BACKGROUND: Naloxone has a significant arousal effect on many types of comas. It is usually believed that this is because its inhibition on endogenous opioid peptides. But depth of coma is not necessarily positively correlated to endorphin (EP).OBJECTIVE: Based on existing findings on direct stimulating effect of naloxone on cerebral cortex, further studies need to be done to explore whether it is dose-dependent or not.DESIGN: Single-factor design based on cells.SETTING: Neurology department in a university hospital and the neurology department in a hospital of a military medical university of Chinese PLA.MATERIALS: This study was completed in the Laboratory Center of Tongji Medical College, Huazhong University of Science and Technology. Thirty healthy new born Wistar rats, regardless of their gender, aging 8 - 12 days and weighing 150 -250 g, were selected.METHODS: The experiment was performed at room temperature. The perfusion slot were placed on the microscope stage, and cells with smooth surfaces, triangle or pyramidal shapes, strong refraction and more than one neurites were selected for patch clamp experiment. Patch clamp whole-cell recording technique was used to measure the pyramidal cells of the frontal lobe immediately after separated from the Wistar rats, and to investigate the fluctuations of their membrane potential of cerebral cortex neurons and the frequencies of their spontaneous electric activities after administration of naloxone at different doses.MAIN OUTCOME MEASURES: The neural excitatory reaction rate, depolarization amplitude and increasing rate of spontaneous electric activities after administration of different doses of naloxone were selected as main outcome measurements.RESULTS: The excitatory reaction rates of cerebral cortex neurons immediately after separation to doses of naloxone(100, 50, 10, 1, 0. 1 μmol/L)were 83%, 67%, 86%, 71% and 33%; while the depolarization amplitude of them were 9. 8, 9.6, 8.4, 5.2 and 1. 3 mV respectively; and the corresponding spontaneous electric activity were increased by 587% , 375% ,291%, 125% and 69%.CONCLUSION: Naloxone can induce excitatory reactions in cerebral cortex neurons directly, and the reactions have proved to be dose-dependent.

ObjectiveTo investigate the heterogeneity and transcriptomic characteristics of T-cell subsets in the liver of mice with nonalcoholic steatohepatitis （NASH） at the single-cell level using single-cell RNA sequencing （scRNA-seq）， and to provide a reference for studying the mechanism of action of T cells in NASH. MethodsSix male C57BL/6 mice were randomly divided into control group fed with regular diet and NASH group fed with methionine-choline-deficient （MCD） diet， with three mice in each group， and liver tissue was collected for scRNA-seq after 6 weeks of modeling. Specific differentially expressed genes were analyzed between T-cell subsets， and related analyses were performed， including dimension clustering， cell type annotation， t-distributed stochastic neighbor embedding （t-SNE）， violin plot， gene ontology （GO） functional enrichment analysis， and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis. Immunofluorescent staining was used to observe the expression of the T cell marker Tcrα and the specific marker genes Tcf7 and Cxcr6 in the liver of mice in the two groups. The independent-samples t test was used for comparison of continuous data between two groups. ResultsTwo T cell subsets were identified in the liver of mice， and the percentage of cluster 6 decreased from 58.5% in the control group to 48.7% in the NASH group. The top four specific genes were Nsg2， Cd8b1， Cd8a， and Tcf7. Tcf7， a characteristic marker gene for cluster 6， was expressed in 65% of cells in cluster 6， and therefore， cluster 6 was defined as Tcf7⁺ T cells. The GO and KEGG enrichment analyses showed that the differentially expressed genes of cluster 6 were involved in T cell activation， leukocyte adhesion， binding ubiquitin-like protein ligase， and the signaling pathways for Th17， Th1， and Th2 cell differentiation. The percentage of cluster 7 increased from 41.5% in the control group to 51.3% in the NASH group. The top four specific genes of cluster 7 were Cd40lg， Tcrg-C1， Il2rα， and Cxcr6. Cxcr6 was expressed in 90% of cells in cluster 7， and therefore， cluster 7 was defined as Cxcr6⁺ T cells. The GO and KEGG enrichment analyses showed that cluster 7 was involved in T cell activation， cytokine production， the T cell receptor signaling pathway， and the Th17 cell differentiation and MAPK signaling pathway. Immunofluorescence assay showed that compared with the control group， the NASH group showed a significant reduction in the area with positive co-expression of Tcf7 protein and Tcrα protein （1.80%±0.67% vs 0.33%±0.13%， P<0.05） and a significant increase in the area with positive co-expression of Cxcr6 protein and Tcrα protein （0.50%±0.09% vs 2.66%± 0.33%， P<0.001）. ConclusionThere is a reduction in the percentage of Tcf7⁺ T cells and an increase in the percentage of Cxcr6⁺ T cells in NASH mice， revealing the characteristics and differences of T cells in the liver of NASH mice.

Objective: The TCGA molecular subtype of endometrial cancer (EC) is widely applied, among which the copy-number high (CNH) subtype has the poorest prognosis. However, the heterogeneity of this subtype remains elusive. In this study, we aimed to identify heterogeneous immune subtypes in CNH EC and explore their prognostic significance. Methods: We collected 60 CNH EC cases in the TCGA database and performed unsupervised cluster analysis based on the enrichment scores of immune-related gene signatures to identify immune subtypes. We described their immune characteristics and prognoses and conducted differential gene analysis and lasso regression to identify a prognostic biomarker, GZMM. For experimental validation, we performed immunohistochemical staining of GZMM in 39 p53-positive EC surgical samples. Results: We defined two immune subtypes, immune-hot (IH) and immune-cold (IC), which differed in immune cell infiltration, cytokine and chemokine expression and prognosis. The IH subtype has significantly stronger immune activation than the IC subtype, showing a significant infiltration of immune effector cells and high expression of relevant chemokines, with better prognosis. Moreover, the immunohistochemical staining of GZMM in a cohort of 39 p53-positive EC surgical samples confirmed GZMM as a unique prognostic biomarker, with high expression in both tumor cells and lymphocytes predicting a better prognosis. Conclusion Our study revealed heterogeneous immune subtypes in CNH EC and identified GZMM as a prognostic biomarker. The stratified classification strategy combining molecular and immune subtypes provides valuable insights for future clinical practice.

Objective: The TCGA molecular subtype of endometrial cancer (EC) is widely applied, among which the copy-number high (CNH) subtype has the poorest prognosis. However, the heterogeneity of this subtype remains elusive. In this study, we aimed to identify heterogeneous immune subtypes in CNH EC and explore their prognostic significance. Methods: We collected 60 CNH EC cases in the TCGA database and performed unsupervised cluster analysis based on the enrichment scores of immune-related gene signatures to identify immune subtypes. We described their immune characteristics and prognoses and conducted differential gene analysis and lasso regression to identify a prognostic biomarker, GZMM. For experimental validation, we performed immunohistochemical staining of GZMM in 39 p53-positive EC surgical samples. Results: We defined two immune subtypes, immune-hot (IH) and immune-cold (IC), which differed in immune cell infiltration, cytokine and chemokine expression and prognosis. The IH subtype has significantly stronger immune activation than the IC subtype, showing a significant infiltration of immune effector cells and high expression of relevant chemokines, with better prognosis. Moreover, the immunohistochemical staining of GZMM in a cohort of 39 p53-positive EC surgical samples confirmed GZMM as a unique prognostic biomarker, with high expression in both tumor cells and lymphocytes predicting a better prognosis. Conclusion Our study revealed heterogeneous immune subtypes in CNH EC and identified GZMM as a prognostic biomarker. The stratified classification strategy combining molecular and immune subtypes provides valuable insights for future clinical practice.

Objective: The TCGA molecular subtype of endometrial cancer (EC) is widely applied, among which the copy-number high (CNH) subtype has the poorest prognosis. However, the heterogeneity of this subtype remains elusive. In this study, we aimed to identify heterogeneous immune subtypes in CNH EC and explore their prognostic significance. Methods: We collected 60 CNH EC cases in the TCGA database and performed unsupervised cluster analysis based on the enrichment scores of immune-related gene signatures to identify immune subtypes. We described their immune characteristics and prognoses and conducted differential gene analysis and lasso regression to identify a prognostic biomarker, GZMM. For experimental validation, we performed immunohistochemical staining of GZMM in 39 p53-positive EC surgical samples. Results: We defined two immune subtypes, immune-hot (IH) and immune-cold (IC), which differed in immune cell infiltration, cytokine and chemokine expression and prognosis. The IH subtype has significantly stronger immune activation than the IC subtype, showing a significant infiltration of immune effector cells and high expression of relevant chemokines, with better prognosis. Moreover, the immunohistochemical staining of GZMM in a cohort of 39 p53-positive EC surgical samples confirmed GZMM as a unique prognostic biomarker, with high expression in both tumor cells and lymphocytes predicting a better prognosis. Conclusion Our study revealed heterogeneous immune subtypes in CNH EC and identified GZMM as a prognostic biomarker. The stratified classification strategy combining molecular and immune subtypes provides valuable insights for future clinical practice.

Gestational trophoblastic neoplasia (GTN) patients can be classified into low-risk and high-risk patients based on the International Federation of Gynecology and Obstetrics(FIGO) staging and scoring system. High-risk patients have a higher risk of resistance to single-agent chemotherapy and require combination chemotherapy. Even worse, there are some high-risk patients who develop resistance to combination chemotherapy or experience recurrence after cure. These patients exhibit strong heterogeneity and pose significant challenges to subsequent treatment, thus attracting considerable attention in recent years. Therefore, this article aims to review the risk factors and treatment strategies for high-risk resistant and recurrent GTN patients, in order to provide a basis for early identification of high-risk patients and their precise treatment.