Vasoprotective Effects of Hyperoside against Cerebral Ischemia/Reperfusion Injury in Rats: Activation of Large-Conductance Ca2+-Activated K+ Channels.

Hyperoside (Hyp), a kind of Chinese herbal medicine, exerts multiple therapeutic effects on many diseases. However, the role and mechanisms of Hyp in vascular pathophysiology in ischemic stroke need to be further established. The study aimed to investigate the role of (large-conductance Ca2+-activated K+) BK channels on the vasoprotection of Hyp against cerebral ischemia and reperfusion (I/R) injury in rats. The concentration gradient of Hyp was pretreated in both the middle cerebral artery occlusion and reperfusion model and oxygen-glucose deprivation/reoxygenation (OGD/R) model of primary vascular smooth muscle cells (VSMCs) in rats. A series of indicators were detected, including neurological deficit score, infarct volume, malondialdehyde (MDA), superoxide dismutase (SOD), cerebral blood flow (CBF), cell viability, membrane potential, and BK channels α- and ß1-subunits expression. The results showed that Hyp significantly reduced infarct volume and ameliorated neurological dysfunction in I/R-injured rats. Besides, the effects of I/R-induced reduction of BK channels α- and ß1-subunits expression were significantly reversed by Hyp in endothelial-denudated cerebral basilar arteries. Furthermore, the protective effect against I/R-induced increases of MDA and reduction of SOD as well as CBF induced by Hyp was significantly reversed by iberiotoxin (IbTX). In OGD/R-injured VSMCs, downregulated cellular viability and BK channels ß1-subunits expression were remarkably reversed by Hyp. However, neither OGD/R nor Hyp affected BK channels α-subunits expression, and Hyp failed to induced hyperpolarization of VSMCs. Moreover, the protective effect against OGD/R-induced reduction of cell viability and SOD level and increases of MDA production induced by Hyp was significantly reversed by IbTX in VSMCs. The study indicates that Hyp has the therapeutic potential to improve vascular outcomes, and the mechanism is associated with suppressing oxidative stress and improving CBF through upregulating BK channels.

The phospholipase effector Tle1Vc promotes Vibrio cholerae virulence by killing competitors and impacting gene expression.

Vibrio cholerae utilizes the Type VI secretion system (T6SS) to gain an advantage in interbacterial competition by delivering anti-prokaryotic effectors in a contact-dependent manner. However, the impact of T6SS and its secreted effectors on physiological behavior remains poorly understood. In this study, we present Tle1Vc, a phospholipase effector in atypical pathogenic V. cholerae E1 that is secreted by T6SS via its interaction with VgrG1E1. Tle1Vc contains a DUF2235 domain and belongs to the Tle1 (type VI lipase effector) family. Bacterial toxicity assays, lipase activity assays and site-directed mutagenesis revealed that Tle1Vc possessed phospholipase A1 activity and phospholipase A2 activity, and that Tle1Vc-induced toxicity required a serine residue (S356) and two aspartic acid residues (D417 and D496). Cells intoxication with Tle1Vc lead to membrane depolarization and alter membrane permeability. Tli1tox-, a cognate immunity protein, directly interacts with Tle1Vc to neutralize its toxicity. Moreover, Tle1Vc can kill multiple microorganisms by T6SS and promote in vivo fitness of V. cholerae through mediating antibacterial activity. Tle1Vc induces bacterial motility by increasing the expression of flagellar-related genes independently of functional T6SS and the tit-for-tat (TFT) response, where Pseudomonas aeruginosa uses its T6SS-H1 cluster to counterattack other offensive attackers. Our study also demonstrated that the physical puncture of E1 T6SS can induce a moderate TFT response, which is essential to the Tle1Vc-mediated strong TFT response, maximizing effector functions. Overall, our study characterized the antibacterial mechanism of phospholipase effector Tle1Vc and its multiple physiological significance.

Pesticin-Like Effector VgrG3cp Targeting Peptidoglycan Delivered by the Type VI Secretion System Contributes to Vibrio cholerae Interbacterial Competition.

Vibrio cholerae can utilize a type VI secretion system (T6SS) to increase its intra- and interspecies competition. However, much still remains to be understood about the underlying mechanism of this intraspecies competition. In this study, we isolated an environmental V. cholerae strain E1 that lacked the typical virulence factors toxin-coregulated pilus and cholera toxin and that encoded a functional T6SS. We identified an evolved VgrG3 variant with a predicted C-terminal pesticin-like domain in V. cholerae E1, designated VgrG3cp. Using heterologous expression, protein secretion, and peptidoglycan-degrading assays, we demonstrated that VgrG3cp is a T6SS-dependent effector harboring cell wall muramidase activity and that its toxicity can be neutralized by cognate immunity protein TsiV3cp. Site-directed mutagenesis proved that the aspartic acid residue at position 867 is crucial for VgrG3cp-mediated antibacterial activity. Bioinformatic analysis showed that genes encoding VgrG3cp-like homologs are distributed in Vibrio species, are linked with T6SS structural genes and auxiliary genes, and the vgrG3cp-tsiV3cp gene pair of V. cholerae probably evolved from Vibrio anguillarum and Vibrio fluvialis via homologous recombination. Through a time-lapse microscopy assay, we directly determined that cells accumulating VgrG3cp disrupted bacterial division, while the cells continued to increase in size until the loss of membrane potential and cell wall breakage and finally burst. The results of the competitive killing assay showed that VgrG3cp contributes to V. cholerae interspecies competition. Collectively, our study revealed a novel T6SS E-I pair representing a new T6SS toxin family which allows V. cholerae to gain dominance within polymicrobial communities by T6SS. IMPORTANCE The type VI secretion system used by a broad range of Gram-negative bacteria delivers toxic proteins to target adjacent eukaryotic and prokaryotic cells. Diversification of effector proteins determines the complex bacterium-bacterium interactions and impacts the health of hosts and environmental ecosystems in which bacteria reside. This work uncovered an evolved valine-glycine repeat protein G3, carrying a C-terminal pesticin-like domain (VgrG3cp), which has been suggested to harbor cell wall hydrolase activity and is able to affect cell division and the integrity of cell wall structure. Pesticin-like homologs constitute a family of T6SS-associated effectors targeting bacterial peptidoglycan which are distributed in Vibrio species, and genetic loci of them are linked with T6SS structural genes and auxiliary genes. T6SS-delivered VgrG3cp mediated broad-spectrum antibacterial activity for several microorganisms tested, indicating that VgrG3cp-mediated antimicrobial activity is capable of conferring bacteria a competitive advantage over competitors in the same niches.

Single cell sequencing revealed the mechanism of CRYAB in glioma and its diagnostic and prognostic value.

Background: We explored the characteristics of single-cell differentiation data in glioblastoma and established prognostic markers based on CRYAB to predict the prognosis of glioblastoma patients. Aberrant expression of CRYAB is associated with invasive behavior in various tumors, including glioblastoma. However, the specific role and mechanisms of CRYAB in glioblastoma are still unclear. Methods: We assessed RNA-seq and microarray data from TCGA and GEO databases, combined with scRNA-seq data on glioma patients from GEO. Utilizing the Seurat R package, we identified distinct survival-related gene clusters in the scRNA-seq data. Prognostic pivotal genes were discovered through single-factor Cox analysis, and a prognostic model was established using LASSO and stepwise regression algorithms. Moreover, we investigated the predictive potential of these genes in the immune microenvironment and their applicability in immunotherapy. Finally, in vitro experiments confirmed the functional significance of the high-risk gene CRYAB. Results: By analyzing the ScRNA-seq data, we identified 28 cell clusters representing seven cell types. After dimensionality reduction and clustering analysis, we obtained four subpopulations within the oligodendrocyte lineage based on their differentiation trajectory. Using CRYAB as a marker gene for the terminal-stage subpopulation, we found that its expression was associated with poor prognosis. In vitro experiments demonstrated that knocking out CRYAB in U87 and LN229 cells reduced cell viability, proliferation, and invasiveness. Conclusion: The risk model based on CRYAB holds promise in accurately predicting glioblastoma. A comprehensive study of the specific mechanisms of CRYAB in glioblastoma would contribute to understanding its response to immunotherapy. Targeting the CRYAB gene may be beneficial for glioblastoma patients.

Comparable outcomes in patients with B-cell acute lymphoblastic leukemia receiving haploidentical hematopoietic stem cell transplantation: Pretransplant minimal residual disease-negative complete remission following chimeric antigen receptor T-cell therapy versus chemotherapy.

Introduction: Chimeric antigen receptor (CAR) T-cell (CAR-T) therapy followed by haploidentical hematopoietic stem cell transplantation (haplo-HSCT) markedly improves the long-term survival of patients with refractory/relapsed (R/R) B-cell acute lymphoblastic leukemia (B-ALL). Methods: We performed a parallel comparison of transplant outcomes in 168 B-ALL patients undergoing haplo-HSCT after achieving minimal residual disease (MRD)-negative complete remission (CR) from CAR-T therapy (n = 28) or chemotherapy (n = 140) between January 2016 and August 2021. We further divided the chemotherapy group into the first CR group (chemo+CR1, n = 118) and a second or more CR group (chemo+≥CR2, n = 22). Results: With a median follow-up period of 31.0 months, the 2-year overall survival (OS), leukemia-free survival (LFS), non-relapse mortality (NRM), and relapse rates in the CAR-T and chemotherapy groups did not differ significantly (OS, 87.9% vs. 71.5 %; LFS, 72.0% vs. 66.8%; NRM, 3.9% vs. 13.7%; relapse, 24.1% vs. 19.4%). Multivariate analysis confirmed that ≥CR2 at transplantation following chemotherapy was an independent risk factor associated with poor OS (hazard ratio (HR) 4.22 [95% CI, 1.34-13.293], p = 0.014) and LFS (HR 2.57 [95% CI, 1.041-6.343], p = 0.041). The probabilities of OS and LFS at 2 years in the CAR-T group were comparable to those in the chemo+CR1 group but significantly higher than those in the chemo+≥CR2 group (OS, 87.9% vs. 37.8%, p = 0.007; LFS, 72.0% vs. 41.7%, p = 0.043). No significant differences in the incidences of NRM were noted among the three groups. Conclusions: Our results demonstrated that patients with R/R B-ALL receiving haplo-HSCT after CAR-T therapy achieved comparable outcomes to patients transplanted post-chemotherapy-based MRD-negative CR1, without increased risk of transplant-related mortality and toxicity.

Efficacy and Safety of Berberine Alone or Combined with Statins for the Treatment of Hyperlipidemia: A Systematic Review and Meta-Analysis of Randomized Controlled Clinical Trials.

To systematically evaluate the efficacy and safety of berberine for the treatment of hyperlipidemia, six electronic literature databases including SinoMed, CNKI, WanFang Data, PubMed, Embase and The Cochrane Library were searched to collect clinical randomized controlled trials (RCTs) of berberine alone or combined with statins for the treatment of hyperlipidemia from the inception to 8 March 2018. Two reviewers independently screened literature, extracted data and assessed the risk of bias of included RCTs. Then, meta-analysis was performed by using RevMan 5.3 software. A total of 11 RCTs involving 1386 patients were finally included. The results of meta-analysis showed that compared with the placebo group, berberine could significantly reduce the total cholesterol and low-density lipoprotein levels and elevate the high density lipoprotein level ( P<0.05 ). Compared with the simvastatin group, berberine was effective only in reducing the level of triglyceride ( MD=-0.37 , 95% CI: - 0.66, - 0.07, P=0.02 ). There, however, was no statistical significance between the BBR group and simvastatin group in the low density lipoprotein and high density lipoprotein levels. Compared with the simvastatin group, berberine plus simvastatin was more effective in reducing the level of triglyceride ( MD=-0.33 , 95% CI: - 0.46, - 0.20, P<0.00001 ) and total cholesterol ( MD=-0.36 , 95% CI: - 0.60, - 0.12, P=0.003 ). In terms of adverse reactions, the incidence of adverse reactions including transaminase elevation and muscle aches was lower in the berberine alone or combined with simvastatin group than that in the control group, while the instance of constipation was higher. This study suggests that berberine is effective for hyperlipidemia. The quality and quantity of included studies, however, were dissatisfactory, which might decrease the reliability of the results. Higher quality studies are needed to provide more high quality evidence.