Ovalbumin promotes innate immune response of Caenorhabditis elegans through DAF-16 and SKN-1 pathways in insulin/IGF-1 signaling.

Ovalbumin (OVA) is a major allergen in eggs and could induce severe allergic reactions in sensitive individuals, where the innate immune system works as a regulator. The mechanism of how innate immunity adjusts to food allergy is relatively well-studied, however, the effects of allergen uptake on the innate immune system remain unclear. Therefore, the Caenorhabditis elegans (C. elegans) model was utilized to assess the effects of OVA on its innate immune system. OVA enhanced the immune response of C. elegans with higher survival rates under Pseudomonas aeruginosa infection. Moreover, sustaining OVA treatment improved the health states that were reflected in the prolonged lifespan, alleviated oxidative stress, accelerated growth, and promoted motility. RNA-sequencing analysis and the slow-killing assays in the mutants of insulin/IGF-1 signaling (IIS)-related genes confirmed that IIS was necessary for OVA to regulate innate immunity. Besides, OVA activated SKN-1 temporarily and facilitated the nuclear localization of DAF-16 for improving immunity and health status in C. elegans. Together, OVA could enhance the innate immune responses via DAF-16 and SKN-1 pathways in the IIS of C. elegans, and this work will provide novel insights into the regulation of innate immunity by OVA in higher organisms.

Diversity Patterns of Eukaryotic Phytoplankton in the Medog Section of the Yarlung Zangbo River.

As one of the important biodiversity conservation areas in China, the ecosystem in the lower reaches of the Yarlung Zangbo River is fragile, and is particularly sensitive to global changes. To reveal the diversity pattern of phytoplankton, the metabarcode sequencing was employed in the Medog section of the lower reaches of the Yarlung Zangbo River during autumn 2019 in present study. The phytoplankton assemblies can be significantly divided into the main stem and the tributaries; there are significant differences in the phytoplankton biomass, alpha and beta diversity between the main stem and the tributaries. While both the main stem and the tributaries are affected by dispersal limitation, the phytoplankton assemblages in the entire lower reaches are primarily influenced by heterogeneous selection. Community dissimilarity and assembly process were significantly correlated with turbidity, electrical conductivity, and nitrogen nutrition. The tributaries were the main source of the increase in phytoplankton diversity in the lower reaches of the Yarlung Zangbo River. Such diversity pattern of phytoplankton in the lower reach may be caused by the special habitat in Medog, that is, the excessive flow velocity, and the significant spatial heterogeneity in physical and chemical factors between stem and tributaries. Based on the results and conclusions obtained in present study, continuous long-term monitoring is essential to assess and quantify the impact of global changes on phytoplankton.

Assessment of sepsis-associated encephalopathy by quantitative magnetic resonance spectroscopy in a rat model of cecal ligation and puncture.

Proton magnetic resonance spectroscopy (1H-MRS) is the only non-invasive technique to quantify neurometabolic compounds in the living brain. We used 1H-MRS to evaluate the brain metabolites in a rat model of Sepsis-associated encephalopathy (SAE) established by cecal ligation and puncture (CLP). 36 male Sprague-Dawley rats were randomly divided into sham and CLP groups. Each group was further divided into three subgroups: subgroup O, subgroup M, and subgroup N. Neurological function assessments were performed on the animals in the subgroup O and subgroup N at 24 h, 48 h, and 72 h. The animals in the subgroup M were examined by magnetic resonance imaging (MRI) at 12 h after CLP. Compared with the sham group, the ratio of N-acetylaspartate (NAA) to creatine (Cr) in the hippocampus was significantly lower in the CLP group. The respective ratios of lactate (Lac), myo-inositol (mIns), glutamate and glutamine (Glx), lipid (Lip), and choline (Cho) to Cr in the CLP group were clearly higher than those in the sham group. Cytochrome c, intimately related to oxidative stress, was elevated in the CLP group. Neurofilament light (NfL) chain and glial fibrillary acidic protein (GFAP) scores in the CLP group were significantly higher than those in the sham group, while zonula occludens-1 (ZO-1) was downregulated. Compared with the sham group, the CLP group displayed higher values of oxygen extraction fraction (OEF), central venous-arterial partial pressure of carbon dioxide (P (cv-a) CO2), and central venous lactate (VLac). In contrast, jugular venous oxygen saturation (SjvO2) declined. In the present study, 1H-MRS could be used to quantitatively assess brain injury in terms of microcirculation disorder, oxidative stress, blood-brain barrier disruption, and glial cell activation through changes in metabolites within brain tissue.

Nanosystem Delivers Senescence Activators and Immunomodulators to Combat Liver Cancer.

CD47 blockade has emerged as a promising immunotherapy against liver cancer. However, the optimization of its antitumor effectiveness using efficient drug delivery systems or combinations of therapeutic agents remains largely incomplete. Here, patients with liver cancer co-expressing CD47 and CDC7 (cell division cycle 7, a negative senescence-related gene) are found to have the worst prognosis. Moreover, CD47 is highly expressed, and senescence is inhibited after the development of chemoresistance, suggesting that combination therapy targeting CD47 and CDC7 to inhibit CD47 and induce senescence may be a promising strategy for liver cancer. The efficacy of intravenously administered CDC7 and CD47 inhibitors is limited by low uptake and short circulation times. Here, inhibitors are coloaded into a dual-targeted nanosystem. The sequential release of the inhibitors from the nanosystem under acidic conditions first induces cellular senescence and then promotes immune responses. In an in situ liver cancer mouse model and a chemotherapy-resistant mouse model, the nanosystem effectively inhibited tumor growth by 90.33% and 85.15%, respectively. Overall, the nanosystem in this work achieved the sequential release of CDC7 and CD47 inhibitors in situ to trigger senescence and induce immunotherapy, effectively combating liver cancer and overcoming chemoresistance.

Comparative genomic analysis of the Growth-Regulating Factors-Interacting Factors (GIFs) in six Salicaceae species and functional analysis of PeGIF3 reveals their regulatory role in Populus heteromorphic leaves.

BACKGROUND: The growth-regulating factor-interacting factor (GIF) gene family plays a vital role in regulating plant growth and development, particularly in controlling leaf, seed, and root meristem homeostasis. However, the regulatory mechanism of heteromorphic leaves by GIF genes in Populus euphratica as an important adaptative trait of heteromorphic leaves in response to desert environment remains unknown. RESULTS: This study aimed to identify and characterize the GIF genes in P. euphratica and other five Salicaceae species to investigate their role in regulating heteromorphic leaf development. A total of 27 GIF genes were identified and characterized across six Salicaceae species (P. euphratica, Populus pruinose, Populus deltoides, Populus trichocarpa, Salix sinopurpurea, and Salix suchowensis) at the genome-wide level. Comparative genomic analysis among these species suggested that the expansion of GIFs may be derived from the specific Salicaceae whole-genome duplication event after their divergence from Arabidopsis thaliana. Furthermore, the expression data of PeGIFs in heteromorphic leaves, combined with functional information on GIF genes in Arabidopsis, indicated the role of PeGIFs in regulating the leaf development of P. euphratica, especially PeGIFs containing several cis-acting elements associated with plant growth and development. By heterologous expression of the PeGIF3 gene in wild-type plants (Col-0) and atgif1 mutant of A. thaliana, a significant difference in leaf expansion along the medial-lateral axis, and an increased number of leaf cells, were observed between the overexpressed plants and the wild type. CONCLUSION: PeGIF3 enhances leaf cell proliferation, thereby resulting in the expansion of the central-lateral region of the leaf. The findings not only provide global insights into the evolutionary features of Salicaceae GIFs but also reveal the regulatory mechanism of PeGIF3 in heteromorphic leaves of P. euphratica.

Anthropogenic impacts and quantitative sources of nitrate in a rural-urban canal using a combined PMF, δ15N/δ18O-NO3-, and MixSIAR approach.

Nitrate (NO3-) pollution in irrigation canals is of great concern because it threatens canal water use; however, little is known about it at present. Herein, a combination of positive matrix factorization (PMF), isotope tracers, and Mixing Stable Isotope Analysis in R (MixSIAR) was developed to identify anthropogenic impacts and quantitative sources of NO3- in a rural-urban canal in China. The NO3- concentration (0.99-1.93 mg/L) of canal water increased along the flow direction and was higher than the internationally recognized eutrophication risk value in autumn and spring. The inputs of the Fuhe River, NH4+ fertilizer, soil nitrogen, manure & sewage, and rainfall were the main driving factors of canal water NO3- based on principal component analysis and PMF, which was supported by evidence from δ15N/δ18O-NO3-. According to the chemical and isotopic analyses, nitrogen transformation was weak, highlighting the potential of δ15N/δ18O-NO3- to trace NO3- sources in canal water. The MixSIAR and PMF results with a <15% divergence emphasized the predominance of the Fuhe River (contributing >50%) and anthropogenic impacts (NH4+ fertilizer plus manure & sewage, >37%) on NO3- in the entire canal, reflecting the effectiveness of the model analysis. According to the MixSIAR model, (1) higher NO3- concentration in canal water was caused by the general enhancement of human activities in spring and (2) NO3- source contributions were associated with land-use patterns. The high contributions of NH4+ fertilizer and manure & sewage showed inverse spatial variations, suggesting the necessity of reducing excessive fertilizer use in the agricultural area and controlling blind wastewater release in the urban area. These findings provide valuable insights into NO3- dynamics and fate for sustainable management of canal water resources. Nevertheless, long-term chemical and isotopic monitoring with alternative modeling should be strengthened for the accurate evaluation of canal NO3- pollution in future studies.

The chromosome-scale genome and population genomics reveal the adaptative evolution of Populus pruinosa to desertification environment.

The Populus pruinosa is a relic plant that has managed to survive in extremely harsh desert environments. Owing to intensifying global warming and desertification, research into ecological adaptation and speciation of P. pruinosa has attracted considerable interest, but the lack of a chromosome-scale genome has limited adaptive evolution research. Here, a 521.09 Mb chromosome-level reference genome of P. pruinosa was reported. Genome evolution and comparative genomic analysis revealed that tandemly duplicated genes and expanded gene families in P. pruinosa contributed to adaptability to extreme desert environments (especially high salinity and drought). The long terminal repeat retrotransposons (LTR-RTs) inserted genes in the gene body region might drive the adaptive evolution of P. pruinosa and species differentiation in saline-alkali desert environments. We recovered genetic differentiation in the populations of the northern Tianshan Mountain and southern Tianshan Mountain through whole-genome resequencing of 156 P. pruinosa individuals from 25 populations in China. Further analyses revealed that precipitation drove the local adaptation of P. pruinosa populations via some genetic sites, such as MAG2-interacting protein 2 (MIP2) and SET domain protein 25 (SDG25). This study will provide broad implications for adaptative evolution and population studies by integrating internal genetic and external environmental factors in P. pruinosa.

The structure and potential allergenicity of peanut allergen monomers after roasting.

Given that roasted peanut (Ro) products are commonly used in daily life, peanut allergenicity is a foremost concern. Analyzing the changes in the structure and potential allergenicity of individual allergens can promote the exploration of the structural basis of the alterations in the potential allergenicity of Ro. This work focused on four major allergens in raw peanut (Ra) and Ro. Structural changes were analyzed on the basis of circular dichroism, ultraviolet and fluorescence spectroscopy, and molecular dynamic simulation. The IgE recognition capability of allergens was assessed via western blot analysis. The IgE binding capacity of allergens was detected by conducting enzyme-linked immunosorbent assay. The potential allergenicity of allergens was evaluated using the KU812 cell degranulation model. The results showed that roasting induced different changes in the overall structures of allergens and altered the structures and electrostatic potential of IgE epitopes, especially Ara h 1 and Ara h 6. These alterations affected the potential allergenicity of allergens. Ara h 1 and Ara h 6 in Ro showed significantly enhanced IgE binding capacities and abilities to elicit KU812 cell degranulation, while Ara h 2 and Ara h 3 did not change significantly. For total protein, the roasted peanut protein showed decreased abilities to elicit KU812 cell degranulation. The results indicated that different allergens in Ro showed different changes of structures and potential allergenicity and that the conformational structure plays a crucial role in potential allergenicity of allergens.

Mass Spectrometry Analysis on the Breakage of Allergens in High-Molecular-Mass Polymer of Roasted Peanuts.

Peanut allergy is a prevalent and concerning food allergy. Roasting can introduce structural changes to peanut allergens, affecting their allergenicity, but the structure on the primary structure is unclear. Here, the breakage sites were identified by mass spectrometry and software tools, and structural changes were simulated by molecular dynamics and displayed by PyMOL software. Results revealed that the appearance frequencies of L, Q, F, and E were high at the N-terminal of the breakage site, while S and E were dominant at the C-terminal. In the conformational structure, breakage sites were found close to disulfide bonds and the Cupin domains of Ara h 1 and Ara h 3. The breakage of allergens destroyed linear epitopes and might change the conformation of epitopes, which could influence peanuts' potential allergenicity.

The flavor substances changes in Fuliang green tea during storage monitoring by GC-MS and GC-IMS.

To study the effect of storage (for 0, 3, 6, and 12 months) on the flavor of green tea (GT), we monitored the volatile organic compounds (VOCs) in GT through gas chromatography (GC) combined with ion mobility spectrometry and headspace solid-phase micro extraction, GC-MS (mass spectrometry). Then, relative odor activity value (ROAV) was applied to analyze the aroma contribution of the VOCs. During storage, the polyphenol and caffeine contents gradually decreased from 22.38 % to 18.51 % and from 4.37 % to 3.74 %, respectively, and the total soluble sugar first increased and then decreased (from 4.89 % to 7.16 % and then 5.02 %). Although the total free amino acid contents showed a fluctuating trend, the content of cysteamine increased gradually. The contents of VOCs with positive contribution to GT aroma, including linalool, geraniol, nonanal, and 6-methyl-5-hepten-2-one, decreased. They also contributed less in the ROAV after storage. The ROAVs of nonanal, linalool, and geraniol decreased from 3.37 to 0.79, from 100 to 38.21, and from 2.98 to 1.8, respectively, after 12 months of storage. Principal component analysis can be used to identify the samples with different storage durations based on these data. Given the increase in amount of cysteamine and decrease in that of linalool oxide, oxidation may be not the only factor responsible for tea quality in storage.

Investigation of peanut allergen-procyanidin non-covalent interactions: Impact on protein structure and in vitro allergenicity.

Interactions between plant polyphenols and food allergens may be a new way to alleviate food allergies. The non-covalent interactions between the major allergen from peanut (Ara h 2) with procyanidin dimer (PA2) were therefore characterized using spectroscopic, thermodynamic, and molecular simulation analyses. The main interaction between the Ara h 2 and PA2 was hydrogen bonding. PA2 statically quenched the intrinsic fluorescence intensity and altered the conformation of the Ara h 2, leading to a more disordered polypeptide structure with a lower surface hydrophobicity. In addition, the in vitro allergenicity of the Ara h 2-PA2 complex was investigated using enzyme-linked immunosorbent assay (ELISA) kits. The immunoglobulin E (IgE) binding capacity of Ara h 2, as well as the release of allergenic cytokines, decreased after interacting with PA2. When the ratio of Ara h 2-to-PA2 was 1:50, the IgE binding capacity was reduced by around 43 %. This study provides valuable insights into the non-covalent interactions between Ara h 2 and PA2, as well as the potential mechanism of action of the anti-allergic reaction caused by binding of the polyphenols to the allergens.

Shedding light on the interaction of ovalbumin and resveratrol: structure, digestibility, transport, and allergenicity assessment of OVA-RES complexes.

BACKGROUND: The interaction between food allergens and plant polyphenols has become a safe and effective management strategy to prevent food allergies. Ovalbumin (OVA) is the most abundant allergen in egg whites. Resveratrol (RES) is a plant polyphenol that is abundant in red grapes, berries, and peanuts, and has an anti-allergic effect on allergy-related immune cells. However, there is little information about the effect of RES on the allergenicity of OVA. In this study, the effect of RES on the allergenicity of OVA was investigated. RESULTS: Molecular docking and spectroscopic studies indicated that the addition of RES changed the structure of OVA. The digestion and transfer rate of OVA-RES were effectively improved with an in vitro gastrointestinal digestion model and Caco-2 cell model, especially when the molar ratio of OVA-RES was 1:20. Meanwhile, the KU812 cell degranulation assay proved that the potential allergenicity was remarkably decreased while the molar ratios of OVA-RES were increased to 1:20. Furthermore, hydrogen bonds and van der Waals forces were the dominating forces to stabilize the OVA-RES complexes. CONCLUSION: All the findings demonstrated that the potential allergenicity of OVA was reduced when interacting with RES, and RES can be a potential food material for preparing a hypoallergenic protein, especially for egg allergy. © 2023 Society of Chemical Industry.

Active polysaccharides: a new roadmap for the prevention and treatment of food allergy.

Active polysaccharides are extensively utilized in the fields of food and medicine because of their rich functional properties and structural plasticity. However, there are still few systematic studies and reviews on active polysaccharides for allergy. Allergy, especially food allergy, occurs frequently around the world and is related to a variety of factors such as age, genetics and dietary habits. Currently in medicine, avoiding allergens and desensitizing can effectively relieve allergy symptoms, but these are difficult to maintain over the long term and come with risks. Based on the supplementation of dietary nutrition to these two treatments, it has been discovered in recent years that the use of active ingredients from natural substances can effectively intervene in allergies. Considering the potential of active polysaccharides in this regard, we systematically characterize the latent patterns of polysaccharides in allergic symptoms and pathogenesis, including the aspects of gut, immunomodulatory, oxidative stress and signaling pathways, as well as the application prospect of them in allergy. It can be found that active polysaccharides have excellent anti-allergic potential, especially from the ocean. We believe that the active polysaccharides are associated with the treatment of allergic diseases, which may provide the benefits to allergy sufferers in the future.

Genes expression profiles in vascular cambium of Eucalyptus urophylla × Eucalyptus grandis at different ages.

BACKGROUND: Wood is a secondary xylem generated by vascular cambium. Vascular cambium activities mainly include cambium proliferation and vascular tissue formation through secondary growth, thereby producing new secondary phloem inward and secondary xylem outward and leading to continuous tree thickening and wood formation. Wood formation is a complex biological process, which is strictly regulated by multiple genes. Therefore, molecular level research on the vascular cambium of different tree ages can lead to the identification of both key and related genes involved in wood formation and further explain the molecular regulation mechanism of wood formation. RESULTS: In the present study, RNA-Seq and Pac-Bio Iso-Seq were used for profiling gene expression changes in Eucalyptus urophylla × Eucalyptus grandis (E. urograndis) vascular cambium at four different ages. A total of 59,770 non-redundant transcripts and 1892 differentially expressed genes (DEGs) were identified. The expression trends of the DEGs related to cell division and differentiation, cell wall biosynthesis, phytohormone, and transcription factors were analyzed. The DEGs encoding expansin, kinesin, cycline, PAL, GRP9, KNOX, C2C2-dof, REV, etc., were highly expressed in E. urograndis at three years old, leading to positive effects on growth and development. Moreover, some gene family members, such as NAC, MYB, HD-ZIP III, RPK, and RAP, play different regulatory roles in wood formation because of their sophisticated transcriptional network and function redundantly. CONCLUSIONS: These candidate genes are a potential resource to further study wood formation, especially in fast-growing and adaptable eucalyptus. The results may also serve as a basis for further research to unravel the molecular mechanism underlying wood formation.

Stress-induced endocytosis from chloroplast inner envelope membrane is mediated by CHLOROPLAST VESICULATION but inhibited by GAPC.

Clathrin-mediated vesicular formation and trafficking are responsible for molecular cargo transport and signal transduction among organelles. Our previous study shows that CHLOROPLAST VESICULATION (CV)-containing vesicles (CVVs) are generated from chloroplasts for chloroplast degradation under abiotic stress. Here, we show that CV interacts with the clathrin heavy chain (CHC) and induces vesicle budding toward the cytosol from the chloroplast inner envelope membrane. In the defective mutants of CHC2 and the dynamin-encoding DRP1A, CVV budding and releasing from chloroplast are impeded. The mutations of CHC2 inhibit CV-induced chloroplast degradation and hypersensitivity to water stress. Moreover, CV-CHC2 interaction is impaired by the oxidized GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPC). GAPC1 overexpression suppresses CV-mediated chloroplast degradation and hypersensitivity to water stress, while CV silencing alleviates the hypersensitivity of the gapc1gapc2 plant to water stress. Together, our work identifies a pathway of clathrin-assisted CVV budding outward from chloroplast, which is involved in chloroplast degradation and stress response.

A Comprehensive Analysis of Chloroplast Genome Provides New Insights into the Evolution of the Genus Chrysosplenium.

Chrysosplenium, a perennial herb in the family Saxifragaceae, prefers to grow in low light and moist environments and is divided into two sections of Alternifolia and Oppositifolia based on phyllotaxy. Although there has been some progress in the phylogeny of Chrysosplenium over the years, the phylogenetic position of some species is still controversial. In this study, we assembled chloroplast genomes (cp genomes) of 34 Chrysosplenium species and performed comparative genomic and phylogenetic analyses in combination with other cp genomes of previously known Chrysosplenium species, for a total of 44 Chrysosplenium species. The comparative analyses revealed that cp genomes of Chrysosplenium species were more conserved in terms of genome structure, gene content and arrangement, SSRs, and codon preference, but differ in genome size and SC/IR boundaries. Phylogenetic analysis showed that cp genomes effectively improved the phylogenetic support and resolution of Chrysosplenium species and strongly supported Chrysosplenium species as a monophyletic taxon and divided into three branches. The results also showed that the sections of Alternifolia and Oppositifolia were not monophyletic with each other, and that C. microspermum was not clustered with other Chrysosplenium species with alternate leaves, but with C. sedakowii into separate branches. In addition, we identified 10 mutational hotspot regions that could serve as potential DNA barcodes for Chrysosplenium species identification. In contrast to Peltoboykinia, the clpP and ycf2 genes of Chrysosplenium were subjected to positive selection and had multiple significant positive selection sites. We further detected a significant positive selection site on the petG gene between the two sections of Chrysosplenium. These evolutionary characteristics may be related to the growth environment of Chrysosplenium species. This study enriches the cp genomes of Chrysosplenium species and provides a reference for future studies on its evolution and origin.

Single-cell RNA-seq reveals a microenvironment and an exhaustion state of T/NK cells in acute myeloid leukemia.

Acute myeloid leukemia (AML) is a heterogeneous blood cancer. Effective immunotherapies for AML are hindered by a lack of understanding of the tumor microenvironment (TME). Here, we retrieved published single-cell RNA sequencing data for 128,688 cells derived from 29 bone marrow aspirates, including 21 AML patients and eight healthy donors. We established a global tumor ecosystem including nine main cell types. Myeloid, T, and NK cells were further re-clustered and annotated. Developmental trajectory analysis indicated that exhausted CD8+ T cells might develop via tissue residual memory T cells (TRM) in the AML TME. Significantly higher expression levels of exhaustion molecules in AML TRM cells suggested that these cells were influenced by the TME and entered an exhausted state. Meanwhile, the upregulation of checkpoint molecules and downregulation of granzyme were also observed in AML NK cells, suggesting an exhaustion state. In conclusion, our comprehensive profiling of T/NK subpopulations provides deeper insights into the AML immunosuppressive ecosystem, which is critical for immunotherapies.

Pollution source identification of nitrogen and phosphorus in the lower West Main Canal, the Ganfu Plain irrigation district (South China).

The degradation of surface water quality has been a widespread concern around the world. However, irrigation canal water does not attract much attention although it is important to agriculture and population. In this study, a 5-year water quality monitoring of surface water was conducted in the lower West Main Canal of the Ganfu Plain irrigation district to identify the levels and pollution sources of nitrogen and phosphorus.Over 75% of samples had total phosphorus (TP) concentrations of > 0.02 mg/L, and all samples had total nitrogen (TN) concentrations of > 0.2 mg/L, indicating a risk of eutrophication. The concentrations of NO3--N and NH4+-N averagely occupied 57% and 18% of TN, respectively. PCA analysis showed that phosphorus and nitrogen in canal water were associated with meteorological factors, urban life and surface runoff, agricultural cultivation, livestock-poultry breeding, and water-sediment interaction in the wet season, whereas they were affected by meteorological factors, industrial effluent, urban domestic sewage, and livestock-poultry breeding in the dry season. Absolute principal component score-multiple linear regression (APCS-MLR) model results revealed that (1) agricultural cultivation plus livestock-poultry breeding contributed 43.2% of TP in canal water in the wet season, while livestock-poultry breeding contributed 52.9% in the dry season, and (2) domestic sewage plus surface runoff contributed 29.4% of TN in the wet season, while livestock-poultry breeding contributed 45.9% in the dry season. The unidentified sources had significant contributions of > 20% for almost all variables. So further investigations are required for determining unidentified sources, and anthropogenic pollution control is imperative for canal water quality protection.

Development of a Two-Step Hydrolysis Hypoallergenic Cow's Milk Formula and Evaluation of Residue Allergenicity by Peptidomics and Immunoreactivity Analysis.

Cow's milk allergy (CMA) is an abnormal immune response that severely affects the nutritional supplementation of allergic infants. Currently, only a limited number of hypoallergenic formulas are available on the market, and these are only categorized according to their degree of hydrolysis, which still poses an allergy risk and cannot be consumed by CMA patients, especially infants. To address this issue, we developed a two-step hydrolysis hypoallergenic formula targeting destruction of allergen epitope from whey protein. Then, a comprehensive evaluation system was constructed, including peptidomics analysis, in vivo and in vitro allergenicity assessments, revealing allergic changes in the product from the epitope structure level to the immunological level. The results showed that 97.14% of hydrolyzed peptides from α-lactalbumin and ß-lactoglobulin did not contain allergenic epitopes after treatment with trypsin and flavourzyme. In vitro and in vivo allergenicity assessment results confirmed that the two-step hydrolysis method effectively reduced the allergenicity of whey protein. Compared with the common milk powder, the hypoallergenic formula induced lower levels of basophil degranulation and relieved the body's anaphylactic symptoms caused by cow milk. This study provides a promising solution to the limited hypoallergenic formula problem and may benefit allergic infants who require nutritional supplements.

Dissolving microneedles-based programmed delivery system for enhanced chemo-immunotherapy of melanoma.

Immune checkpoint blockade, especially the programmed cell death ligand 1 (PD-L1) blockade, has revolutionized the treatment of melanoma. However, PD-1/PD-L1 monotherapy leads to unsatisfactory therapeutic outcomes. The immunotherapy of melanoma could be improved by adding doxorubicin (DOX), which triggers immunogenic cell death (ICD) effect to activate anti-tumor immunity. Additionally, microneedles, especially dissolving microneedles (dMNs), can further enhance outcomes of chemo-immunotherapy due to the physical adjuvant effect of dMNs. Herein, we developed the dMNs-based programmed delivery system that incorporated pH-sensitive and melanoma-targeting liposomes to co-deliver DOX and siPD-L1, achieving enhanced chemo-immunotherapy of melanoma (si/DOX@LRGD dMNs). The incorporated si/DOX@LRGD LPs demonstrated uniform particle size, pH-sensitive drug release, high in vitro cytotoxicity and targeting ability. Besides, si/DOX@LRGD LPs effectively downregulated the expression of PD-L1, induced tumor cell apoptosis and triggered ICD effect. The si/DOX@LRGD LPs also showed deep penetration (approximately 80 µm) in 3D tumor spheroids. Moreover, si/DOX@LRGD dMNs dissolved rapidly into the skin and had sufficient mechanical strength to penetrate skin, reaching a depth of approximately 260 µm in mice skin. In mice model of melanoma tumor, si/DOX@LRGD dMNs exhibited better anti-tumor efficacy than monotherapy by dMNs and tail intravenous injection at the same dose. This was due to the higher cytotoxic CD8+ T cells and the secreted cytotoxic cytokine IFN-γ evoked by si/DOX@LRGD dMNs, thereby eliciting strong T-cell mediated immune response and resulted in enhanced anti-tumor effects. In conclusion, these findings suggested that si/DOX@LRGD dMNs provided a promising and effective strategy for enhanced chemo-immunotherapy of melanoma.