Microencapsulation of fish oil by spray drying, spray freeze-drying, freeze-drying, and microwave freeze-drying: Microcapsule characterization and storage stability.

The objective of this paper was to evaluate the effect of spray drying (SD), spray freeze-drying (SFD), freeze-drying (FD), and microwave freeze-drying (MFD) on the characteristics of fish oil (FO) microcapsules. The physicochemical properties, morphology, fatty acid composition, and stability of the microcapsules were analyzed. The encapsulation efficiencies of microcapsules dried by SD, SFD, FD, and MFD were 86.98%, 77.79%, 63.29%, and 57.89%, respectively. SD microcapsules exhibited superior properties in terms of effective loading capacity, color, and flowability. Conversely, SFD microcapsules demonstrated improved solubility. Microencapsulation positively affected the thermal stability of FO, but the content of unsaturated fatty acids decreased. The findings from the storage experiment indicated that the oxidative stability of SD fish oil microcapsules was marginally lower compared to microcapsules produced through three alternative drying techniques, all of which were based on the FD concept. The comparison of various drying methods and their effects on the quality of FO microcapsules offers valuable insights that can serve as a foundation for the industrial production of high-quality microcapsules.

A circular single-stranded DNA mycovirus infects plants and confers broad-spectrum fungal resistance.

Circular single-stranded DNA (ssDNA) viruses have been rarely found in fungi, and the evolutionary and ecological relationships among ssDNA viruses infecting fungi and other organisms remain unclear. In this study, a novel circular ssDNA virus, tentatively named Diaporthe sojae circular DNA virus 1 (DsCDV1), was identified in the phytopathogenic fungus Diaporthe sojae isolated from pear trees. DsCDV1 has a monopartite genome (3185 nt in size) encapsidated in isometric virions (21-26 nm in diameter). The genome comprises seven putative open reading frames encoding a discrete replicase (Rep) split by an intergenic region, a putative capsid protein (CP), several proteins of unknown function (P1-P4), and a long intergenic region. Notably, the two split parts of DsCDV1 Rep share high identities with the Reps of Geminiviridae and Genomoviridae, respectively, indicating an evolutionary linkage with both families. Phylogenetic analysis based on Rep or CP sequences placed DsCDV1 in a unique cluster, supporting the establishment of a new family, tentatively named Gegemycoviridae, intermediate to both families. DsCDV1 significantly attenuates fungal growth and nearly erases fungal virulence when transfected into the host fungus. Remarkably, DsCDV1 can systematically infect tobacco and pear seedlings, providing broad-spectrum resistance to fungal diseases. Subcellular localization analysis revealed that DsCDV1 P3 is systematically localized in the plasmodesmata, while its expression in trans-complementation experiments could restore systematic infection of a movement-deficient plant virus, suggesting that P3 is a movement protein. DsCDV1 exhibits unique molecular and biological traits not observed in other ssDNA viruses, serving as a link between fungal and plant ssDNA viruses and presenting an evolutionary connection between ssDNA viruses and fungi. These findings contribute to expanding our understanding of ssDNA virus diversity and evolution, offering potential biocontrol applications for managing crucial plant diseases.

Induced Mesenchymal Stem Cells-Small Extracellular Vesicles Alleviate Post-stroke Cognitive Impairment by Rejuvenating Senescence of Neural Stem Cells.

Ischemic stroke is typified by hypoxia and a cascade of pathophysiological events, including metabolic dysfunction, ionic dysregulation, excitotoxicity, inflammatory infiltration, and oxidative stress. These ultimately result in neuronal apoptosis or necrosis with constrained neuroregenerative capabilities. In this study, neural stem cells (NSCs) under conditions of oxygen-glucose deprivation (OGD) in vitro and following middle cerebral artery occlusion (MCAO) in vivo were explored. Transcriptome sequencing revealed a decline in NSC differentiation and neurogenesis after OGD exposure, which was related to cellular senescence. This observation was corroborated by increased senescence markers in the MCAO mouse model, reduction in NSC numbers, and decline in neurogenesis. Importantly, iMSC-sEVs (induced mesenchymal stem cells-small extracellular vesicles) have the therapeutic potential to alleviate NSC senescence and rejuvenate their regenerative capacities both in vitro and in vivo. Moreover, iMSC-sEVs contribute to the recovery of cognitive function and synapse loss caused by MCAO.

The roles of templates consisting of amino acids in the synthesis and application of gold nanoclusters.

Gold nanoclusters (AuNCs) with low toxicity, high photostability, and facile synthesis have attracted great attention. The ligand is of great significance in stabilizing AuNCs and regulating their properties. Ligands consisting of amino acids (proteins and peptides) are an ideal template for synthesizing applicative AuNCs due to their inherent bioactivity, biocompatibility, and accessibility. In this review, we summarize the correlation of the template consisting of amino acids with the properties of AuNCs by analyzing different peptide sequences. The selection of amino acids can regulate the fluorescence excitation/emission and intensity, size, cell uptake, and light absorption. By analyzing the role played by AuNCs stabilized by proteins and peptides in the application, universal rules and detailed performances of sensors, antibacterial agents, therapeutic reagents, and light absorbers are reviewed. This review can guide the template design and application of AuNCs when selecting proteins and peptides as ligands.

Cost-effectiveness analysis of continuing bevacizumab plus chemotherapy versus chemotherapy alone after first progression of metastatic colorectal cancer.

BACKGROUND: Continuation of bevacizumab plus second-line chemotherapy has significantly improved overall and progression-free survival in patients with metastatic colorectal cancer (mCRC). However, the cost-effectiveness of such high cost therapy is still uncertain in China; so this analysis was performed to evaluate the cost-effectiveness of these treatment options from the Chinese health care system perspective. METHODS: A cost-effectiveness analysis was conducted using data from the ML18147 trial (ClinicalTrials.gov identifier NCT00700102) by modeling a partitioned survival model. Main evaluation indicators were quality-adjusted life years (QALYs) and incremental cost-effectiveness ratio (ICER) with a willingness to pay (WTP) threshold of $38,201 per QALY. One-way and probabilistic sensitivity analyses were conducted to assess the robustness and stability of the model. Subgroup and scenario analyses were also performed to make our study more relevant. RESULTS: Bevacizumab plus chemotherapy increased 0.12 QALYs and an incremental cost of $22,761.62 compared with chemotherapy, resulting in an ICER of $188,904.09 per QALY. The model was most sensitive to the utility of progression-free survival and the cost of bevacizumab. Compared with chemotherapy, bevacizumab plus chemotherapy had a 0% cost-effectiveness probability, and no cost-effectiveness in subgroups at the WTP threshold of $38,201 per QALY. The scenario analysis found that bevacizumab biosimilar gained an ICER of $126,397.38 per QALY when assuming the cost of drugs was calculated at the most affordable price. CONCLUSIONS: At the WTP threshold of $38,201 per QALY, continuation of bevacizumab plus chemotherapy is unlikely considered cost-effective for patients after first progression of mCRC.

A transcriptomics-based investigation of the mechanism of pulmonary fibrosis induced by nickel oxide nanoparticles.

Nickel oxide nanoparticles (NiONPs) are an emerging nanomaterial, which poses a huge threat to the health of workplace population. Nanoparticles induce pulmonary fibrosis, and its mechanisms are associated with noncoding RNAs (ncRNAs). However, ncRNAs and competing endogenous RNA (ceRNA) networks which involved in NiONP-induced pulmonary fibrosis are still unclear. This study aimed to identify ncRNA-related ceRNA networks and investigate the role of the Wnt/ß-catenin pathway in pulmonary fibrosis. Male Wistar rats were intratracheally instilled with 0.015, 0.06, and 0.24 mg/kg NiONPs twice a week for 9 weeks. First, we found there were 93 circularRNAs (circRNAs), 74 microRNAs (miRNAs), 124 long non-coding RNAs (lncRNAs), and 1675 messenger RNAs (mRNAs) differentially expressed through microarray analysis. Second, we constructed ceRNA networks among lncRNAs/circRNAs, miRNAs and mRNAs and identified two ceRNA networks (lncMelttl16/miR-382-5p/Hsd17b7 and circIqch/miR-181d-5p/Stat1) after real time-quantitative polymerase chain reaction (RT-qPCR) validation. Furthermore, based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses, ncRNAs were found to be involved in biological processes and signaling pathways related to pulmonary fibrosis. KEGG analysis showed that NiONPs activated the Wnt/ß-catenin pathway in rats. In vitro, HFL1 cells were treated with 0, 50, 100, and 200 µg/mL NiONPs for 24 h. We found that NiONPs induced collagen deposition and Wnt/ß-catenin pathway activation. Moreover, a blockade of Wnt/ß-catenin pathway alleviated NiONP-induced collagen deposition. In conclusion, these observations suggested that ncRNAs were crucial in pulmonary fibrosis development and that the Wnt/ß-catenin pathway mediated the deposition of collagen.

Small extracellular vesicles from irradiated lung epithelial cells promote the activation of fibroblasts in pulmonary fibrosis.

BACKGROUND: Alveolar epithelial injury and dysfunction are the risk factors for radiation-induced pulmonary fibrosis (RIPF). However, it is not clear about the relationship between RIPF and the small extracellular vesicles (sEV) secreted by irradiated alveolar epithelial cells. Based on the activation of fibroblasts, this study explored the role of sEV derived from alveolar epithelial cells in RIPF and the potential mechanisms. METHODS: Transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blotting were used to characterize sEV. Western blotting was used to detect fibrosis-associated proteins. Cell counts and transwell assays were used to evaluate the proliferation and migration ability of fibroblasts. RT-PCR was used to observe the extracellular matrix (ECM) synthesized by fibroblasts, miRNA changes in the sEV were determined by second-generation sequencing. RESULTS: TEM, NTA, and western blotting showed the extracellular vesicles with a double-layer membrane structure of approximately 100 nm in diameter. The sEV derived from irradiated A549, HBEC3-KT, and MLE12 cells upregulated FN1 and alpha-SMA proteins expression in fibroblasts and drove the fibroblast to myofibroblast transition, and the sEV from irradiated mouse bronchoalveolar lavage fluid (BALF) affirmed the same results. In addition, the sEV derived from irradiated alveolar epithelial cells significantly increased the migration ability of fibroblasts and the expression of extracellular matrix proteins such as FN1. The results of miRNA sequencing of sEV in BALF of rats with RIPF showed that the metabolic pathway may be important for miRNA to regulate the activation of fibroblasts. CONCLUSION: The sEV derived from radiated pulmonary epithelial cells promote the activation, migration and extracellular matrix proteins expression of lung fibroblasts; miRNA in sEV may be an important molecular that affects the activation of lung fibroblasts.

Hirsutella sinensis mycelium polysaccharides attenuate the TGF-ß1-induced epithelial-mesenchymal transition in human intrahepatic bile duct epithelial cells.

Hirsutella sinensis is the anamorph of Ophiocordyceps sinensis, and its mycelia has been used to effectively treat a variety of hepatobiliary diseases in clinical practice. In the present study, we performed a systematic study on the composition and structure of its polysaccharides, and then employed a TGF-ß1-induced human intrahepatic bile duct epithelial cell-epithelial-mesenchymal transition (HIBEC-EMT) model to investigate their effects on treating primary biliary cholangitis (PBC) based on hepatic bile duct fibrosis. Four polysaccharide fractions were obtained from H. sinensis mycelia by hot-water extraction, DEAE-cellulose column and gradient ethanol precipitation separation. HSWP-1a was an α-(1,4)-D-glucan; HSWP-1b and HSWP-1d mainly consisted of mannoglucans with a backbone composed of 1,4-linked α-D-Glcp and 1,4,6-linked α-D-Manp residues branched at O-6 of the 1,4-linked α-D-Glcp with a 1-linked α-D-Glcp as a side chain; and HSWP-1c mainly contained galactomannoglucans. These polysaccharide fractions protected HIBECs from a TGF-ß1-induced EMT, according to HIBEC morphological changes, cell viability, decreased E-cadherin and ZO-1 expression, and increased vimentin and collagen I expression. Furthermore, the effects of the polysaccharides might be mediated by inhibiting the activation of the TGF-ß/Smad signaling pathway, which attenuated hepatic bile duct fibrosis and potential PBC effects.

Association of congenital iodine deficiency syndrome and differentiated thyroid cancer: a Mendelian randomization study.

BACKGROUND: The effect of iodine deficiency, especially during the fetal period, on thyroid cancer risk remains unclear. The evidence from observational studies is controversial because of the inevitable confounding factors. We studied the causal effect of congenital iodine deficiency on differentiated thyroid cancer (DTC) based on Mendelian randomization (MR). METHODS: Two-Sample MR analysis was performed using data from published genome-wide association studies, including congenital iodine deficiency syndrome (CIDS) (353 cases, 187,684 controls) and DTC (649 cases, 431 controls) data. RESULTS: There was a causal relationship between CIDS and DTC (P < 0.05), with CIDS increasing the DTC risk by 37.4% (OR = 1.374, 95%CI = 1.110-1.700). Heterogeneity tests and tests of multiple validities indicated that the results were solid and reliable (all P < 0.05). CONCLUSIONS: Fetal iodine deficiency increases the risk of DTC, so future clinical studies should focus on the effect of iodine supplementation during pregnancy to reduce the risk of thyroid cancer in the offspring.

Didymin alleviates metabolic dysfunction-associated fatty liver disease (MAFLD) via the stimulation of Sirt1-mediated lipophagy and mitochondrial biogenesis.

BACKGROUND: Metabolic dysfunction-associated fatty liver disease (MAFLD) is one of the most prevalent metabolic syndromes worldwide. However, no approved pharmacological treatments are available for MAFLD. Chenpi, one kind of dried peel of citrus fruits, has traditionally been utilized as a medicinal herb for liver diseases. Didymin is a newly identified oral bioactive dietary flavonoid glycoside derived from Chenpi. In this study, we investigated the therapeutic potential of Didymin as an anti-MAFLD drug and elucidated its underlying mechanisms. METHODS: High-fat diet (HFD)-induced MAFLD mice and alpha mouse liver 12 (AML12) cells were utilized to evaluate the effects and mechanisms of Didymin in the treatment of MAFLD. Liver weight, serum biochemical parameters, and liver morphology were examined to demonstrate the therapeutic efficacy of Didymin in MAFLD treatment. RNA-seq analysis was performed to identify potential pathways that could be affected by Didymin. The impact of Didymin on Sirt1 was corroborated through western blot, molecular docking analysis, microscale thermophoresis (MST), and deacetylase activity assay. Then, a Sirt1 inhibitor (EX-527) was utilized to confirm that Didymin alleviates MAFLD via Sirt1. Western blot and additional assays were used to investigate the underlying mechanisms. RESULTS: Our results suggested that Didymin may possess therapeutic potential against MAFLD in vitro and in vivo. By promoting Sirt1 expression as well as directly binding to and activating Sirt1, Didymin triggers downstream pathways that enhance mitochondrial biogenesis and function while reducing apoptosis and enhancing lipophagy. CONCLUSIONS: These suggest that Didymin could be a promising medication for MAFLD treatment. Furthermore, its therapeutic effects are mediated by Sirt1.

RMI1 facilitates repair of ionizing radiation-induced DNA damage and maintenance of genomic stability.

Ionizing radiation (IR) causes a wide variety of DNA lesions, of which DNA double-stranded breaks (DSBs) are the most deleterious. Homologous recombination (HR) is a crucial route responsible for repairing DSBs. RecQ-mediated genome instability protein 1 (RMI1) is a member of an evolutionarily conserved Bloom syndrome complex, which prevents and resolves aberrant recombination products during HR, thereby promoting genome stability. However, little is known about the role of RMI1 in regulating the cellular response to IR. This study aimed to understand the cellular functions and molecular mechanisms by which RMI1 maintains genomic stability after IR exposure. Here, we showed IR upregulated the RMI1 protein level and induced RMI1 relocation to the DNA damage sites. We also demonstrated that the loss of RMI1 in cells resulted in enhanced levels of DNA damage, sustained cell cycle arrest, and impaired HR repair after IR, leading to reduced cell viability and elevated genome instability. Taken together, our results highlighted the direct roles of RMI1 in response to DNA damage induced by IR and implied that RMI1 might be a new genome safeguard molecule to radiation-induced damage.

Puerarin ameliorates metabolic dysfunction-associated fatty liver disease by inhibiting ferroptosis and inflammation.

Metabolic dysfunction-associated fatty liver disease (MAFLD) is frequently linked to type 2 diabetes mellitus (T2DM), and both conditions exacerbate the progression of the other. However, there is currently no standardized treatment or drug for MAFLD. In this study, A MAFLD animal model through a high-fat diet (HFD) along with administration of streptozotocin (STZ), and palmitic acid (PA)-induced AML12 cells were treated by puerarin. The objective of this study was to assess the therapeutic effect of puerarin, a flavonoid substance that possesses various pharmacological properties, on MAFLD. The results showed that puerarin administration enhanced glucose tolerance and insulin sensitivity, while also mitigating liver dysfunction and hyperlipidemia in MAFLD mice. Moreover, puerarin attenuated oxidative stress levels and inflammation in the liver. Transmission electron microscopy and Western blot analysis indicated that puerarin inhibited ferroptosis in vivo. Further mechanistic investigations revealed that puerarin upregulated SIRT1 expression, increased nuclear factor erythroid 2-related factor 2 (Nrf2) protein levels, and facilitated translocation into the nucleus. The protective effect of puerarin on PA-induced AML12 cells was diminished by the utilization of EX-527 (a SIRT1 inhibitor) and Nrf2 siRNA. Overall, the results demonstrate that puerarin ameliorates MAFLD by suppressing ferroptosis and inflammation via the SIRT1/Nrf2 signaling pathway. The results emphasize the possible medicinal application of puerarin for managing MAFLD.

Attenuated Salmonella carrying siRNA-CD24 improved the effect of oxaliplatin on HCC.

Oxaliplatin is a chemotherapy drug currently utilized in the treatment of advanced cancer patients. However, its tolerability poses a limitation to its clinical application. Studies have demonstrated that the presence of tumor-associated macrophages is positively correlated with poor prognosis in various solid tumors, including hepatocellular carcinoma (HCC), and is a significant factor contributing to oxaliplatin resistance. Therefore, targeting tumor-associated macrophages may be an effective strategy to improve the efficacy of oxaliplatin in the treatment of HCC patients. CD24 is a novel target for tumor therapy that can interact with the inhibitory receptor Siglec-10 on tumor-associated macrophages, transmitting immune inhibitory signals and inhibiting macrophage phagocytosis function. In this study, we utilized RNAi technology to inhibit the expression of CD24 in tumor cells and combined it with oxaliplatin, resulting in reduced tumor invasion, migration, and proliferation, as well as increased cell apoptosis. Furthermore, immunofluorescence and flow cytometry results indicated that both the single treatment group and combination treatment group enhanced the infiltration of immune cells. This study presents a novel approach to identifying combination therapy and targets for the clinical treatment of HCC with oxaliplatin.

BRAFV600E genetic testing should be recommended for Bethesda III or V thyroid nodules based on fine-needle aspiration.

The preoperative diagnosis of thyroid nodules now routinely includes BRAFV600E genetic testing in most provincial and municipal hospitals in China. This study identified the most suitable patients of thyroid nodule for BRAFV600E genetic testing. We retrospectively collected data of patients from the Hospital Information System that had undergone fine needle aspiration biopsy (FNAB) from May 2019 to December 2021. Data of FNAB, BRAFV600E genetic testing, and post-surgical pathological diagnosis were collected. A total of 12,392 patients were included in this study. Among them, 7,010 patients underwent solely FNAB, while 5,382 patients had both FNAB and BRAFV600E genetic testing. In the FNAB group, 2,065 thyroid nodules were surgically removed, with a 93.12% malignancy rate. In the FNAB + BRAF group, 2,005 nodules were dissected, and the malignancy rate was 98.20%. However, it was evident that in the subgroups, the combination of FNAB and BRAFV600E genetic testing only benefited Bethesda III (p < 0.001) and V (p = 0.001) nodules. Overall, the combination of FNAB with BRAFV600E genetic testing significantly improved the malignancy rate of surgical thyroid nodes at our hospital when compared to FNAB alone. The subgroup analysis showed that BRAFV600E genetic testing only benefited Bethesda III and V nodules. These findings provide a clinical reference for rationally selecting the most suitable population for BRAFV600E genetic testing.

Protective and immunomodulatory effects of mesenchymal stem cells on multiorgan injury in male rats with heatstroke.

Heatstroke (HS) causes multiple organ dysfunction syndrome (MODS) with a mortality rate of 60% after hospitalization. Currently, there is no effective and targeted approach for the treatment of HS. Despite growing evidence that mesenchymal stem cells (MSCs) may reduce multiorgan damage and improve survival through immunomodulatory effects in several diseases, no one has tested whether MSCs have immunomodulatory effects in heatstroke. The present study focused on pathological changes and levels of the cytokines and immunoglobulins to investigate the mechanisms underlying the protective effect and the anti-inflammatory effects of MSCs. We found that MSCs treatment significantly reduced the 28-day mortality rate (P < 0.05), the levels of hepatic and renal function markers on day 1 (P < 0.01) and the pathological lesion scores of multiple organs in HS rats. The levels of IgG1, IgM, and IgA of the HS + MSC group was significantly higher than that in HS group on days 3 and 28(P < 0.05). In conclusion, MSCs contribute to protecting against multiorgan injury, reducing pro-inflammatory cytokines, stabilizing immunoglobulins, and reducing the mortality rate of HS rats.

Analysis of differences in tobacco leaf microbial communities after redrying in Chinese provinces and from abroad.

Microorganisms play an important role in the tobacco aging process. Before the aging process, raw tobacco leaves must be threshed and redried. In order to explore the differences of microbial community structure of threshed and redried tobacco leaves from different origins at home and abroad, 14 groups of tobacco leaves from 8 different countries were tested by high-throughput DNA sequencing and microbiology analysis. Then, through amplicon sequence variants (ASV) cluster analysis, Venn diagram and species labeling and other microbial diversity analysis, the dominant bacteria and fungi on the surface of threshed and redried tobacco leaves were obtained. The results showed that there were significant differences in the composition of tobacco bacteria and fungi after threshing and redrying from different geographical areas. The relative abundance of Microbacterium and Sphingomonas in domestic tobacco leaves was significantly higher than that of foreign tobacco leaves. The relative abundance of Pseudomonas in foreign tobacco bacterial colonies was significantly higher than that of domestic tobacco leaves. In terms of fungi, the relative abundance of Aspergillus and Alternaria in domestic tobacco leaves was significantly higher than that of foreign tobacco leaves. Septoria, Sampaiozyma, Cladosporium and Phoma account for significantly higher proportions of foreign tobacco leaves. These microorganisms may be indispensable in aging process to form different flavors of tobacco leaves. It provides an important theoretical basis for the further use of microorganisms to promote tobacco leaf aging.

Characterization of bacterial community in tobacco leaves at flue-curing and redrying processing stages.

During the processing of tobacco leaves, flue-curing and redrying can affect the structure of bacterial community, having an effect on the aging quality of tobacco leaves. In order to characterize the effects of flue-curing and redrying on the bacterial community of tobacco leaves, the bacterial community of samples at different processing stages (before flue-curing, after flue-curing, before redrying and after redrying) was analyzed using Illumina sequencing. A total of 33 phyla, 79 classes, 195 orders, 344 families, 826 genera and 7922 ASVs were obtained from 36 samples. There was no significant difference in the core bacterial groups of tobacco leaf at four processing stages. Proteobacteria dominated at the phylum level. Sphingomonas, Pseudomonas and Methylobacterium were the main genera shared by all samples. The functional prediction by PICRUSt showed an increase in the relative abundance of pathway related to metabolism after flue-curing and pathway related to environmental information processing after redrying. This study, we analyzed the changes of bacterial community and structural composition of tobacco leaves from flue-curing to redrying, and found that flue-curing had a greater effect on the microbial community than redrying. This is conducive for the exploration of microbial resources and improvement of tobacco leaf quality.

Synchronous targeted delivery of TGF-ß siRNA to stromal and tumor cells elicits robust antitumor immunity against triple-negative breast cancer by comprehensively remodeling the tumor microenvironment.

The poor permeability of therapeutic drugs, limited T-cell infiltration, and strong immunosuppressive tumor microenvironment of triple-negative breast cancer (TNBC) acts as a prominent barrier to the delivery of drugs and immunotherapy including programmed cell death ligand-1 antibody (anti-PD-L1). Transforming growth factor (TGF)-ß, an important cytokine produced by cancer-associated fibroblasts (CAFs) and tumor cells contributes to the pathological vasculature, dense tumor stroma and strong immunosuppressive tumor microenvironment (TME). Herein, a nanomedicine platform (HA-LSL/siTGF-ß) employing dual-targeting, alongside hyaluronidase (HAase) and glutathione (GSH) triggered release was elaborately constructed to efficiently deliver TGF-ß small interference RNA (siTGF-ß). It was determined that this system was able to improve the efficacy of anti-PD-L1. The siTGF-ß nanosystem efficiently silenced TGF-ß-related signaling pathways in both activated NIH 3T3 cells and 4T1 cells in vitro and in vivo. This occurred firstly, through CD44-mediated uptake, followed by rapid escape mediated by HAase in endo/lysosomes and release of siRNA mediated by high GSH concentrations in the cytoplasm. By simultaneous silencing of TGF-ß in stromal and tumor cells, HA-LSL/siTGF-ß dramatically reduced stroma deposition, promoted the penetration of nanomedicines for deep remodeling of the TME, improved oxygenation, T cells infiltration and subsequent anti-PD-L1 deep penetration. The double suppression of TGF-ß has been demonstrated to promote blood vessel normalization, inhibit an epithelial-to-mesenchymal transition (EMT), and further modify the immunosuppressive TME, which was supported by an overall increase in the proportion of dendritic cells and cytotoxic T cells. Further, a reduction in the proportion of immunosuppression cells such as regulatory T cells and myeloid-derived suppressor cells was also observed in the TME. Based on the comprehensive remodeling of the tumor microenvironment by this nanosystem, subsequent anti-PD-L1 therapy elicited robust antitumor immunity. Specifically, this system was able to suppress the growth of both primary and distant tumor while preventing tumor metastasis to the lung. Therefore, the combination of the dual-targeted siTGF-ß nanosystem, alongside anti-PD-L1 may serve as a novel method to enhance antitumor immunotherapy against stroma-rich TNBC.

Molecularly Imprinted Polymers with Shape-Memorable Imprint Cavities for Efficient Separation of Hemoglobin from Blood.

Efficient separation and purification of hemoglobin from blood and other complicated biological fluids still remains a big challenge. Molecularly imprinted polymers (MIPs) of hemoglobin are potential choices; however, they suffer from severe problems including difficult template removal and low imprinting efficiency like other protein-imprinted polymers. Herein, a novel MIP of bovine hemoglobin (BHb) was designed in which a peptide crosslinker (PC), instead of the commonly used crosslinkers, was used. The PC, a random copolymer of lysine and alanine, adopts an α-helical conformation at pH 10 but transits to a random coil conformation at pH 5. The introduction of alanine residues lowers the pH range at which the PC undergoes helix-coil transition. The imprint cavities in the polymers are shape-memorable due to the reversible and precise helix-coil transition of the peptide segments in the polymers. They can be enlarged by lowering pH from 10 to 5, thus allowing complete removal of the template protein under mild conditions. When the pH is adjusted back to 10, their original size and shape will be recovered. Therefore, the MIP binds the template protein BHb with high affinity. Compared with the MIP crosslinked with the commonly used crosslinker, the imprinting efficiency of the PC-crosslinked MIP is significantly improved. In addition, both the maximum adsorption capacity (641.9 mg/g) and imprinting factor (7.2) are much higher than the BHb MIPs reported previously. The new BHb MIP also exhibits high selectivity toward BHb and good reusability. Thanks to the high adsorption capacity and high selectivity of the MIP, when it was applied to extract BHb from bovine blood, BHb in the blood sample was extracted almost completely, and high purity product was obtained.

Peptide-crosslinked molecularly imprinted polymers for efficient separation of immunoglobulin G from human serum.

Low-cost and highly effective methods are highly desirable to replace the costly ethanol fractionation and affinity chromatography in IgG isolation from human plasma. Molecularly imprinted polymers (MIPs) of IgG are potential candidates, however, they still suffer from severe problems such as difficult template removal and low imprinting efficiency. Here, a recently developed strategy was adopted to overcome these problems. The MIPs were synthesized using poly(L-glutamic acid) (PLGA) peptide crosslinkers instead of commonly used crosslinkers, such as N,N-methylenebisacrylamide (BIS). Because of the pH-induced helix-coil transition and the precise folding of the peptide segments in the polymers, the imprint cavities can be enlarged by adjusting the pH from 5.0 to 7.0, but their original size and shape are restored when the pH is adjusted back. Therefore, the IgG template can be eluted completely under mild conditions, and significantly improved imprinting efficiency can be achieved. Compared with BIS-crosslinked MIP, 8.6 times more binding sites can be created by molecular imprinting PLGA-crosslinked MIP. The factors influencing the performance of the MIP were studied systematically. An optimized MIP with a high adsorption capacity (612.5 mg g-1), high IF (4.92), and high selectivity was obtained. The adsorption capacity and selectivity of the MIP are much higher than the previously reported IgG MIPs. Because of its high adsorption capacity and selectivity, it can separate IgG from human serum effectively, affording high purity products.