy development in mice by oral delivery of β-lactoglobulin-derived peptides loaded PLGA nanoparticles is associated with systemic whey-specific immune silencing

Background: Two to four percentage of infants are affected by cow's milk allergy (CMA), which persists in 20% of cases. Intervention approaches using early oral exposure to cow's milk protein or hydrolysed cow's milk formula are being studied for CMA prevention. Yet, concerns regarding safety and/or efficacy remain to be tackled in particular for high-risk non-exclusively breastfed infants. Therefore, safe and effective strategies to improve early life oral tolerance induction may be considered. Objective: We aim to investigate the efficacy of CMA prevention using oral pre-exposure of two selected 18-AA β-lactoglobulin-derived peptides loaded poly (lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) in a whey-protein induced CMA murine model. Methods: The peptides were loaded in PLGA NPs via a double emulsion solvent evaporation technique. In vivo, 3-week-old female C3H/HeOuJ mice received 6 daily gavages with PBS, whey, Peptide-mix, a high- or low-dose Peptide-NPs or empty-NP plus Peptide-mix, prior to 5 weekly oral sensitizations with cholera toxin plus whey or PBS (sham). One week after the last sensitization, the challenge induced acute allergic skin response, anaphylactic shock score, allergen-specific serum immunoglobulins and ex vivo whey-stimulated cytokine release by splenocytes was measured. Results: Mice pre-treated with high-dose Peptide-NPs but not low-dose or empty-NP plus Peptide-mix, were protected from anaphylaxis and showed a significantly lower acute allergic skin response upon intradermal whey challenge compared to whey-sensitized mice. Compared with the Peptide-mix or empty-NP plus Peptide-mix pre-treatment, the high-dose Peptide-NPs-pre-treatment inhibited ex vivo whey-stimulated pro-inflammatory cytokine TNF-α release by splenocytes. Conclusion & Clinical relevance: Oral pre-exposure of mice to two β-lactoglobulin-derived peptides loaded PLGA NPs induced a dose-related partial prevention of CMA symptoms upon challenge to whole whey protein and silenced whey-specific systemic immune response. These findings encourage further development of the concept of peptide-loaded PLGA NPs for CMA prevention towards clinical application

Oral pretreatment with β-lactoglobulin derived peptide and CpG co-encapsulated in PLGA nanoparticles prior to sensitizations attenuates cow's milk allergy development in mice

Cow's milk allergy is a common food allergy among infants. Improved hygiene conditions and loss of microbial diversity are associated with increased risk of allergy development. The intestinal immune system is essential for oral tolerance induction. In this respect, bacterial CpG DNA is known to drive Th1 and regulatory T-cell (Treg) development via Toll-Like-Receptor 9 (TLR-9) signaling, skewing away from the allergic Th2 phenotype. We aimed to induce allergen specific tolerance via oral delivery of poly (lactic-co-glycolic acid) nanoparticles (NP) co-encapsulated with a selected β-lactoglobulin derived peptide (BLG-Pep) and TLR-9 ligand CpG oligodeoxynucleotide (CpG). In vivo, 3-4-week-old female C3H/HeOuJ mice housed in individually ventilated cages received 6-consecutive-daily gavages of either PBS, whey, BLG-Pep/NP, CpG/NP, a mixture of BLG-Pep/NP plus CpG/NP or co-encapsulated BLG-Pep+CpG/NP, before 5-weekly oral sensitizations with whey plus cholera toxin (CT) or only CT (sham) and were challenged with whey 5 days after the last sensitization. The co-encapsulated BLG-Pep+CpG/NP pretreatment, but not BLG-Pep/NP, CpG/NP or the mixture of BLG-Pep/NP plus CpG/NP, prevented the whey-induced allergic skin reactivity and prevented rise in serum BLG-specific IgE compared to whey-sensitized mice. Importantly, co-encapsulated BLG-Pep+CpG/NP pretreatment reduced dendritic cell (DC) activation and lowered the frequencies of PD-L1+ DC in the mesenteric lymph nodes compared to whey-sensitized mice. By contrast, co-encapsulated BLG-Pep+CpG/NP pretreatment increased the frequency of splenic PD-L1+ DC compared to the BLG-Pep/NP plus CpG/NP recipients, in association with lower Th2 development and increased Treg/Th2 and Th1/Th2 ratios in the spleen. Oral administration of PLGA NP co-encapsulated with BLG-Pep and CpG prevented rise in serum BLG-specific IgE and symptom development while lowering splenic Th2 cell frequency in these mice which were kept under strict hygienic conditions

Polyphenol-rich oolong tea alleviates obesity and modulates gut microbiota in high-fat diet-fed mice

Obesity is a major public health issue worldwide. Oolong tea (OT), which is partially fermented from Camellia sinensis leaves, has proven health benefits and potential preventive applications in multiple studies. However, research on the role of OT in obesity prevention and potential mechanisms is still limited. The purpose of this study was to investigate the modulatory effects of OT intervention on high-fat diet (HFD)-induced obesity and gut microbiota dysbiosis using an obese mouse model. Our results showed that 8-week OT supplementation with 93.94% polyphenols significantly decreased body weight gain, adipose tissue mass, and serum levels of triglyceride (2.60 mmol/L), cholesterol (5.49 mmol/L), and low-density lipoprotein cholesterol (0.61 mmol/L) in HFD-fed mice. Meanwhile, OT intervention was observed to improve fat accumulation, hepatic damage, glucose intolerance, and endotoxemia and alleviate inflammation by decreasing the levels of pro-inflammatory factors. OT also upregulated the expression of genes including Srebf1, Ppara, Lxra, Pgc1a, and Hsl and downregulated the expression of genes including Leptin, Il-6, and Il-1b. In addition, the gut dysbiosis characterized by decreased flora diversity and increased Firmicutes/Bacteroidetes ratio in obese mice was recovered by OT intervention. Certain differentially abundant microbes caused by HFD feeding, including Enterococcus, Intestinimonas, Blautia, and Bilophila, were also improved by OT treatment. This study demonstrated that OT, as a novel resource of dietary polyphenols, exhibited a protective effect on HFD-induced obesity and gut microbiota disorder

Live Cell Imaging by Förster Resonance Energy Transfer Fluorescence to Study Trafficking of PLGA Nanoparticles and the Release of a Loaded Peptide in Dendritic Cells

Our previous study demonstrated that a selected β-lactoglobulin-derived peptide (BLG-Pep) loaded in poly(lactic- co-glycolic acid) (PLGA) nanoparticles protected mice against cow's milk allergy development. However, the mechanism(s) responsible for the interaction of the peptide-loaded PLGA nanoparticles with dendritic cells (DCs) and their intracellular fate was/were elusive. Förster resonance energy transfer (FRET), a distance-dependent non-radioactive energy transfer process mediated from a donor to an acceptor fluorochrome, was used to investigate these processes. The ratio of the donor (Cyanine-3)-conjugated peptide and acceptor (Cyanine-5) labeled PLGA nanocarrier was fine-tuned for optimal (87%) FRET efficiency. The colloidal stability and FRET emission of prepared NPs were maintained upon 144 h incubation in PBS buffer and 6 h incubation in biorelevant simulated gastric fluid at 37 °C. A total of 73% of Pep-Cy3 NP was internalized by DCs as quantified using flow cytometry and confirmed using confocal fluorescence microscopy. By real-time monitoring of the change in the FRET signal of the internalized peptide-loaded nanoparticles, we observed prolonged retention (for 96 h) of the nanoparticles-encapsulated peptide as compared to 24 h retention of the free peptide in the DCs. The prolonged retention and intracellular antigen release of the BLG-Pep loaded in PLGA nanoparticles in murine DCs might facilitate antigen-specific tolerance induction

Oral pretreatment with β-lactoglobulin derived peptide and CpG co-encapsulated in PLGA nanoparticles prior to sensitizations attenuates cow’s milk allergy development in mice

Cow’s milk allergy is a common food allergy among infants. Improved hygiene conditions and loss of microbial diversity are associated with increased risk of allergy development. The intestinal immune system is essential for oral tolerance induction. In this respect, bacterial CpG DNA is known to drive Th1 and regulatory T-cell (Treg) development via Toll-Like-Receptor 9 (TLR-9) signaling, skewing away from the allergic Th2 phenotype. We aimed to induce allergen specific tolerance via oral delivery of poly (lactic-co-glycolic acid) nanoparticles (NP) co-encapsulated with a selected β-lactoglobulin derived peptide (BLG-Pep) and TLR-9 ligand CpG oligodeoxynucleotide (CpG). In vivo, 3-4-week-old female C3H/HeOuJ mice housed in individually ventilated cages received 6-consecutive-daily gavages of either PBS, whey, BLG-Pep/NP, CpG/NP, a mixture of BLG-Pep/NP plus CpG/NP or co-encapsulated BLG-Pep+CpG/NP, before 5-weekly oral sensitizations with whey plus cholera toxin (CT) or only CT (sham) and were challenged with whey 5 days after the last sensitization. The co-encapsulated BLG-Pep+CpG/NP pretreatment, but not BLG-Pep/NP, CpG/NP or the mixture of BLG-Pep/NP plus CpG/NP, prevented the whey-induced allergic skin reactivity and prevented rise in serum BLG-specific IgE compared to whey-sensitized mice. Importantly, co-encapsulated BLG-Pep+CpG/NP pretreatment reduced dendritic cell (DC) activation and lowered the frequencies of PD-L1+ DC in the mesenteric lymph nodes compared to whey-sensitized mice. By contrast, co-encapsulated BLG-Pep+CpG/NP pretreatment increased the frequency of splenic PD-L1+ DC compared to the BLG-Pep/NP plus CpG/NP recipients, in association with lower Th2 development and increased Treg/Th2 and Th1/Th2 ratios in the spleen. Oral administration of PLGA NP co-encapsulated with BLG-Pep and CpG prevented rise in serum BLG-specific IgE and symptom development while lowering splenic Th2 cell frequency in these mice which were kept under strict hygienic conditions

Spatio-Temporal Traffic Flow Prediction Based on Coordinated Attention

Traffic flow prediction can provide effective support for traffic management and control and plays an important role in the traffic system. Traffic flow has strong spatio-temporal characteristics, and existing traffic flow prediction models tend to extract long-term dependencies of traffic flow in the temporal and spatial dimensions individually, often ignoring the potential correlations existing between spatio-temporal information of traffic flow. In order to further improve the prediction accuracy, this paper proposes a coordinated attention-based spatio-temporal graph convolutional network (CVSTGCN) model for simultaneously and dynamically capturing the long-term dependencies existing between the spatio-temporal information of traffic flows. CVSTGCN is composed of a full convolutional network structure, which combines coordinate methods to specify the influence degrees of different feature information in different spatio-temporal dimensions, and the spatio-temporal information of different spatio-temporal dimensions by the graph convolutional network. In addition, the hard-swish activation function is introduced to replace the Rectified Linear Unit (ReLU) activation function in the prediction of traffic flow. Finally, evaluation experiments are conducted on two real datasets to demonstrate that the proposed model has the best prediction performance in both short-term and long-term forecasting

A Quantitative Structure-Property Relationship Model Based on Chaos-Enhanced Accelerated Particle Swarm Optimization Algorithm and Back Propagation Artificial Neural Network

A quantitative structure-property relationship (QSPR) model is proposed to explore the relationship between the pKa of various compounds and their structures. Through QSPR studies, the relationship between the structure and properties can be obtained. In this study, a novel chaos-enhanced accelerated particle swarm algorithm (CAPSO) is adopted to screen molecular descriptors and optimize the weights of back propagation artificial neural network (BP ANN). Then, the QSPR model based on CAPSO and BP ANN is proposed and named the CAPSO BP ANN model. The prediction experiment showed that the CAPSO algorithm was a reliable method for screening molecular descriptors. The five molecular descriptors obtained by the CAPSO algorithm could well characterize the molecular structure of each compound in pKa prediction. The experimental results also showed that the CAPSO BP ANN model exhibited good performance in predicting the pKa values of various compounds. The absolute mean relative error, root mean square error, and square correlation coefficient are respectively 0.5364, 0.0632, and 0.9438, indicating the high prediction accuracy. The proposed hybrid intelligent model can be applied in engineering design and the prediction of physical and chemical properties

DCCaps-UNet: A U-Shaped Hyperspectral Semantic Segmentation Model Based on the Depthwise Separable and Conditional Convolution Capsule Network

Traditional hyperspectral image semantic segmentation algorithms can not fully utilize the spatial information or realize efficient segmentation with less sample data. In order to solve the above problems, a U-shaped hyperspectral semantic segmentation model (DCCaps-UNet) based on the depthwise separable and conditional convolution capsule network was proposed in this study. The whole network is an encoding–decoding structure. In the encoding part, image features are firstly fully extracted and fused. In the decoding part, images are then reconstructed by upsampling. In the encoding part, a dilated convolutional capsule block is proposed to fully acquire spatial information and deep features and reduce the calculation cost of dynamic routes using a conditional sliding window. A depthwise separable block is constructed to replace the common convolution layer in the traditional capsule network and efficiently reduce network parameters. After principal component analysis (PCA) dimension reduction and patch preprocessing, the proposed model was experimentally tested with Indian Pines and Pavia University public hyperspectral image datasets. The obtained segmentation results of various ground objects were analyzed and compared with those obtained with other semantic segmentation models. The proposed model performed better than other semantic segmentation methods and achieved higher segmentation accuracy with the same samples. Dice coefficients reached 0.9989 and 0.9999. The OA value can reach 99.92% and 100%, respectively, thus, verifying the effectiveness of the proposed model