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Mucosal vaccines offer potential benefits over parenteral vaccines for they can trigger both systemic immune protection and immune responses at the predominant sites of pathogen infection. However, the defense function of mucosal barrier remains a challenge for vaccines to overcome. Here, we show that surface modification of exosomes with the fragment crystallizable (Fc) part from IgG can deliver the receptor-binding domain (RBD) of SARS-CoV-2 to cross mucosal epithelial layer and permeate into peripheral lung through neonatal Fc receptor (FcRn) mediated transcytosis. The exosomes F-L-R-Exo are generated by genetically engineered dendritic cells, in which a fusion protein Fc-Lamp2b-RBD is expressed and anchored on the membrane. After intratracheally administration, F-L-R-Exo is able to induce a high level of RBD-specific IgG and IgA antibodies in the animals’ lungs. Furthermore, potent Th1 immune-biased T cell responses were also observed in both systemic and mucosal immune responses. F-L-R-Exo can protect the mice from SARS-CoV-2 pseudovirus infection after a challenge. These findings hold great promise for the development of a novel respiratory mucosal vaccine approach.

Abstract Background Cucurbita pepo cv Dayangua (CPD) is an edible plant with diverse pharmacological properties. The current research on CPD has primarily focused on initial investigations of its chemical composition and pharmacological effects, and no comprehensive toxicity assessment has been conducted to date. Methods In the present study, the toxicity of CPD was evaluated through both acute and sub-chronic oral toxicity tests in mice. 16S rDNA sequencing was used to analyze the composition of the gut microbiota of mice at different time points to observe the effect of CPD on these microbial communities. Results In the acute toxicity test, CPD exhibited low toxicity, with a median lethal dose (LD50) > 2000 mg/kg. The sub-chronic toxicity test indicated that CPD administration at doses of 200, 400, and 600 mg/kg did not cause mortality or significant organ damage in mice. Furthermore, analysis of the gut microbiota after gavage administration of CPD at 400 and 600 mg/kg revealed an improved abundance of some beneficial gut bacteria. Conclusions In summary, no acute or sub-chronic toxic effects were observed in mice following the oral administration of CPD. CPD did not affect the structure and diversity of the gut microbiota and may contribute to an increase in the number of beneficial gut bacteria.

Rice is one of the most important food crops in the world and is important for global food security. However, damage caused by striped stem borer (SSB) seriously threatens rice production and can cause significant yield losses. The development and use of resistant rice varieties or genes is currently the most effective strategy for controlling SSB. We genotyped 201 rice samples using 2849855 high-confidence single nucleotide polymorphisms (SNPs). We conducted a genome-wide association study (GWAS) based on observed variation data of 201 rice cultivars resistant to SSB. We obtained a quantitative trait locus (QTL)-qRSSB4 that confers resistance to SSB. Through annotation and analysis of genes within the qRSSB4 locus, as well as qRT-PCR detection in resistant rice cultivars, we ultimately selected the candidate gene LOC_Os04g34140 (named OsRSSB4) for further analysis. Next, we overexpressed the candidate gene OsRSSB4 in Nipponbare through transgenic methods, resulting in OsRSSB4 overexpressing lines (OsRSSB4OE). In addition, we evaluated the insect resistance of OsRSSB4OE lines using wild type (Nipponbare) as a control. The bioassay experiment results of live plants showed that after 20 days of inoculation with SSB, the withering heart rate of OsRSSB4OE-34 and OsRSSB4OE-39 lines was only 8.3% and 0%, with resistance levels of 1 and 0, respectively; however, the withering heart rate of the wild-type reached 100%, with a resistance level of 9. The results of the in vitro stem bioassay showed that, compared with the wild-type, the average corrected mortality rate of the SSB fed on the OsRSSB4OE line reached 94.3%, and the resistance reached a high level. In summary, we preliminarily confirmed that OsRSSB4 positively regulates the defense of rice against SSB. This research findings reveal new SSB resistance gene resources, providing an important genetic basis for SSB resistance breeding in rice crops.

Carbohydrate-binding modules (CBMs) are essential virulence factors in phytopathogens, particularly the extensively studied members from the CBM50 gene family, which are known as lysin motif (LysM) effectors and which play crucial roles in plant–pathogen interactions. However, the function of CBM50 in Tilletia horrida has yet to be fully studied. In this study, we identified seven CBM50 genes from the T. horrida genome through complete sequence analysis and functional annotation. Their phylogenetic relationships, conserved motifs, promoter elements, and expression profile were further analyzed. The phylogenetic analysis indicated that these seven ThCBM50 genes were divided into three groups, and close associations were observed among proteins with similar protein motifs. The promoter cis-acting elements analysis revealed that these ThCBM50 proteins may be involved in the regulation of the phytohormones, stress response, and meristem expression of the host plant during T. horrida infection. The transcriptome data indicated that four ThCBM50 genes were upregulated during T. horrida infection. We further found that ThCBM50_1 caused cell death in the leaves of Nicotiana benthamiana, and its signal peptide (SP) had a secreting function. These results offer important clues that highlight the features of T. horrida CBM50 family proteins and set the stage for further investigation into their roles in the interactions between T. horrida and rice.

The phyllosphere, comprising the aerial portions of plants, is a vibrant ecosystem teeming with diverse microorganisms crucial for plant health and productivity. This review examines the functional roles of phyllosphere microorganisms in rice (Oryza sativa), focusing on their importance in nutrient uptake, disease resistance, and growth promotion. The molecular mechanisms underlying these interactions are explored along with their potential applications in enhancing sustainable rice production. The symbiotic relationships between rice plants and their associated microorganisms are highlighted, offering insights into improved agricultural practices. Furthermore, this review addresses the challenges and future developments in translating laboratory findings into practical applications. By synthesizing current research, this comprehensive analysis serves as a valuable resource for leveraging phyllosphere microbes in rice farming and related fields.

Background/Objectives: Omicron, the predominant variant of SARS-CoV-2, exhibits strong immune-evasive properties, leading to the reduced efficacy of existing vaccines. Consequently, the development of versatile vaccines is imperative. Intranasal mRNA vaccines offer convenient administration and have the potential to enhance mucosal immunity. However, delivering vaccines via the nasal mucosa requires overcoming complex physiological barriers. The aim of this study is to modify PEGylated lipids to enhance the mucosal immune efficacy of the vaccine. Methods: The PEGylated lipid component of lipid nanoparticle (LNP) delivery vectors was modified with chitosan or mannose to generate novel LNPs that enhance vaccine adhesion or targeting on mucosal surfaces. The impact of the mRNA encoding the receptor-binding domain of Omicron BA.4/BA.5 on the immune response was examined. Results: Compared to the unmodified LNP group, the IgG and IgA titers in the chitosan or mannose-modified LNP groups showed an increasing trend. The chitosan-modified group showed better effects. Notably, the PEGylated lipid with 1.5 mol% of chitosan modification produced high levels of IgG1 and IgG2a antibodies, promoting Th1/Th2 responses while also generating high levels of IgA, which can induce stronger cellular immunity, humoral immunity, and mucosal immunity. Conclusions: The 1.5 mol% of chitosan-modified LNPs (mRNA-LNP-1.5CS) can serve as a safe and effective carrier for intranasal mRNA vaccines, offering a promising strategy for combating the Omicron variant.

Immunotherapy has revolutionized the landscape of cancer treatment. However, single immunotherapy only works well in a small subset of patients. Combined immunotherapy with antitumor synergism holds considerable potential to boost the therapeutic outcome. Nevertheless, the synergistic, additive or antagonistic antitumor effects of combined immunotherapies have been rarely explored. Herein, we established a novel combined cancer treatment modality by synergizing p21-activated kinase 4 (PAK4) silencing with immunogenic phototherapy in engineered extracellular vesicles (EVs) that were fabricated by coating M1 macrophage-derived EVs on the surface of the nano-complex cores assembled with siRNA against PAK4 and a photoactivatable polyethyleneimine. The engineered EVs induced potent PAK4 silencing and robust immunogenic phototherapy, thus contributing to effective antitumor effects in vitro and in vivo. Moreover, the antitumor synergism of the combined treatment was quantitatively determined by the CompuSyn method. The combination index (CI) and isobologram results confirmed that there was an antitumor synergism for the combined treatment. Furthermore, the dose reduction index (DRI) showed favorable dose reduction, revealing lower toxicity and higher biocompatibility of the engineered EVs. Collectively, the study presents a synergistically potentiated cancer treatment modality by combining PAK4 silencing with immunogenic phototherapy in engineered EVs, which is promising for boosting the therapeutic outcome of cancer immunotherapy.

Abstract Rice kernel smut (RKS), caused by the fungus Tilletia horrida, has become a major disease in rice-growing areas worldwide, especially since the widespread cultivation of high-yielding hybrid rice varieties. The disease causes a significant yield loss during the production of rice male sterile lines by producing masses of dark powdery teliospores. This review mainly summarizes the pathogenic differentiation, disease cycle, and infection process of the T. horrida, as well as the decoding of the T. horrida genome, functional genomics, and effector identification. We highlight the identification and characterization of virulence-related pathways and effectors of T. horrida, which could foster a better understanding of the rice–T. horrida interaction and help to elucidate its pathogenicity molecular mechanisms. The multiple effective disease control methods for RKS are also discussed, included chemical fungicides, the mining of resistant rice germplasms/genes, and the monitoring and early warning signs of this disease in field settings.

Abstract Background The sustainable development of rice production is facing severe threats by a variety of pathogens, such as necrotrophic Rhizoctonia solani and hemibiotrophic Xanthomonas oryzae pv. oryzae (Xoo). Mining and applying resistance genes to increase the durable resistance of rice is an effective method that can be used to control these diseases. Results In this research, we isolated and characterized CYP716A16, which is a positive regulator of rice to R. solani AG1-IA and Xoo, and belongs to the cytochrome P450 (CYP450) protein 716A subfamily. Overexpression (OE) of CYP716A16 resulted in enhanced resistance to R. solani AG1-IA and Xoo, while RNA interference (RNAi) of CYP716A16 resulted in increased susceptibility compared with wild-type (WT) plants. Additionally, jasmonic acid (JA)-dependent defense responses and reactive oxygen species (ROS) were activated in the CYP716A16-OE lines after R. solani AG1-IA inoculation. The comparative transcriptomic and metabolomics analysis of CYP716A16-OE and the WT lines showed that OE of CYP716A16 activated the biosynthesis of flavonoids and increased the amounts of narcissoside, methylophiopogonanone A, oroxin A, and amentoflavone in plants. Conclusion Based on these results, we suggest that JA-dependent response, ROS level, multiple resistance-related proteins, and flavonoid contents play an important role in CYP716A16-regulated R. solani AG1-IA and Xoo resistance. Our results broaden our knowledge regarding the function of a P450 protein 716A subfamily in disease resistance and provide new insight into the molecular mechanism of rice immune response.

Abstract Bacillus thuringiensis (Bt) is an important biological insecticide used to management of different agricultural pests by producing toxic parasporal crystals proteins. Strain HD521 has an antagonistic effect against Rhizoctonia solani AG1IA, the causal agent of rice sheath blight. This strain with three cry7 genes can the formation of bipyramidal parasporal crystals (BPCs). BPCs are used for insecticidal activities against Henosepilachna vigintioctomaculata larva (Coleoptera). Strain HS18-1 contains different types of BPCs encoding genes and has effective toxicity for Lepidoptera and Diptera insects. Here we report the whole genome sequencing and assembly of HD521 and HS18-1 strains and analyzed the genome constitution covering virulence factors, types of plasmid, insertion sequences, and prophage sequences. The results showed that the genome of strain HD521 contains a circular chromosome and six circular plasmids, encoding eight types of virulence protein factors [Immune Inhibitor A, Hemolytic Enterotoxin, S-layer protein, Phospholipase C, Zwittermicin A-resistance protein, Metalloprotease, Chitinase, and N-acyl homoserine lactonase (AiiA)], four families of insertion sequence, and comprises six pro-phage sequences. The genome of strain HS18-1 contains one circular chromosome and nine circular plasmids, encoding five types of virulence protein factors [Hemolytic Enterotoxin, S-layer protein, Phospholipase C, Chitinase, and N-acyl homoserine lactonase (AiiA)] and four families of insertion sequence, and comprises of three pro-phage sequences. The obtained results will contribute to deeply understand the B. thuringiensis strain HD521 and HS18-1 at the genomic level.

Ovarian cancer is currently one of the most common malignant tumors in females with poor survival rates around the world, killing about 200,000 women each year. Although great progress has been made in treatment, most patients receiving first-line therapy experience tumor recurrence. The tumor microenvironment plays an important role in regulating the progression and prognosis of ovarian cancer. Cancer-associated mesothelial cells are the main cell population in the tumor microenvironment, which affect the progression, prognosis and chemical resistance of ovarian cancer. Cancer-associated mesothelial cells can also interact with other microenvironmental components, such as exosomes, macrophages, and adipocytes. Some studies have developed drugs targeting cancer-associated mesothelial cells in ovarian cancer to evaluate the therapeutic efficiency. In this review we highlighted the key role of cancer-associated mesothelial cells in the progression and prognosis of ovarian cancer. We also described the progress of cancer-associated mesothelial cells targeted therapy for ovarian cancer. Continued insight into the role of cancer-associated mesothelial cells in ovarian cancer will potentially contribute to the development of new and effective therapeutic regiments.

Metastasis is responsible for 90% of deaths in cancer patients. Most patients diagnosed with metastatic cancer will die within 5 years. PA is good for health and has become an emerging adjuvant therapy for cancer survivors. Regular moderate exercise substantially lowers the incidence and recurrence of several cancers, alleviates cancer-related adverse events, enhances the efficacy of anti-cancer treatments, and improves the quality of life of cancer patients. Revealing the mechanisms of PA inhibiting tumor metastasis could upgrade our understanding of cancer biology and help researchers explore new therapeutic strategies to improve survival in cancer patients. However, it remains poorly understood how physical activity prevents metastasis by modulating tumor behavior. The immune system is involved in each step of tumor metastasis. From invasion to colonization, immune cells interact with tumor cells to secret cytokines and proteases to remodel the tumor microenvironment. Substantial studies demonstrated the ability of physical activity to induce antitumor effects of immune cells. This provides the possibility that physical activity can modulate immune cells behavior to attenuate tumor metastasis. The purpose of this review is to discuss and summarize the critical link between immune function and exercise in metastasis prevention.

The present review sets out to discuss recent developments of the effects and mechanisms of carrier properties on their circulation time. For most drugs, sufficient in vivo circulation time is the basis of high bioavailability. Drug carrier plays an irreplaceable role in helping drug avoid being quickly recognized and cleared by mononuclear phagocyte system, to give drug enough time to arrive at targeted organ and tissue to play its therapeutic effect. The physical and chemical properties of drug carriers, such as size, shape, surface charge and surface modification, would affect their in vivo circulation time, metabolic behavior and biodistribution. The final circulation time of carriers is determined by the balance between macrophage recognitions, blood vessel penetration and urine excretion. Therefore, when designing the drug delivery system, we should pay much attention to the properties of drug carriers to get enough in vivo circulation time to arrive at target site eventually. This article mainly reviews the effect of carrier size, size, surface charge and surface properties on its circulation time in vivo, and discusses the mechanism of these properties affecting circulation time. This review has reference significance for the research of long-circulation drug delivery system.

Abstract Background Rice (Oryza sativa) bacterial leaf blight (BLB), caused by the hemibiotrophic Xanthomonas oryzae pv. oryzae (Xoo), is one of the most devastating diseases affecting the production of rice worldwide. The development and use of resistant rice varieties or genes is currently the most effective strategy to control BLB. Results Here, we used 259 rice accessions, which are genotyped with 2 888 332 high-confidence single nucleotide polymorphisms (SNPs). Combining resistance variation data of 259 rice lines for two Xoo races observed in 2 years, we conducted a genome-wide association study (GWAS) to identify quantitative trait loci (QTL) conferring plant resistance against BLB. The expression levels of genes, which contains in GWAS results were also identified between the resistant and susceptible rice lines by transcriptome analysis at four time points after pathogen inoculation. From that 109 candidate resistance genes showing significant differential expression between resistant and susceptible rice lines were uncovered. Furthermore, the haplotype block structure analysis predicted 58 candidate genes for BLB resistance based on Chr. 7_707158 with a minimum P-value (–log 10 P = 9.72). Among them, two NLR protein-encoding genes, LOC_Os07g02560 and LOC_Os07g02570, exhibited significantly high expression in the resistant line, but had low expression in the susceptible line of rice. Conclusions Together, our results reveal novel BLB resistance gene resources, and provide important genetic basis for BLB resistance breeding of rice crops.

3D printing is a promising technology used in the fabrication of complex oral dosage delivery pharmaceuticals. This study first reports an innovative color jet 3D printing (CJ-3DP) technology to produce colorful cartoon levetiracetam pediatric preparations with high accuracy and reproducibility. For this study, the ideal printing ink consisted of 40% (v/v) isopropanol aqueous solution containing 0.05% (w/w) polyvinylpyrrolidone and 4% (w/w) glycerin, which was satisfied with scale-up of the production. The external and internal spatial structures of the tablets were designed to control the appearance and release, and cartoon tablets with admirable appearances and immediate release characteristics were printed. The dosage model showed a good linear relationship between the model volume and the tablet strength (r > 0.999), which proved the potential of personalized administration. The surface roughness indicated that the appearance of the CJ-3DP tablets was significantly better than the first listed 3D printed drug (SpritamⓇ). Moreover, the scanning electron microscopy and porosity results further showed that the tablets have a structure of loose interior and tight exterior, which could ensure good mechanical properties and rapid dispersion characteristics simultaneously. In conclusion, the innovative CJ-3DP technology can be used to fabricate personalized pediatric preparations for improved compliance. Due to the stable formulation and fabrication process, this technology has the potential in scale-up production.

Brain-targeting delivery of 1,1′-methylenebis[4-[(hydroxyimino)methyl]-pyridinium] dimethanesulfonate (MMB4 DMS) is limited by its hydrophilic property and chemical instability. In order to solve this problem, herein, we develop a facile protocol through combining antisolvent precipitation and emulsion-solvent evaporation method to synthesize midazolam (MDZ) coated MMB4 DMS (MMB4@MDZ) nanoparticles. The as-prepared MMB4@MDZ had a MMB4 DMS nanocrystal (MMB4-NC) core and a MDZ shell. The MDZ shell prevented the MMB4-NC core from contacting the aqueous environment, and thus, guaranteed the chemical stability of MMB4 DMS. Most charmingly, the iron mimic cyclic peptide CRTIGPSVC (CRT) was modified on MMB4@MDZ surfaces to produce CRT-MMB4@MDZ which was endowed with ability to absorb transferrin (Tf)-abundant corona. Taking advantages of the Tf-abundant corona, CRT-MMB4@MDZ achieved transferrin receptor (TfR)-mediated brain-targeting delivery. With the fascinating chemical stability and brain-targeting delivery effect, CRT-MMB4@MDZ showed great clinical transform prospect as a brand-new nanomedicine. Of particular importance, this work promised not only a core–shell carrier-free nanomedicine platform for effective delivery of unstable water-soluble drug, but also a protein corona-manipulating strategy for targeting delivery.

In recent years, nanocrystal technology has been extensively investigated. Due to the submicron particle size and unique physicochemical properties of nanocrystals, they overcome the problems of low drug solubility and poor bioavailability. Although the structures of nanocrystals are simple, the further development of these materials is hindered by their stability. Drug nanocrystals with particle sizes of 1∼1000 nm usually require the addition of stabilizers such as polymers or surfactants to enhance their stability. The stability of nanocrystal suspensions and the redispersibility of solid nanocrystal drugs are the key factors for the large-scale production of nanocrystal preparations. In this paper, the factors that affect the stability of drug nanocrystal preparations are discussed, and related methods for solving the stability problem are put forward.

Poly(lactic-co-glycolic acid) (PLGA) has garnered increasing attention as a candidate drug delivery polymer owing to its favorable properties, including its excellent biocompatibility, biodegradability, non-toxicity, non-immunogenicity, and mechanical strength. PLAG are specifically used as microspheres for the sustained/controlled and targeted delivery of hydrophilic or hydrophobic drugs, as well as biological therapeutic macromolecules, including peptide and protein drugs. PLGAs with different molecular weights, lactic acid (LA)/glycolic acid (GA) ratios, and end groups exhibit unique release characteristics, which is beneficial for obtaining diverse therapeutic effects. This review aims to analyze the composition of PLGA microspheres, and understand the manufacturing process involved in their production, from a quality by design perspective. Additionally, the key factors affecting PLGA microsphere development are explored as well as the principles involved in the synthesis and degradation of PLGA and its interaction with active drugs. Further, the effects elicited by microcosmic conditions on PLGA macroscopic properties, are analyzed. These conditions include variations in the organic phase (organic solvent, PLGA, and drug concentration), continuous phase (emulsifying ability), emulsifying stage (organic phase and continuous phase interaction, homogenization parameters), and solidification process (relationship between solvent volatilization rate and curing conditions). The challenges in achieving consistency between batches during manufacturing are addressed, and continuous production is discussed as a potential solution. Finally, potential critical quality attributes are introduced, which may facilitate the optimization of process parameters.

For efficient intranasal transport of parathyroid hormone (1-34) [PTH(1-34)], there is a great medical need to investigate permeation enhancers for intranasal formulations. In this study, the development of PTH(1-34) intranasal formulations was conducted. Based on conformation and chemical stability studies, the most preferable aqueous environment was determined to be 0.008 M acetate buffer solution (ABS). Subsequently, citric acid and Kolliphor® HS·15 were compared as permeation enhancers. The mechanisms of action of citric acid and Kolliphor® HS·15 were investigated using an in vitro model of nasal mucosa, and Kolliphor® HS·15 led to higher permeability of fluorescein isothiocyanate-labeled PTH(1-34) (FITC-PTH) by enhancing both the transcellular and paracellular routes. Moreover, citric acid showed severe mucosal toxicity resulting in cilia shedding, while Kolliphor® HS·15 did not cause obvious mucosa damage. Finally, Kolliphor® HS·15 was studied as a permeation enhancer using a liquid chromatography tandem mass spectrometry (LC-MS/MS) method. The results showed that 5% and 10% Kolliphor® HS·15 increased the bioavailability of PTH(1-34) to 14.76% and 30.87%, respectively. In conclusion, an effective and biosafe PTH(1-34) intranasal formulation was developed by using 10% Kolliphor® HS·15 as a permeation enhancer. Intranasal formulations with higher concentrations of Kolliphor® HS·15 for higher bioavailability of PTH(1-34) could be further researched.

Pollen fertility is an agronomic trait that strongly influences rice yield. Recent studies have revealed that the development of the pollen wall is required for pollen fertility and is regulated by several genes. However, the mechanisms underlying pollen and pollen wall development in rice remain largely unknown. In the present study, a point mutation in a gene on chromosome 1 was identified that resulted in the production of less and shrunken pollen (LSP) and led to defects in pollen wall formation. This gene was named LSP1 and was found to encode a member of the adenosine triphosphate-binding cassette (ABC) transporter G subfamily, OsABCG3. Two other loss-of-function mutants of LSP1/OsABCG3, generated using CRISPR/Cas9 technology, showed the same male sterile phenotype. The LSP1/OsABCG3 gene showed a spatio-temporal expression pattern in the developing anthers, and is an ortholog of the Arabidopsis genes AtABCG1 and AtABCG16, which play an important role in pollen wall development. Mutation of LSP1/OsABCG3 affected the expression of several genes involved in pollen and pollen wall formation. These results suggest that LSP1/OsABCG3 is critical for normal pollen fertility and shed light on the molecular mechanisms underlying rice pollen wall development.

Currently, there is a shortage of pediatric medicines on the market, and 3D printing technology can more flexibly produce personalized medicines to meet individual needs. The study developed a child-friendly composite gel ink (carrageenan-gelatin), created 3D models by computer-aided design technology, then produced personalized medicines using 3D printing to improve the safety and accuracy of medication for pediatric patients. An in-depth understanding of the printability of different formulations was obtained by analyzing the rheological and textural properties of different gel inks and observing the microstructure of different gel inks, which guided the formulation optimization. Through formulation optimization, the printability and thermal stability of gel ink were improved, and F6 formulation (carrageenan: 0.65%; gelatin: 12%) was selected as the 3D printing inks. Additionally, a personalized dose linear model was established with the F6 formulation for the production of 3D printed personalized tablets. Moreover, the dissolution tests showed that the 3D printed tablets were able to dissolve more than 85% within 30 min and had similar dissolution profiles to the commercially available tablets. This study demonstrates that 3D printing is an effective manufacturing technique that allows for flexible, rapid, and automated production of personalized formulations.

In the last few decades, RNA-based drugs have emerged as a promising candidate to specifically target and modulate disease-relevant genes to cure genetic defects. The key to applying RNA therapy in clinical trials is developing safe and effective delivery systems. Exosomes have been exploited as a promising vehicle for drug delivery due to their nanoscale size, high stability, high biocompatibility, and low immunogenicity. We reviewed and summarized the progress in the strategy and application of exosome-mediated RNA therapy. The challenges of exosomes as a carrier for RNA drug delivery are also elucidated in this article. RNA molecules can be loaded into exosomes and then delivered to targeted cells or tissues via various biochemical or physical approaches. So far, exosome-mediated RNA therapy has shown potential in the treatment of cancer, central nervous system disorders, COVID-19, and other diseases. To further exploit the potential of exosomes for RNA delivery, more efforts should be made to overcome both technological and logistic problems.

Some physical processes such as oil and gas development, metal deposit collection, and groundwater resource migration can cause density changes, for which microgravity monitoring is the most intuitive method to monitor the density change process. Based on the basic principle of microgravity measurement and the idea of multiscale separation, a multiscale, second-order, surface-fitting, residual gravity anomaly extraction method is proposed to separate superimposed microgravity fields. In this method, regional fields of different scales are fitted and calculated successively with the measurement points as the center, so as to separate the gravity anomalies produced by different-depth density bodies. Results from actual data show that this method extracts the reservoir’s residual density characteristics of plane gravity anomaly on the basis of remaining oil distribution characteristics, consistent with reservoir numerical simulation results. A three-dimensional least-squares inversion of the method for extracting residual gravity anomaly was carried out, with the inversion results consistent with the results of vertical remaining oil distribution characteristics and well-test production results.

The molecular motors are protein molecules which can be teamed up to form the active mobile nanomachine with special functions. As a new type of therapeutic target different from normal enzymes or receptors, molecular motor plays a unique role in the transportation of intracellular substances, mitosis, and the construction of cytoskeleton. It has also been reported that molecular motors were capable of regulate the tumor cells' proliferation, related to the occurrence of neurodegenerative diseases, and can be used as a nano carrier to deliver drugs. Molecular motors could be divided into linear and rotary molecular motors according to their different forms of motion. The former includes dynein, kinesin, and myosin, and the latter includes ATP synthase and flagellar motors. In addition, there are also some chemically or biologically modified molecular motors to expand or enhance their therapeutic applications. In this review, after the introduction of the classification, structures, main functions and motion mechanism of molecular motors in detail, the application progress of molecular motors in recent years is summarized. Finally, the current problems and future prospect are also discussed in this paper.

Three-dimensional printing technology, also called additive manufacturing technology, is used to prepare personalized 3D-printed drugs through computer-aided model design. In recent years, the use of 3D printing technology in the pharmaceutical field has become increasingly sophisticated. In addition to the successful commercialization of Spritam® in 2015, there has been a succession of Triastek’s 3D-printed drug applications that have received investigational new drug (IND) approval from the Food and Drug Administration (FDA). Compared with traditional drug preparation processes, 3D printing technology has significant advantages in personalized drug manufacturing, allowing easy manufacturing of preparations with complex structures or drug release behaviors and rapid manufacturing of small batches of drugs. This review summaries the mechanisms of the most commonly used 3D printing technologies, describes their characteristics, advantages, disadvantages, and applications in the pharmaceutical industry, analyzes the progress of global commercialization of 3D printed drugs and their problems and challenges, reflects the development trends of the 3D printed drug industry, and guides researchers engaged in 3D printed drugs.

Three-dimensional (3D) printing is an additive manufacturing technique that creates objects under computer control. Owing to the rapid advancement of science and technology, 3D printing technology has been widely utilized in processing and manufacturing but rarely used in the pharmaceutical field. The first commercial form of Spritam® immediate-release tablet was approved by FDA in 2015, which promoted the advancement of 3D printing technology in pharmaceutical development. Three-dimensional printing technology is able to meet individual treatment demands with customized size, shape, and release rate, which overcomes the difficulties of traditional pharmaceutical technology. This paper intends to discuss the critical process parameters of binder jet 3D printing technology, list its application in pharmaceutical manufacturing in recent years, summarize the still-open questions, and demonstrate its great potential in the pharmaceutical industry.

Purpose: This study aimed to explore the ability of texture parameters combining with machine learning methods in distinguishing intrahepatic cholangiocarcinoma (ICCA) and hepatic lymphoma (HL). Method: A total of 28 patients with HL and 101 patients with ICCA were included. A total of 45 texture features were extracted by the software LifeX from contrast-enhanced computer tomography (CECT) images and 38 of them were eligible. A total of 5 feature selection methods and 9 feature classification methods were used to build the best diagnostic models, combining with the 10-fold cross-validation to assess the accuracy of these models. The discriminative ability of each model was evaluated by receiver operating characteristic analysis. Result: A total of 45 predictive models were built by the cross combination of each selection and classification method to differentiate ICCA from HL. According to the results of test group, most of the models performed well with a large area under the curve (AUC) (>0.85) and high accuracy (>0.85). Random Forest (RF)_Linear Discriminant Analysis (LDA) (AUC = 0.997, accuracy = 0.969) was the best model among all the 45 models. Conclusion: Combining texture parameters from CECT with multiple machine learning models can differentiate ICCA and HL effectively, and RF_LDA performed the best in this process.

Mitochondria are the major energy source for cell functions. However, for the plant fungal pathogens, mitogenome variations and their roles during the host infection processes remain largely unknown. Rhizoctonia solani, an important soil-borne pathogen, forms different anastomosis groups (AGs) and adapts to a broad range of hosts in nature. Here, we reported three complete mitogenomes of AG1-IA RSIA1, AG1-IB RSIB1, and AG1-IC, and performed a comparative analysis with nine published Rhizoctonia mitogenomes (AG1-IA XN, AG1-IB 7/3/14, AG3, AG4, and five Rhizoctonia sp. mitogenomes). These mitogenomes encoded 15 typical proteins (cox1-3, cob, atp6, atp8-9, nad1-6, nad4L, and rps3) and several LAGLIDADG/GIY-YIG endonucleases with sizes ranging from 109,017 bp (Rhizoctonia sp. SM) to 235,849 bp (AG3). We found that their large sizes were mainly contributed by repeat sequences and genes encoding endonucleases. We identified the complete sequence of the rps3 gene in 10 Rhizoctonia mitogenomes, which contained 14 positively selected sites. Moreover, we inferred a robust maximum-likelihood phylogeny of 32 Basidiomycota mitogenomes, representing that seven R. solani and other five Rhizoctonia sp. lineages formed two parallel branches in Agaricomycotina. The comparative analysis showed that mitogenomes of Basidiomycota pathogens had high GC content and mitogenomes of R. solani had high repeat content. Compared to other strains, the AG1-IC strain had low substitution rates, which may affect its mitochondrial phylogenetic placement in the R. solani clade. Additionally, with the published RNA-seq data, we investigated gene expression patterns from different AGs during host infection stages. The expressed genes from AG1-IA (host: rice) and AG3 (host: potato) mainly formed four groups by k-mean partitioning analysis. However, conserved genes represented varied expression patterns, and only the patterns of rps3-nad2 and nad1-m3g18/mag28 (an LAGLIDADG endonuclease) were conserved in AG1-IA and AG3 as shown by the correlation coefficient analysis, suggesting regulation of gene repertoires adapting to infect varied hosts. The results of variations in mitogenome characteristics and the gene substitution rates and expression patterns may provide insights into the evolution of R. solani mitogenomes.

The necrotrophic phytopathogen Rhizoctonia solani (R. solani) is a fungus that causes disease in a wide range of plant species. Fungal genomes encode abundant, small cysteine-rich (SCR) secreted proteins, and the probable importance of these to pathogenesis has been highlighted in various pathogens. However, there are currently no reports of an R. solani SCR-secreted protein with evidential elicitor activity. In this study, the molecular function of 10 SCR-secreted protein genes from R. solani was explored by agroinfiltration into Nicotiana benthamiana (N. benthamiana) leaves, and a novel SCR protein RsSCR10 was identified that triggered cell death and oxidative burst in tobacco. RsSCR10 comprises 84 amino acids, including a signal peptide (SP) of 19 amino acids that is necessary for RsSCR10 to induce tobacco cell death. Elicitation of cell death by RsSCR10 was dependent on Hsp90 but not on RAR1, proving its effector activity. Two cysteine residues have important effects on the function of RsSCR10 in inducing cell death. Furthermore, RsSCR10 showed cross-interaction with five rice molecules, and the inferred functions of these rice proteins suggest they are instrumental in how the host copes with adversity. Overall, this study demonstrates that RsSCR10 is a potential effector that has a critical role in R. solani AG1 IA-host interactions.

Abstract Background Tilletia horrida is a basidiomycete fungus that causes rice kernel smut, one of the most important rice diseases in hybrid rice growing areas worldwide. However, little is known about its mechanisms of pathogenicity. We previously reported the genome of T. horrida, and 597 genes that encoded secreted proteins were annotated. Among these were some important effector genes related to pathogenicity. Results A secretome analysis suggested that five Tilletia fungi shared more gene families than were found in other smuts, and there was high conservation between them. Furthermore, we screened 597 secreted proteins from the T. horrida genome, some of which induced expression in host-pathogen interaction processes. Through transient expression, we demonstrated that two putative effectors could induce necrosis phenotypes in Nicotiana benthamiana. These two encoded genes were up-regulated during early infection, and the encoded proteins were confirmed to be secreted using a yeast secretion system. For the putative effector gene smut_5844, a signal peptide was required to induce non-host cell death, whereas ribonuclease catalytic active sites were required for smut_2965. Moreover, both putative effectors could induce an immune response in N. benthamiana leaves. Interestingly, one of the identified potential host interactors of smut_5844 was laccase-10 protein (OsLAC10), which has been predicted to be involved in plant lignification and iron metabolism. Conclusions Overall, this study identified two secreted proteins in T. horrida that induce cell death or are involved in defense machinery in non-host plants. This research provides a useful foundation for understanding the interaction between rice and T. horrida.

Anastomosis group AG-1 IA of Rhizoctonia solani Khün has a wide host range and threatens crop production. Various glycosyltransferases secreted by phytopathogenic fungi play an essential role in pathogenicity. Previously, we identified a glycosyltransferase RsIA_GT (AG11A_09161) as a secreted protein-encoding gene of R. solani AG-1 IA, whose expression levels increased during infection in rice. In this study, we further characterized the virulence function of RsIA_GT. It is conserved not only in Basidiomycota, including multiple anastomosis groups of R. solani, but also in other primary fungal taxonomic categories. RsIA_GT possesses a signal peptide (SP) for protein secretion, and its functionality was proven using yeast and Nicotiana benthamiana. The SP-truncated form of RsIA_GT (RsIA_GT(ΔS)) expressed in Escherichia coli-induced lesion-like phenotype in rice leaves when applied to punched leaves. However, Agrobacterium-mediated transient expressions of both the full-length RsIA_GT and RsIA_GT(ΔS) did not induce cell death in N. benthamiana leaves. Instead, only RsIA_GT(ΔS) suppressed the cell death induced by two reference cell death factors BAX and INF1 in N.benthamiana. RsIA_GT(ΔS)R154A D168A D170A, a mutant RsIA_GT(ΔS) for the glycosyltransferase catalytic domain, still suppressed the BAX- or INF1-induced cell death, suggesting that the cell death suppression activity of RsIA_GT(ΔS) would be independent from its enzymatic activity. RsIA_GT(ΔS) also suppressed the H2O2 production and callose deposition and showed an effect on the induction of defense genes associated with the expression of BAX and INF1. The transient expression of RsIA_GT(ΔS) in N. benthamiana enhanced the lesion area caused by R. solani AG-1 IA. The secreted glycosyltransferase, RsIA_GT, of R. solani AG-1 IA is likely to have a dual role in virulence inside and outside of host cells.

The quality of active pharmaceutical ingredients (APIs) is an important factor which can affect the safety and efficacy of pharmaceuticals. This study was designed to investigate the nature of paliperidone palmitate (PP) obtained by different crystallization processes, then compare the characteristics between test formulations which prepared PP of different crystallization and reference formulations (Invega Sustenna®) in vitro and in vivo. Two different PPs, namely PP-1 and PP-2, were prepared by different crystallization methods. Contact angle, morphology, and crystallinity of the PPs were characterized. Taking the particle sizes and distribution of Invega Sustenna® as reference, test formulations were prepared by the wet milling method using either a PP-1 or PP-2 sample. Their release behavior, stability in vitro, and pharmacokinetics in vivo were subsequently investigated. The results indicated that PP-2 had a higher surface free energy (SFE). More small particles were attached to the PP-1 surface under the influence of crystallization temperature. Different crystallization processes did not change the crystal of PP, but changed the crystallinity of PP. There was no obvious difference in in vitro releases between test formulations. However, the stability and state of formulation containing PP-2 were better compared to formulations containing PP-1, indicated by differences in crystallinity and SFE. Meanwhile, pharmacokinetic in vivo results demonstrated that the pharmacokinetic profiles and parameters of formulation containing PP-2 and Invega Sustenna® tended to be consistent, but those of formulations containing PP-1 were significantly different from those of formulations containing PP-2 or Invega Sustenna®, and there was burst release phenomenon of formulations containing PP-1 in rats. PP made by different crystallization processes could induce changes in appearance, SFE, and crystallinity, and further affect the stability, state, and pharmacokinetic in vivo formulation.

Physical modeling is critical to study the performance of certain operation in heavy oil reservoirs. A well-designed experiment should guarantee the information gathered from lab would be applied to predict the thermal process in the field. To meet this requirement, the initial and boundary condition similarity between lab and field should be satisfied. It is reasonable to follow certain scaling criteria to fabricate the physical model. In addition to these conventional guidelines, this paper makes following recommendations to ensure a successful thermal recovery experiment, • To control and mitigate the steam channeling between the sand-pack and apparatus wall, the back wall is designed as it can be pushed enough to increase contact pressure. • Heat loss should be handled carefully, which impacting steam chamber growing and causing heat accumulation around the model. • A data acquisition system, based on PXI platform and Labview software, for the thermal recovery experiments had been proved valuable in evaluating the spreading progress of steam chamber.

It is challenging to enhance heavy oil recovery in the late stages of steam flooding. This challenge is due to reduced residual oil saturation, high steam-oil ratio, and lower profitability. A field test of the CO2-assisted steam flooding technique was carried out in the steam-flooded heavy oil reservoir in the J6 block of the Xinjiang oil field (China). In the field test, a positive response to the CO2-assisted steam flooding treatment was observed, including a gradually increasing heavy oil production, an increase in the formation pressure, and a decrease in the water cut. The production wells in the test area mainly exhibited four types of production dynamics, and some of the production wells exhibited production dynamics that were completely different from those during steam flooding. After being flooded via CO2-assisted steam flooding, these wells exhibited a gravity drainage pattern without steam channeling issues, and hence, they yielded stable oil production. In addition, emulsified oil and CO2 foam were produced from the production well, which agreed well with the results of laboratory-scale tests. The reservoir-simulation-based prediction for the test reservoir shows that the CO2-assisted steam flooding technique can reduce the steam-oil ratio from 12 m3 (CWE)/t to 6 m3 (CWE)/t and can yield a final recovery factor of 70%.

Background We aimed to investigate the association between blood pressure (BP) and outcomes in intracerebral hemorrhage (ICH) subtypes with different etiologies. Methods and Results A total of 5656 in‐hospital patients with spontaneous ICH were included between January 2012 and December 2016 in a prospective multicenter cohort study. Etiological subtypes of ICH were assigned using SMASH‐U (structural lesion, medication, amyloid angiopathy, systemic/other disease, hypertension, undetermined) classification. Elevated systolic BP was defined as ≥140 mm Hg. Hypertension was defined as elevated BP for >1 month before the onset of ICH. The primary outcomes were measured as 1‐month survival rate and 3‐month mortality. A total of 5380 patients with ICH were analyzed, of whom 4052 (75.3%) had elevated systolic BP on admission and 3015 (56.0%) had hypertension. In multinomial analysis of patients who passed away by 3 months, systolic BP on admission was significantly different in cerebral amyloid angiopathy (P

Purpose: The purpose of the current study was to evaluate the ability of magnetic resonance (MR) radiomics-based machine-learning algorithms in differentiating glioblastoma (GBM) from primary central nervous system lymphoma (PCNSL).Method: One-hundred and thirty-eight patients were enrolled in this study. Radiomics features were extracted from contrast-enhanced MR images, and the machine-learning models were established using five selection methods (distance correlation, random forest, least absolute shrinkage and selection operator (LASSO), eXtreme gradient boosting (Xgboost), and Gradient Boosting Decision Tree) and three radiomics-based machine-learning classifiers [linear discriminant analysis (LDA), support vector machine (SVM), and logistic regression (LR)]. Sensitivity, specificity, accuracy, and areas under curves (AUC) of models were calculated, with which the performances of classifiers were evaluated and compared with each other.Result: Brilliant discriminative performance would be observed among all classifiers when combined with the suitable selection method. For LDA-based models, the optimal one was Distance Correlation + LDA with AUC of 0.978. For SVM-based models, Distance Correlation + SVM was the one with highest AUC of 0.959, while for LR-based models, the highest AUC was 0.966 established with LASSO + LR.Conclusion: Radiomics-based machine-learning algorithms potentially have promising performances in differentiating GBM from PCNSL.

ObjectiveTo obtain molecular information in slides directly from H&E staining slides, which apparently display morphological information, to show that some differences in molecular level have already encoded in morphology.MethodsIn this paper, we selected Ki-67-expression as the representative of molecular information. We proposed a method that can predict Ki-67 positive cells directly from H&E stained slides by a deep convolutional network model. To train this model, we constructed a dataset containing Ki-67 negative or positive cell images and background images. These images were all extracted from H&E stained WSIs and the Ki-67 expression was acquired from the corresponding IHC stained WSIs. The trained model was evaluated both on classification performance and the ability to quantify Ki-67 expression in H&E stained images.ResultsThe model achieved an average accuracy of 0.9371 in discrimination of Ki-67 negative cell images, positive cell images and background images. As for evaluation of quantification performance, the correlation coefficient between the quantification results of H&E stained images predicted by our model and that of IHC stained images obtained by color channel filtering is 0.80.Conclusion and SignificanceOur study indicates that the deep learning model has a good performance both on prediction of Ki-67 positive cells and quantification of Ki-67 expression in cancer samples stained by H&E. More generally, this study shows that deep learning is a powerful tool in exploring the relationship between morphological information and molecular information.Availability and ImplementationThe main program is available at https://github.com/liuyiqing2018/predict_Ki-67_from_HE

Rhizoctonia solani AG1 IA is a necrotrophic fungus that causes rice sheath blight, one of the most significant rice diseases in the world. However, little is known about the pathogenic mechanisms and functions of effectors in R. solani AG1 IA. We performed functional studies on effectors in R. solani AG1 IA and found that, of 11 putative effectors tested, only RsIA_NP8 caused necrosis in the leaves of Nicotiana benthamiana. The predicted signal peptide of this protein was required to induce cell death, whereas predicted N-glycosylation sites were not required. RsIA_NP8 was upregulated during early infection, and the encoded protein was secreted. Furthermore, the ability of RsIA_NP8 to trigger cell death in N. benthamiana depended on suppressor of G2 allele of Skp1 (SGT1) and heat shock protein 90 (HSP90), but not on Mla12 resistance (RAR1) and somatic embryogenesis receptor-like kinase (SERK3). A natural variation that prevents the triggering of cell death in N. benthamiana was found in RsIA_NP8 in 25 R. solani AG1 IA strains. It is important to note that RsIA_NP8 induced the immune response in N. benthamiana leaves. Collectively, these results show that RsIA_NP8 is a possible effector that plays a key role in R. solani AG1 IA–host interactions.

Meloxicam (MLX) is a non-steroidal anti-inflammatory drug used to treat rheumatoid arthritis and osteoarthritis. However, its poor water solubility limits the dissolution process and influences absorption. In order to solve this problem and improve its bioavailability, we prepared it in nanocrystals with three different particle sizes to improve solubility and compare the differences between various particle sizes. The nanocrystal particle sizes were studied through dynamic light scattering (DLS) and laser scattering (LS). Transmission electron microscopy (TEM) was used to characterize the morphology of nanocrystals. The sizes of meloxicam-nanocrystals-A (MLX-NCs-A), meloxicam-nanocrystals-B (MLX-NCs-B), and meloxicam-nanocrystals-C (MLX-NCs-C) were 3.262 ± 0.016 μm, 460.2 ± 9.5 nm, and 204.9 ± 2.8 nm, respectively. Molecular simulation was used to explore the distribution and interaction energy of MLX molecules and stabilizer molecules in water. The results of differential scanning calorimetry (DSC) and powder X-ray diffraction (PXRD) proved that the crystalline state did not change in the preparation process. Transport studies of the Caco-2 cell model indicated that the cumulative degree of transport would increase as the particle size decreased. Additionally, plasma concentration–time curves showed that the AUC0–∞ of MLX-NCs-C were 3.58- and 2.92-fold greater than those of MLX-NCs-A and MLX-NCs-B, respectively. These results indicate that preparing MLX in nanocrystals can effectively improve the bioavailability, and the particle size of nanocrystals is an important factor in transmission and absorption.

During the development of a pharmaceutical formulation, a powerful tool is needed to extract the key points from the complicated process parameters and material attributes. Artificial neural networks (ANNs), a promising and more flexible modeling technique, can address real intricate questions in a high parallelism and distributed pattern in the manner of biological neural networks. The data mined and analyzing based on ANNs have the ability to replace hundreds of trial and error experiments. ANNs have been used for data analysis by pharmaceutics researchers since the 1990s and it has now become a research method in pharmaceutical science. This review focuses on the latest application progress of ANNs in the prediction, characterization and optimization of pharmaceutical formulation to provide a reference for the further interdisciplinary study of pharmaceutics and ANNs.

Chitosan is a natural polysaccharide, mainly derived from the shell of marine organisms. At present, chitosan has been widely used in the field of biomedicine due to its special characteristics of low toxicity, biocompatibility, biodegradation and low immunogenicity. Chitosan nanoparticles can be easily prepared. Chitosan nanoparticles with positive charge can enhance the adhesion of antigens in nasal mucosa and promote its absorption, which is expected to be used for intranasal vaccine delivery. In this study, we prepared chitosan nanoparticles by a gelation method, and modified the chitosan nanoparticles with mannose by hybridization. Bovine serum albumin (BSA) was used as the model antigen for development of an intranasal vaccine. The preparation technology of the chitosan nanoparticle-based intranasal vaccine delivery system was optimized by design of experiment (DoE). The DoE results showed that mannose-modified chitosan nanoparticles (Man-BSA-CS-NPs) had high modification tolerance and the mean particle size and the surface charge with optimized Man-BSA-CS-NPs were 156 nm and +33.5 mV. FTIR and DSC results confirmed the presence of Man in Man-BSA-CS-NPs. The BSA released from Man-BSA-CS-NPs had no irreversible aggregation or degradation. In addition, the analysis of fluorescence spectroscopy of BSA confirmed an appropriate binding constant between CS and BSA in this study, which could improve the stability of BSA. The cell study in vitro demonstrated the low toxicity and biocompatibility of Man-BSA-CS-NPs. Confocal results showed that the Man-modified BSA-FITC-CS-NPs promote the endocytosis and internalization of BSA-FITC in DC2.4 cells. In vivo studies of mice, Man-BSA-CS-NPs intranasally immunized showed a significantly improvement of BSA-specific serum IgG response and the highest level of BSA-specific IgA expression in nasal lavage fluid. Overall, our study provides a promising method to modify BSA-loaded CS-NPs with mannose, which is worthy of further study.

Recently, the development of Binder Jet 3D printing technology has promoted the research and application of personalized formulations, which are especially useful for children’s medications. Additionally, physiological pharmacokinetic (PBPK) modeling can be used to guide drug development and drug dose selection. Multiple technologies can be used in combination to increase the safety and effectiveness of drug administration. In this study, we performed in vivo pharmacokinetic experiments in dogs with preprepared 3D-printed levetiracetam instant-dissolving tablets (LEV-IDTs). Bioequivalence analysis showed that the tablets were bioequivalent to commercially available preparations (Spritam®) for dogs. Additionally, we evaluated the bioequivalence of 3D-printed LEV-IDTs with Spritam® by a population-based simulation based on the established PBPK model of levetiracetam for Chinese adults. Finally, we established a PBPK model of oral levetiracetam in Chinese children by combining the physiological parameters of children, and we simulated the PK (pharmacokinetics) curves of Chinese children aged 4 and 6 years that were administered the drug to provide precise guidance on adjusting the dose according to the effective dose range of the drug. Briefly, utilizing both Binder jet 3D printing technology and PBPK models is a promising route for personalized drug delivery with various age groups.