[Pharmacokinetic behavior and brain tissue distribution of paeoniflorin combined with normal and toxic doses of strychnine in rats after percutaneous administration].

This study aims to establish a rapid and sensitive UPLC-MS/MS method for simultaneously determining the content of strychnine and paeoniflorin in plasma and brain tissue of rats, and compare the pharmacokinetic behavior and brain tissue distribution of paeoniflorin combined with normal and toxic doses of strychnine in rats after percutaneous administration. Compared with those in the toxic-dose strychnine group, the AUC_(0-t), AUC_(0-∞), and C_(max) of strychnine decreased by 51.51%, 45.68%, and 46.03%, respectively(P<0.01), and the corresponding values of paeoniflorin increased by 91.41%, 102.31%, and 169.32%, respectively(P<0.01), in the compatibility group. Compared with the normal-dose strychnine group, the compatibility group showed insignificantly decreased C_(max), AUC_(0-t), and AUC_(0-∞) of strychnine, increased C_(max) and T_(max) of paeoniflorin(P<0.01), 66.88% increase in AUC_(0-t), and 70.55% increase in AUC_(0-∞) of paeoniflorin. In addition, the brain tissue concentration of strychnine decreased and that of paeoniflorin increased after compatibility. The combination of paeoniflorin with normal dose and toxic dose of strychnine can inhibit the percutaneous absorption of strychnine, and greatly promote the percutaneous penetration of paeoniflorin, whereas the interaction mechanism remains to be explored. The UPLC-MS/MS method established in this study is easy to operate and has good precision. It is suitable for in vivo study of pharmacokinetic behavior and brain tissue distribution of paeoniflorin and strychnine after percutaneous administration in rats, which provides reference for the safe and rational clinical use of strychnine and the combined use of drugs, and lays a solid foundation for the development of external preparations containing Strychni Semen.

Scalable and fast fabrication of holey multilayer graphene via microwave and its application in supercapacitors.

By virtue of its high specific surface area and low tortuosity for ionic storage and transportation, holey graphene has come to be regarded as a promising material for energy storage devices, such as lithium ion batteries, and supercapacitors. For practical applications, a scalable and green preparation method for holey graphene is required. This work proposes a facile preparation method for holey graphene by simply microwaving pristine graphene in air. Compared with previous scalable methods, this method exhibits much greater efficiency, reducing the preparation time from hours to minutes. The mechanism underlying the microwave irradiation-induced formation of nanosized holes involves the interaction between microwaves, electrons, oxygen in air, and carbon atoms in the defect areas of the graphene. The size, density, and distribution of holes can be controlled by tuning the microwave irradiation time and oxygen concentration. Used as a hybrid conductive agent, the as-prepared holey multilayer graphene increases capacitance retention to 96.25% at high current density (8 A g-1), and 96.48% in long cycles (1 A g-1 and 10 000 cycles).

Temperature sensitive properties and preparation of europium complexes with double ligands.

Temperature sensitive paints (TSPs) are a class of rising materials for non-contact temperature measurement technology. Efficient complexes with remarkable luminescent properties play the core role in TSPs. Rare earth organic complexes are often used as probe molecules for TSPs because of their long fluorescence lifetime, narrow fluorescence emission peaks, high fluorescence intensity, and large Stokes shift. Herein, two europium-based complexes, Eu(PCCA)3 phen and Eu (PMCA)3 phen, were synthesized under hydrothermal conditions, using europium(III) oxide (Eu2 O3 ), p-chlorocinnamic acid (PCCA), p-methoxy cinnamic acid (PMCA) and 1,10-phenanthroline (phen) as raw materials. Two temperature sensitive paints (Eu(PCCA)3 phen/PMMA and Eu(PMCA)3 phen/PMMA) were obtained by the polymerization of methyl methacrylate (MMA) with different europium complexes. The structure, morphology, luminescent properties of the europium complexes and temperature quenching properties of the TSPs were characterized by infrared (IR) spectroscopy, scanning electron microscopy (SEM), and fluorescence spectroscopy. The fluorescence results show that both of the two TSPs have good temperature quenching performance in the range 20-100°C with high sensitivity 50-60°C and 80-90°C, respectively. Furthermore, the highest sensitivity of Eu(PCCA)3 phen/PMMA is greater than that of Eu(PMCA)3 phen/PMMA. This work can provide a universal way for preparing efficient TSPs in practical applications.

The Electric Field Responses of Inorganic Ionogels and Poly(ionic liquid)s.

Ionic liquids (ILs) are a class of pure ions with melting points lower than 100 °C. They are getting more and more attention because of their high thermal stability, high ionic conductivity and dielectric properties. The unique dielectric properties aroused by the ion motion of ILs makes ILs-contained inorganics or organics responsive to electric field and have great application potential in smart electrorheological (ER) fluids which can be used as the electro-mechanical interface in engineering devices. In this review, we summarized the recent work of various kinds of ILs-contained inorganic ionogels and poly(ionic liquid)s (PILs) as ER materials including their synthesis methods, ER responses and dielectric analysis. The aim of this work is to highlight the advantage of ILs in the synthesis of dielectric materials and their effects in improving ER responses of the materials in a wide temperature range. It is expected to provide valuable suggestions for the development of ILs-contained inorganics and PILs as electric field responsive materials.

Characterization of CSF2A fusion gene and its effect on Epstein-Barr virus-positive tumor cells.

We build the latent membrane protein gene latent membrane protein 2A (LMP2A) and the granulocyte-macrophase colony-stimulating factor (GM-CSF) gene fusion gene (CSF2A) and discuss how the CSF2A fusion protein influenced the proliferation and apoptosis of Epstein-Barr virus-positive (EBV+ ) tumor cells. Reverse-transcription polymerase chain reaction (RT-PCR) method was used to amplify the LMP2A gene and GM-CSF gene fragments, respectively, according to the principle of overlap extension in the coding (Gly4Ser)3 polypeptide gene fragments of DNA restructured under the connection. The CSF2A gene could be connected with the pIRES2-enhanced green fluorescent protein vector by recombinant DNA technology and identified by enzyme electrophoresis analysis and DNA sequencing. Then, the recombinant vector was transfected into dendritic cells (DCs); RT-PCR and Western blot analysis were used for testing the CSF2A gene messenger RNA and protein expression. The impacts of CSF2A on the proliferation and apoptosis of EBV+ tumor cells were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Hochest 33342 staining. We successfully obtained the recombinant vector named pIRES2-CSF2A. The expression of CSF2A could be detected by transfecting pIRES2-CSF2A into DCs. The DCs were cocultured with T lymphocytes and then acted on the EBV+ CNE2 nasopharyngeal carcinoma cells. MTT assay showed that the inhibiting effect of CSF2A obviously increased and the time dependency (**P < 0.01, *P < 0.05) also existed. Hochest 33342 staining showed apoptosis morphological changes of cells in nucleus staining and generated the apoptotic body. Apoptosis cells of the pIRES2-CSF2A group increased significantly at 48 hours. The results showed that the pIRES2-CSF2A recombinant vector was effectively transfected into DCs and the fusion gene CSF2A could promote EBV+ CNE2 cell apoptosis, laying the foundation for the specificity of EBV+ tumor targeting immune gene therapy in the future.

Preparation and evaluation of a chitosan-coated antioxidant liposome containing vitamin C and folic acid.

Vitamin C (VC) and folic acid (FA) are the important nutrient and antioxidant in human body. In order to improve their stability, their co-loaded liposomes (VCFA-Lip) and chitosan-coated liposomes (CS-VCFA-Lip) are prepared and characterised. The mean particle size of VCFA-Lip and CS-VCFA-Lip is 138 nm and 249 nm, respectively. The encapsulation efficiencies of both drugs for CS-VCFA-Lip are much higher than those for VCFA-Lip. Furthermore, the experimental results show that the antioxidant activity of CS-VCFA-Lip is higher than that of VCFA-Lip. Moreover, the storage stability study reveals that the chitosan coating can efficiently improve the physical stability of VCFA-Lip. These results indicate that stability of VC and FA can be greatly improved after being wrapped by liposomes. In addition, the performance of CS-VCFA-Lip is better than VCFA-Lip, indicating CS-VCFA-Lip can be applied as a promising delivery system for the antioxidant defence system to the food industry and cosmetic industry.

Transposition of the lingual thyroid gland to the submandibular region through a submandibular approach: A case report.

BACKGROUND: Ectopic thyroid at the base of the tongue is a rare congenital condition, and it is even rarer to have clinical symptoms and require surgical intervention. This disease is easily misdiagnosed preoperatively. This article reports the diagnosis, surgical treatment, and follow-up of a case of lingual thyroid. CASE SUMMARY: The patient was a 54-year-old woman who presented with laryngeal foreign body sensation and dysphagia for 20 d. The lingual thyroid was considered for general examination, and surgery was performed to transpose the lingual thyroid to the right submaxillary region. Pathological analysis confirmed thyroid tissue. The patient experienced complete remission after surgery, but developed hypothyroidism and required thyroid hormone replacement therapy, and her thyroid function gradually recovered over time. CONCLUSION: We report a rare case of lingual thyroid with marked laryngeal foreign body sensation and dysphagia. Symptoms were completely relieved by transposition surgery but postoperative hypothyroidism developed.

Compound collagen peptide powder improves skin photoaging by reducing oxidative stress and activating TGF-ß1/Smad pathway.

Fish collagen peptide (FCP) has been extensively investigated as a natural product that can combat photoaging; however, its efficacy is limited by its singular composition. Compound collagen peptide powder (CCPP) is a novel functional food formulation that exhibits photoprotective properties and comprises FCP and a blend of natural botanical ingredients. The objective of this study was to investigate the efficacy of CCPP and its molecular mechanism. CCPP had a low molecular weight, facilitating its efficient absorption, and was abundant in amino acids, total polyphenols, and total flavonoids. The results of in vivo studies demonstrated that CCPP exhibited significant efficacy in reducing skin wrinkles, enhancing the contents of water and oil in the skin, and ameliorating histopathological alterations in mice. The results of in vitro studies demonstrated that CCPP effectively mitigated photoaging in human skin fibroblasts by attenuating oxidative stress and promoting extracellular matrix (ECM) synthesis. Moreover, we clearly demonstrated that the TGF ß1/Smad pathway was involved in the promotion of ECM synthesis and cell proliferation by CCPP in human skin fibroblasts. These findings suggest that, compared with single collagen, CCPP has a more comprehensive range of antiphotoaging properties.

Exploration of Curvature and Stiffness Dual-Regulated Breast Cancer Cell Motility by a Motor-Clutch Model and Cell Traction Force Characterization.

The migration of breast cancer cells is the main cause of death and significantly regulated by physical factors of the extracellular matrix (ECM). To be specific, the curvature and stiffness of the ECM were discovered to effectively guide cell migration in velocity and direction. However, it is not clear what the extent of effect is when these dual-physical factors regulate cell migration. Moreover, the mechanobiology mechanism of breast cancer cell migration in the molecular level and analysis of cell traction force (CTF) are also important, but there is a lack of systematic investigation. Therefore, we employed a microfluidic platform to construct hydrogel microspheres with an independently adjustable curvature and stiffness as a three-dimensional substrate for breast cancer cell migration. We found that the cell migration velocity was negatively correlated to curvature and positively correlated to stiffness. In addition, curvature was investigated to influence the focal adhesion expression as well as the assignment of F-actin at the molecular level. Further, with the help of a motor-clutch mathematical model and hydrogel microsphere stress sensors, it was concluded that cells perceived physical factors (curvature and stiffness) to cause changes in CTF, which ultimately regulated cell motility. In summary, we employed a theoretical model (motor-clutch) and experimental strategy (stress sensors) to understand the mechanism of curvature and stiffness regulating breast cancer cell motility. These results provide evidence of force driven cancer cell migration by ECM physical factors and explain the mechanism from the perspective of mechanobiology.

TRIM29 modulates proteins involved in PTEN/AKT/mTOR and JAK2/STAT3 signaling pathway and suppresses the progression of hepatocellular carcinoma.

Tripartite motif-containing 29 (TRIM29), also known as the ataxia telangiectasia group D-complementing (ATDC) gene, has been reported to play an oncogenic or tumor suppressive role in developing different tumors. So far, its expression and biological functions in hepatocellular carcinoma (HCC) remain unclear. We investigated TRIM29 expression pattern in human HCC samples using quantitative RT-PCR and immunohistochemistry. Relationships between TRIM29 expression level, clinical prognostic indicators, overall survival (OS), and disease-free survival (DFS) were evaluated by Kaplan-Meier analysis and Cox proportional hazards model. A series of in vitro experiments and a xenograft tumor model were conducted to detect the functions of TRIM29 in HCC cells. RNA sequencing, western blotting, and immunochemical staining were performed to assess the molecular regulation of TRIM29 in HCC. We found that the mRNA and protein levels of TRIM29 were significantly reduced in HCC samples, compared with adjacent noncancerous tissues, and were negatively correlated with poor differentiation of HCC tissues. Survival analysis confirmed that lower TRIM29 expression significantly correlated with shorter OS and DFS of HCC patients. TRIM29 overexpression remarkably inhibited cell proliferation, migration, and EMT in HCC cells, whereas knockdown of TRIM29 reversed these effects. Moreover, deactivation of the PTEN/AKT/mTOR and JAK2/STAT3 pathways might be involved in the tumor suppressive role of TRIM29 in HCC. Our findings indicate that TRIM29 in HCC exerts its tumor suppressive effects through inhibition of the PTEN/AKT/mTOR and JAK2/STAT3 signaling pathways and may be used as a potential biomarker for survival in patients with HCC.

Integration of microbiomics, metabolomics, and transcriptomics reveals the therapeutic mechanism underlying Fuzheng-Qushi decoction for the treatment of lipopolysaccharide-induced lung injury in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Fuzheng-Qushi decoction (FZQS) is a practical Chinese herbal formula for relieving cough and fever. Therefore, the action and specific molecular mechanism of FZQS in the treatment of lung injury with cough and fever as the main symptoms need to be further investigated. AIMS OF THE STUDY: To elucidate the protective effects of FZQS against lung injury in mice and reveal its potential targets and key biological pathways for the treatment of lung injury based on transcriptomics, microbiomics, and untargeted metabolomics analyses. MATERIALS AND METHODS: Lipopolysaccharide (LPS) was used to induce a mouse model of lung injury, followed by the administration of FZQS. ELISA was used to detect IL-1ß, IL-6, IL-17A, IL-4, IL-10, and TNF-α, in mouse lung tissues. Macrophage polarization and neutrophil activation were measured by flow cytometry. RNA sequencing (RNA-seq) was applied to screen for differentially expressed genes (DEGs) in lung tissues. RT-qPCR and Western blot assays were utilized to validate key DEGs and target proteins in lung tissues. 16S rRNA sequencing was employed to characterize the gut microbiota of mice. Metabolites in the gut were analyzed using untargeted metabolomics. RESULTS: FZQS treatment significantly ameliorated lung histopathological damage, decreased pro-inflammatory cytokine levels, and increased anti-inflammatory cytokine levels. M1 macrophage levels in the peripheral blood decreased, M2 macrophage levels increased, and activated neutrophils were inhibited in mice with LPS-induced lung injury. Importantly, transcriptomic analysis showed that FZQS downregulated macrophage and neutrophil activation and migration and adhesion pathways by reversing 51 DEGs, which was further confirmed by RT-qPCR and Western blot analysis. In addition, FZQS modulated the dysbiosis of the gut microbiota by reversing the abundance of Corynebacterium, Facklamia, Staphylococcus, Paenalcaligenes, Lachnoclostridium, norank_f_Muribaculaceae, and unclassified_f_Lachnospiraceae. Meanwhile, metabolomics analysis revealed that FZQS significantly regulated tryptophan metabolism by reducing the levels of 3-Indoleacetonitrile and 5-Hydroxykynurenine. CONCLUSION: FZQS effectively ameliorated LPS-induced lung injury by inhibiting the activation, migration, and adhesion of macrophages and neutrophils and modulating gut microbiota and its metabolites.

Dysregulated NOX1-NOS2 activity as hallmark of ileitis in mice.

Inflammation of the ileum, or ileitis, is commonly caused by Crohn's disease (CD) but can also accompany ulcerative colitis (backwash ileitis), infections or drug-related damage. Oxidative tissue injury triggered by reactive oxygen species (ROS) is considered part of the ileitis etiology. However, not only elevated ROS but also permanently decreased ROS are associated with inflammatory bowel disease (IBD). While very early onset IBD (VEO-IBD) is associated with a spectrum of NOX1 variants, how NOX1 inactivation contributes to disease development remains ill-defined. Besides propagating signaling responses, NOX1 provides superoxide for peroxynitrite formation in the epithelial barrier. Here we report that NOX4, an H2O2-generating NADPH oxidase with documented tissue protective effects in the intestine and other tissues, limits the generation of ileal peroxynitrite by NOX1/NOS2. Deletion of NOX4 leads to persistent peroxynitrite excess, hyperpermeability, villus blunting, muscular hypertrophy, chemokine/cytokine upregulation and dysbiosis. Conversely, SAMP1/YitFc mice, a CD-like ileitis model, showed age-dependent NOX1/NOS2 downregulation preventing ileal peroxynitrite formation in homeostasis and LPS-induced acute inflammation. Deficiency in NOX1 correlated with the upregulation of antimicrobial peptides, suggesting that ileal peroxynitrite acts as chemical barrier and microbiota modifier in the ileum.

Developing a Pocket Park Prescription Program for Human Restoration: An Approach That Encourages Both People and the Environment.

Healing through nature has long been confirmed as an efficient way to improve human physical and psychological health in contemporary urban life. This concept evolved into the well-known Park Prescription Program. However, the psychological restoration imparted by nature was not particularly emphasized in the original Park Prescription Program; it primarily addresses the regulation of physical activities. The quality of urban parks may affect how well people pursue these prescriptions, but the program rarely includes designers among its stakeholders. This study is inspired by the Park Prescription Program, and its intent is to develop a Pocket Park Prescription Program that encourages usage by active people and proper landscape design. The inclusion of designers has been found to be extremely important for pocket parks since they are limited in size but have the advantages of high flexibility and accessibility, and their restorative potential needs be maximized with the limited resources available. Ten pocket parks with distinct functional and landscape attributes were selected in Shanghai as research sites. The Restorative Component Scale was designed into a questionnaire-based survey to measure how people perceived restorative experiences in each site. The onsite survey also incorporated questions investigating people's behaviour characteristics of using these pocket parks. Site photos were taken and analysed with semantic image segmentation to indicate the landscape compositions of each site. The relations between people perceived restorativeness and parks' using patterns and landscape characteristics were then explored with correlation analysis to provide cues on instructing how people's visiting behavior and park landscape design can be improved. The results suggest there are better restoration results when people stay longer in pocket parks, and when people visit their neighborhood pocket parks two to three times a week, these benefits are further enhanced. This study also found that when these prescribed health behaviors are uncertain, the restorative experiences perceived by people can be improved with design interventions in regard to landscape elements such as vegetation, person, decorative lamps, pavement and terrain. These interventions should be made also in consideration of specific park functions. The research outcome intends to show that designers should be included as stakeholders in the Pocket Park Prescription Program, and it is expected to guide individuals towards effectively using pocket parks for restoration. This is to ensure that both the design and the people's perspectives will be strengthened through the implementation of this program.

The effects of the natural visual-aural attributes of urban green spaces on human behavior and emotional response.

Introduction: Nature-based solutions (NBS) have been used to address a wide range of urban environmental challenges, an important aspect of which is to improve human health and well-being. However, most relevant studies focus either on what positive influences nature may have or on identifying what natural factors can have these benefits. Few have investigated the sensory composition of nature and the effects of nature in different sensory aspects on human health. Setting out from the multi-sensory perspective, this study aims to explore human behavior and emotional response from visual and aural contact with urban nature. Methods: Taking Jiangjia Art Garden in Chengdu as an example, natural attributes such as its visual (landscape) and aural (sound source) characteristics as well as people's activities (behavioral responses) were measured by on-site mapping analysis. This was done while a questionnaire-based survey was conducted to investigate people's emotional responses regarding their overall satisfaction, pleasantness, calm, and agreeableness. Results: The results indicated that nature-dominated visual landscapes such as grassland, waterscapes, and woodlands, as well as natural sounds such as bird sounds, chirp sounds, and wind sounds were found to be positively correlated to the vitality of activities and people's emotional status. Regarding behavioral responses, it was shown that grasslands and woodlands are more likely to be attractive places for recreation, and the vitality measured became extremely high when these two were paired with lakes. As for the emotional responses, people's perceived overall satisfaction, calm, and agreeableness were equally reflected in their behavioral patterns, suggesting a strong relationship with natural factors. Discussion: The research findings were visually presented in behavior and emotional maps to provide direct cues of informing the future design of high-quality urban green spaces and promoting the application of aural-visual experience in the design of urban nature areas.

Ezetimibe Induces Paraptosis through Niemann-Pick C1-like 1 Inhibition of Mammalian-Target-of-Rapamycin Signaling in Hepatocellular Carcinoma Cells.

Currently, hepatocellular carcinoma (HCC) is characterized by its unfavorable prognosis and resistance to conventional chemotherapy and radiotherapy. Drug repositioning, an approach aimed at identifying novel therapeutic applications for existing drugs, presents a cost-effective strategy for developing new anticancer agents. We explored the anticancer properties of Ezetimibe, a widely used oral lipid-lowering drug, in the context of HCC. Our findings demonstrate that Ezetimibe effectively suppresses HCC cell proliferation through paraptosis, an apoptotic-independent cell death pathway. The examination of HCC cells lines treated with Ezetimibe using light microscopy and transmission electron microscopy (TEM) showed cytoplasmic vacuolation in the perinuclear region. Notably, the nuclear membrane remained intact in both Ezetimibe-treated and untreated HCC cell lines. Probe staining assays confirmed that the cytoplasmic vacuoles originated from dilated endoplasmic reticulum (ER) compartments rather than mitochondria. Furthermore, a dose-dependent accumulation of reactive oxygen species (ROS) was observed in Ezetimibe-treated HCC cell lines. Co-treatment with the general antioxidant NAC attenuated vacuolation and improved cell viability in Ezetimibe-treated HCC cells. Moreover, Ezetimibe induced paraptosis through proteasome activity inhibition and initiation of the unfolded protein response (UPR) in HCC cell lines. In our in vivo experiment, Ezetimibe significantly impeded the growth of HCC tumors. Furthermore, when combined with Sorafenib, Ezetimibe exhibited a synergistic antitumor effect on HCC cell lines. Mechanistically, Ezetimibe induced paraptosis by targeting NPC1L1 to inhibit the PI3K/AKT/mTOR signaling pathway. In conclusion, our study highlights the potential of Ezetimibe as an anticancer agent by triggering paraptosis in HCC cells.

VEO-IBD NOX1 variant highlights a structural region essential for NOX/DUOX catalytic activity.

Inflammatory bowel diseases (IBD) are chronic intestinal disorders that result from an inappropriate inflammatory response to the microbiota in genetically susceptible individuals, often triggered by environmental stressors. Part of this response is the persistent inflammation and tissue injury associated with deficiency or excess of reactive oxygen species (ROS). The NADPH oxidase NOX1 is highly expressed in the intestinal epithelium, and inactivating NOX1 missense mutations are considered a risk factor for developing very early onset IBD. Albeit NOX1 has been linked to wound healing and host defence, many questions remain about its role in intestinal homeostasis and acute inflammatory conditions. Here, we used in vivo imaging in combination with inhibitor studies and germ-free conditions to conclusively identify NOX1 as essential superoxide generator for microbiota-dependent peroxynitrite production in homeostasis and during early endotoxemia. NOX1 loss-of-function variants cannot support peroxynitrite production, suggesting that the gut barrier is persistently weakened in these patients. One of the loss-of-function NOX1 variants, NOX1 p. Asn122His, features replacement of an asparagine residue located in a highly conserved HxxxHxxN motif. Modelling the NOX1-p22phox complex revealed near the distal heme an internal pocket restricted by His119 and Asn122 that is part of the oxygen reduction site. Functional studies in several human NADPH oxidases show that substitution of asparagine with amino acids with larger side chains is not tolerated, while smaller side chains can support catalytic activity. Thus, we identified a previously unrecognized structural feature required for the electron transfer mechanism in human NADPH oxidases.

PD1hi CD200hi CD4+ exhausted T cell increase immunotherapy resistance and tumour progression by promoting epithelial-mesenchymal transition in bladder cancer.

BACKGROUND: Bladder cancer (BLCA) is one of the most diagnosed cancers in humans worldwide. Recently, immunotherapy has become a main treatment option for BC. However, most BLCA patients do not respond to immune checkpoint inhibitors or relapse after immunotherapy. Therefore, it is very important to identify novel biomarkers for the prediction of immunotherapy response in B patients. METHODS: Pancancer single-cell RNA sequencing (scRNA-seq) data were used to identify the clusters of CD4+ T cells in the tumour microenvironment (TME). The clinical significance of key CD4+ T-cell clusters was evaluated based on the survival data of two independent immunotherapy bladder cancer (BLCA) cohorts. We also investigated the function of key clusters of CD4+ T cell in the TME of BC cells in vitro. RESULTS: This study identified two novel exhausted CD4+ T-cell subpopulations with the expression of PD1hi CD200hi or PD1hi CD200low in BC patients. Moreover, BLCA patients with a high level of PD1hi CD200hi CD4+ exhausted T cell showed immunotherapy resistance. Cell function analysis demonstrated that PD1hi CD200hi CD4+ exhausted T cell can promote epithelial-mesenchymal transition (EMT) and angiogenesis in BLCA cells. In addition, PD1hi CD200hi CD4+ exhausted T cells were shown to communicate with malignant BLCA cells through the GAS6-AXL axis. Finally, we also found that GAS6 expression is upregulated in B cells by METTL3-mediated m6A modification. CONCLUSIONS: PD1hi CD200hi CD4+ exhausted T cell may serve as a novel biomarker for poor prognosis and immunotherapy resistance in B. Targeted inhibitors of PD1hi CD200hi CD4+ exhausted T cells may help improve the efficacy of immunotherapy.

Pien Tze Huang alleviates Concanavalin A-induced autoimmune hepatitis by regulating intestinal microbiota and memory regulatory T cells.

BACKGROUND: Traditional Chinese medicine has used the drug Pien Tze Huang (PTH), a classic prescription, to treat autoimmune hepatitis (AIH). However, the precise mode of action is still unknown. AIM: To investigate the mechanism of PTH in an AIH mouse model by determining the changes in gut microbiota structure and memory regulatory T (mTreg) cells functional levels. METHODS: Following induction of the AIH mouse model induced by Concanavalin A (Con A), prophylactic administration of PTH was given for 10 d. The levels of mTreg cells were measured by flow cytometry, and intestinal microbiota was analyzed by 16S rRNA analysis, while western blotting was used to identify activation of the toll-like receptor (TLR)2, TLR4/nuclear factor-κB (NF-κB), and CXCL16/CXCR6 signaling pathways. RESULTS: In the liver of mice with AIH, PTH relieved the pathological damage and reduced the numbers of T helper type 17 cells and interferon-γ, tumor necrosis factor-alpha, interleukin (IL)-1ß, IL-2, IL-6, and IL-21 expression. Simultaneously, PTH stimulated the abundance of helpful bacteria, promoted activation of the TLR2 signal, which may enhance Treg/mTreg cells quantity to produce IL-10, and suppressed activation of the TLR4/NF-κB and CXCL16/CXCR6 signaling pathways. CONCLUSION: PTH regulates intestinal microbiota balance and restores mTreg cells to alleviate experimental AIH, which is closely related to the TLR/CXCL16/CXCR6/NF-κB signaling pathway.

High density and proximity of CD8+ T cells to tumor cells are correlated with better response to nivolumab treatment in metastatic pleural mesothelioma.

BACKGROUND: The efficacy of immune checkpoint inhibitors (ICIs) in pleural mesothelioma has recently been established. The response to ICIs can be predicted by quantitative analysis of cells and their spatial distribution in the tumor microenvironment (TME). However, the detailed composition of the TME in pleural mesothelioma has not been reported. We evaluated the association between the TME and response to ICIs in this cancer. METHODS: A retrospective analysis of 22 pleural mesothelioma patients treated with nivolumab in different centers was performed using surgical specimens. Four patients had a partial response to nivolumab (response group) and 18 patients had stable or progressive disease (nonresponse group). The number of CD4, CD8, FoxP3, CK, and PD-L1 positive cells, cell density, and cell-to-cell distance were analyzed by multiplex immunofluorescence. RESULTS: PD-L1 expression did not differ significantly between the response and nonresponse groups. The density of total T cells and of CD8+ T cells was significantly higher in the response than in the nonresponse group. CD8+ T cells were more clustered and located closer to tumor cells, whereas regulatory T cells were located further from tumor cells in the response than in the nonresponse group. CONCLUSIONS: High density and spatial proximity of CD8+ T cells to tumor cells were associated with better response to nivolumab, whereas the proximity of regulatory T cells to tumor cells was associated with worse response, suggesting that the distinct landscape of the TME could be a potential predictor of ICI efficacy in pleural mesothelioma.

[Research progress in biological basis of cold and heat essence of Chinese medicine].

Cold-heat problem is one core of traditional Chinese medicine theory. This paper summarizes the experimental research related to the biological basis of cold-heat essence in cold-heat syndrome, cold-heat body constitution and cold-heat property of Chinese herbs. In view of the classical physiological and biochemical indices, gene expression, protein expression and metabolic differences, differences in cold-heat syndrome or cold-heat constitution are mainly based on neurotransmitter, thyroid function, sex hormone, cyclic nucleotide system, and energy metabolism relating to the corresponding gene and protein expression. Furthermore, this paper analyses the change of correlation indices that accompany with a dynamic development process of "constitution-syndrome-herbal intervention", implying that the research of biological basis of cold-heat essence has turned from single index to multiple indices, and from dispersion research to system research.