Small extracellular vesicles from hypoxia-preconditioned bone marrow mesenchymal stem cells attenuate spinal cord injury via miR-146a-5p-mediated regulation of macrophage polarization

Spinal cord injury is a disabling condition with limited treatment options.Multiple studies have provided evidence suggesting that small extracellular vesicles(SEVs)secreted by bone marrow mesenchymal stem cells(MSCs)help mediate the beneficial effects conferred by MSC transplantation following spinal cord injury.Strikingly,hypoxia-preconditioned bone marrow mesenchymal stem cell-derived SEVs(HSEVs)exhibit increased therapeutic potency.We thus explored the role of HSEVs in macrophage immune regulation after spinal cord injury in rats and their significance in spinal cord repair.SEVs or HSEVs were isolated from bone marrow MSC supernatants by density gradient ultracentrifugation.HSEV administration to rats via tail vein injection after spinal cord injury reduced the lesion area and attenuated spinal cord inflammation.HSEVs regulate macrophage polarization towards the M2 phenotype in vivo and in vitro.Micro RNA sequencing and bioinformatics analyses of SEVs and HSEVs revealed that mi R-146a-5p is a potent mediator of macrophage polarization that targets interleukin-1 receptor-associated kinase 1.Reducing mi R-146a-5p expression in HSEVs partially attenuated macrophage polarization.Our data suggest that HSEVs attenuate spinal cord inflammation and injury in rats by transporting mi R-146a-5p,which alters macrophage polarization.This study provides new insights into the application of HSEVs as a therapeutic tool for spinal cord injury.

Hepatic arterial infusion chemotherapy with anti-angiogenesis agents and immune checkpoint inhibitors for unresectable hepatocellular carcinoma and meta-analysis

BACKGROUND Hepatic arterial infusion chemotherapy(HAIC)has been proven to be an ideal choice for treating unresectable hepatocellular carcinoma(uHCC).HAIC-based treatment showed great potential for treating uHCC.However,large-scale studies on HAIC-based treatments and meta-analyses of first-line treatments for uHCC are lacking.AIM To investigate better first-line treatment options for uHCC and to assess the safety and efficacy of HAIC combined with angiogenesis inhibitors,programmed cell death of protein 1(PD-1)and its ligand(PD-L1)blockers(triple therapy)under real-world conditions.METHODS Several electronic databases were searched to identify eligible randomized controlled trials for this meta-analysis.Study-level pooled analyses of hazard ratios(HRs)and odds ratios(ORs)were performed.This was a retrospective single-center study involving 442 patients with uHCC who received triple therapy or angiogenesis inhibitors plus PD-1/PD-L1 blockades(AIPB)at Sun Yat-sen University Cancer Center from January 2018 to April 2023.Propensity score matching(PSM)was performed to balance the bias between the groups.The Kaplan-Meier method and cox regression were used to analyse the survival data,and the log-rank test was used to compare the suvival time between the groups.RESULTS A total of 13 randomized controlled trials were included.HAIC alone and in combination with sorafenib were found to be effective treatments(P values for ORs:HAIC,0.95;for HRs:HAIC+sorafenib,0.04).After PSM,176 HCC patients were included in the analysis.The triple therapy group(n=88)had a longer median overall survival than the AIPB group(n=88)(31.6 months vs 14.6 months,P<0.001)and a greater incidence of adverse events(94.3%vs 75.4%,P<0.001).CONCLUSION This meta-analysis suggests that HAIC-based treatments are likely to be the best choice for uHCC.Our findings confirm that triple therapy is more effective for uHCC patients than AIPB.

Neural stem cell-derived exosomes regulate cell proliferation,migration,and cell death of brain microvascular endothelial cells via the miR-9/Hes1 axis under hypoxia

Background:Our previous study found that mouse embryonic neural stem cell(NSC)-derived exosomes(EXOs)regulated NSC differentiation via the miR-9/Hes1 axis.However,the effects of EXOs on brain microvascular endothelial cell(BMEC)dysfunction via the miR-9/Hes1 axis remain unknown.Therefore,the current study aimed to determine the effects of EXOs on BMEC proliferation,migration,and death via the miR-9/Hes1 axis.Methods:Immunofluorescence,quantitative real-time polymerase chain reaction,cell counting kit-8 assay,wound healing assay,calcein-acetoxymethyl/propidium iodide staining,and hematoxylin and eosin staining were used to determine the role and mechanism of EXOs on BMECs.Results:EXOs promoted BMEC proliferation and migration and reduced cell death under hypoxic conditions.The overexpression of miR-9 promoted BMEC prolifera-tion and migration and reduced cell death under hypoxic conditions.Moreover,miR-9 downregulation inhibited BMEC proliferation and migration and also promoted cell death.Hes1 silencing ameliorated the effect of amtagomiR-9 on BMEC proliferation and migration and cell death.Hyperemic structures were observed in the regions of the hippocampus and cortex in hypoxia-induced mice.Meanwhile,EXO treatment improved cerebrovascular alterations.Conclusion:NSC-derived EXOs can promote BMEC proliferation and migra-tion and reduce cell death via the miR-9/Hes1 axis under hypoxic conditions.Therefore,EXO therapeutic strategies could be considered for hypoxia-induced vascular injury.

TuBG1 promotes hepatocellular carcinoma via ATR/P53-apoptosis and cycling pathways

Background:As reported,γ-tubulin(TuBG1)is related to the occurrence and development of various types of malignant tumors.However,its role in hepatocellular cancer(HCC)is not clear.The present study was to investigate the relationship between TuBG1 and clinical parameters and survival in HCC patients.Methods:The correlation between TuBG1 and clinical parameters and survival in HCC patients was ex-plored by bioinformatics analysis.Immunohistochemistry was used for the verification.The molecular function of TuBG1 was measured using colony formation,scratch assay,trans-well assay and flow cytometry.Gene set enrichment analysis(GSEA)was used to pick up the enriched pathways,followed by investigating the target pathways using Western blotting.The tumor-immune system interactions and drug bank database(TISIDB)was used to evaluate TuBG1 and immunity.Based on the TuBG1-related immune genes,a prognostic model was constructed and was further validated internally and externally.Results:The bioinformatic analysis found high expressed TuBG1 in HCC tissue,which was confirmed us-ing immunohistochemistry and Western blotting.After silencing the TuBG1 in HCC cell lines,more G1 arrested cells were found,cell proliferation and invasion were inhibited,and apoptosis was promoted.Furthermore,the silence of TuBG1 increased the expressions of Ataxia-Telangiectasia and Rad-3(ATR),phospho-P38 mitogen-activated protein kinase(P-P38MAPK),phospho-P53(P-P53),B-cell lymphoma-2 associated X protein(Bax),cleaved caspase 3 and P21;decreased the expressions of B-cell lymphoma-2(Bcl-2),cyclin D1,cyclin E2,cyclin-dependent kinase 2(CDK2)and CDK4.The correlation analysis of immunohistochemistry and clinical parameters and survival data revealed that TuBG1 was negatively corre-lated with the overall survival.The constructed immune prognosis model could effectively evaluate the prognosis.Conclusions:The increased expression of TuBG1 in HCC is associated with poor prognosis,which might be involved in the occurrence and development of HCC.

Prohibitin 1 inhibits cell proliferation and induces apoptosis via the p53-mediated mitochondrial pathway in vitro

BACKGROUND Prohibitin 1(PHB1)has been identified as an antiproliferative protein that is highly conserved and ubiquitously expressed,and it participates in a variety of essential cellular functions,including apoptosis,cell cycle regulation,prolifera-tion,and survival.Emerging evidence indicates that PHB1 may play an important role in the progression of hepatocellular carcinoma(HCC).However,the role of PHB1 in HCC is controversial.AIM To investigate the effects of PHB1 on the proliferation and apoptosis of human HCC cells and the relevant mechanisms in vitro.METHODS HCC patients and healthy individuals were enrolled in this study according to the inclusion and exclusion criteria;then,PHB1 levels in the sera and liver tissues of these participates were determined using ELISA,RT-PCR,and immunohistoche-mistry.Human HepG2 and SMMC-7721 cells were transfected with the pEGFP-PHB1 plasmid and PHB1-specific shRNA(shRNA-PHB1)for 24-72 h.Cell prolif-eration was analysed with an MTT assay.Cell cycle progression and apoptosis were analysed using flow cytometry(FACS).The mRNA and protein expression levels of the cell cycle-related molecules p21,Cyclin A2,Cyclin E1,and CDK2 and the cell apoptosis-related molecules cytochrome C(Cyt C),p53,Bcl-2,Bax,caspase 3,and caspase 9 were measured by real-time PCR and Western blot,respectively.RESULTS Decreased levels of PHB1 were found in the sera and liver tissues of HCC patients compared to those of healthy individuals,and decreased PHB1 was positively correlated with low differentiation,TNM stage III-IV,and alpha-fetoprotein≥400μg/L.Overexpression of PHB1 significantly inhibited human HCC cell proliferation in a time-dependent manner.FACS revealed that the overexpression of PHB1 arrested HCC cells in the G0/G1 phase of the cell cycle and induced apoptosis.The proportion of cells in the G0/G1 phase was significantly increased and the proportion of cells in the S phase was decreased in HepG2 cells that were transfected with pEGFP-PHB1 compared with untreated control and empty vector-transfected cells.The percentage of apoptotic HepG2 cells that were transfected with pEGFP-PHB1 was 15.41%±1.06%,which was significantly greater than that of apoptotic control cells(3.65%±0.85%,P<0.01)and empty vector-transfected cells(4.21%±0.52%,P<0.01).Similar results were obtained with SMMC-7721 cells.Furthermore,the mRNA and protein expression levels of p53,p21,Bax,caspase 3,and caspase 9 were increased while the mRNA and protein expression levels of Cyclin A2,Cy-clin E1,CDK2,and Bcl-2 were decreased when PHB1 was overexpressed in human HCC cells.However,when PHB1 was upregulated in human HCC cells,Cyt C expression levels were increased in the cytosol and decreased in the mitochondria,which indicated that Cyt C had been released into the cytosol.Conversely,these effects were reversed when PHB1 was knocked down.CONCLUSION PHB1 inhibits human HCC cell viability by arresting the cell cycle and inducing cell apoptosis via activation of the p53-mediated mitochondrial pathway.

Transcriptome sequencing reveals novel biomarkers and immune cell infiltration in esophageal tumorigenesis

BACKGROUND Esophageal squamous cell carcinoma(ESCC)is one of the most common malignancies worldwide,and its development comprises a multistep process from intraepithelial neoplasia(IN)to carcinoma(CA).However,the critical regulators and underlying molecular mechanisms remain largely unknown.AIM To explore the genes and infiltrating immune cells in the microenvironment that are associated with the multistage progression of ESCC to facilitate diagnosis and early intervention.METHODS A mouse model mimicking the multistage development of ESCC was established by providing warter containing 4-nitroquinoline 1-oxide(4NQO)to C57BL/6 mice.Moreover,we established a control group without 4NQO treatment of mice.Then,transcriptome sequencing was performed for esophageal tissues from patients with different pathological statuses,including low-grade IN(LGIN),high-grade IN(HGIN),and CA,and controlled normal tissue(NOR)samples.Differentially expressed genes(DEGs)were identified in the LGIN,HGIN,and CA groups,and the biological functions of the DEGs were analyzed via Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses.The CIBERSORT algorithm was used to detect the pattern of immune cell infilt-ration.Immunohistochemistry(IHC)was also conducted to validate our results.Finally,the Luminex multiplex cytokine analysis was utilized to measure the serum cytokine levels in the mice.RESULTS Compared with those in the NOR group,a total of 681541,and 840 DEGs were obtained in the LGIN,HGIN,and CA groups,respectively.Using the intersection of the three sets of DEGs,we identified 86 genes as key genes involved in the development of ESCC.Enrichment analysis revealed that these genes were enriched mainly in the keratinization,epidermal cell differentiation,and interleukin(IL)-17 signaling pathways.CIBERSORT analysis revealed that,compared with those in the NOR group,M0 and M1 macrophages in the 4NQO group showed stronger infiltration,which was validated by IHC.Serum cytokine analysis revealed that,compared with those in the NOR group,IL-1βand IL-6 were upregulated,while IL-10 was downregulated in the LGIN,HGIN,and CA groups.Moreover,the expression of the representative key genes,such as S100a8 and Krt6b,was verified in external human samples,and the results of immunohistochemical staining were consistent with the findings in mice.CONCLUSION We identified a set of key genes represented by S100a8 and Krt6b and investigated their potential biological functions.In addition,we found that macrophage infiltration and abnormal alterations in the levels of inflam-mation-associated cytokines,such as IL-1β,IL-6,and IL-10,in the peripheral blood may be closely associated with the development of ESCC.

Revolutionizing tumor immunotherapy:unleashing the power of progenitor exhausted T cells

In exploring persistent infections and malignancies, a distinctive subgroup of CD8^(+) T cells, progenitor exhausted CD8^(+) T(Tpex) cells, has been identified. These Tpex cells are notable for their remarkable self-renewal and rapid proliferation abilities. Recent strides in immunotherapy have demonstrated that Tpex cells expand and differentiate into responsive exhausted CD8^(+) T cells, thus underscoring their critical role in the immunotherapeutic retort. Clinical examinations have further clarified a robust positive correlation between the proportional abundance of Tpex cells and enhanced clinical prognosis. Tpex cells have found noteworthy applications in the formulation of inventive immunotherapeutic approaches against tumors. This review describes the functions of Tpex cells in the tumor milieu, particularly their potential utility in tumor immunotherapy. Precisely directing Tpex cells may be essential to achieving successful outcomes in immunotherapy against tumors.

Efficacy and predictive factors of transarterial chemoembolization combined with lenvatinib plus programmed cell death protein-1 inhibition for unresectable hepatocellular carcinoma

BACKGROUND The efficacy and safety of transarterial chemoembolization(TACE)combined with lenvatinib plus programmed cell death protein-1(PD-1)for unresectable hepato-cellular carcinoma(HCC)have rarely been evaluated and it is unknown which factors are related to efficacy.AIM To evaluate the efficacy and independent predictive factors of TACE combined with lenvatinib plus PD-1 inhibitors for unresectable HCC.METHODS This study retrospectively enrolled patients with unresectable HCC who received TACE/lenvatinib/PD-1 treatment between March 2019 and April 2022.Overall survival(OS)and progression-free survival(PFS)were determined.The objective response rate(ORR)and disease control rate(DCR)were evaluated in accordance with the modified Response Evaluation Criteria in Solid Tumors.Additionally,the prognostic factors affecting the clinical outcome were assessed.RESULTS One hundred and two patients were enrolled with a median follow-up duration of 12.63 months.The median OS was 26.43 months(95%CI:17.00-35.87),and the median PFS was 10.07 months(95%CI:8.50-11.65).The ORR and DCR were 61.76%and 81.37%,respectively.The patients with Barcelona Clinic Liver Cancer Classification(BCLC)B stage,early neutrophil-to-lymphocyte ratio(NLR)response(decrease),or early alpha-fetoprotein(AFP)response(decrease>20%)had superior OS and PFS than their counterparts.CONCLUSION This study showed that TACE/lenvatinib/PD-1 treatment was well tolerated with encouraging efficacy in patients with unresectable HCC.The patients with BCLC B-stage disease with early NLR response(decrease)and early AFP response(decrease>20%)may achieve better clinical outcomes with this triple therapy.

Effectiveness and tolerability of programmed cell death protein-1 inhibitor+chemotherapy compared to chemotherapy for upper gastrointestinal tract cancers

BACKGROUND The combination of programmed cell death protein-1(PD-1)inhibitor and che-motherapy is approved as a standard first-or second-line treatment in patients with advanced oesophageal or gastric cancer.However,it is unclear whether this combination is superior to chemotherapy alone.AIM To assess the comparative effectiveness and tolerability of combining PD-1 inhibitors with chemotherapy vs chemotherapy alone in patients with advanced gastric cancer,gastroesophageal junction(GEJ)cancer,or oesophageal carcinoma.METHODS We searched the PubMed and Embase databases for studies that compared the efficacy and tolerance of PD-1 inhibitors in combination with chemotherapy vs chemotherapy alone in patients with advanced oesophageal or gastric cancer.We employed either random or fixed models to analyze the outcomes of each clinical trial,en-compassing data on overall survival(OS),progression-free survival(PFS),objective response rate,and adverse events(AEs).RESULTS Nine phase 3 clinical trials(7016 advanced oesophageal and gastric cancer patients)met the inclusion criteria.Our meta-analysis demonstrated that the pooled PD-1 inhibitor+chemotherapy group had a significantly longer OS than the chemotherapy-alone group[hazard ratio(HR)=0.76,95%confidence interval(CI):0.71-0.81];the pooled PFS result was consistent with that of OS(HR=0.67,95%CI:0.61-0.74).The count of patients achieving an objective response in the PD-1 inhibitor+chemotherapy group surpassed that of the chemotherapy-alone group[odds ratio(OR)=1.86,95%CI:1.59-2.18].AE incidence was also higher in the combination-therapy group than in the chemotherapy-alone group,regardless of whether≥grade 3 only(OR=1.30,95%CI:1.07-1.57)or all AE grades(OR=1.88,95%CI:1.39-2.54)were examined.We performed a subgroup analysis based on the programmed death-ligand 1(PD-L1)combined positive score(CPS)and noted extended OS and PFS durations within the CPS≥1,CPS≥5,and CPS≥10 subgroups of the PD-1 inhibitor+chemotherapy group.CONCLUSION In contrast to chemotherapy alone,the combination of PD-1 inhibitor and chemotherapy appears to present a more favorable option for initial or subsequent treatment in patients with gastric cancer,GEJ tumor,or oesophageal cancer.This holds true particularly for individuals with PD-L1 CPS scores of≥5 and≥10.

Cardiac differentiation is modulated by anti-apoptotic signals in murine embryonic stem cells

BACKGROUND Embryonic stem cells(ESCs)serve as a crucial ex vivo model,representing epiblast cells derived from the inner cell mass of blastocyst-stage embryos.ESCs exhibit a unique combination of self-renewal potency,unlimited proliferation,and pluripotency.The latter is evident by the ability of the isolated cells to differ-entiate spontaneously into multiple cell lineages,representing the three primary embryonic germ layers.Multiple regulatory networks guide ESCs,directing their self-renewal and lineage-specific differentiation.Apoptosis,or programmed cell death,emerges as a key event involved in sculpting and forming various organs and structures ensuring proper embryonic development.How-ever,the molecular mechanisms underlying the dynamic interplay between diffe-rentiation and apoptosis remain poorly understood.AIM To investigate the regulatory impact of apoptosis on the early differentiation of ESCs into cardiac cells,using mouse ESC(mESC)models-mESC-B-cell lym-phoma 2(BCL-2),mESC-PIM-2,and mESC-metallothionein-1(MET-1)-which overexpress the anti-apoptotic genes Bcl-2,Pim-2,and Met-1,respectively.METHODS mESC-T2(wild-type),mESC-BCL-2,mESC-PIM-2,and mESC-MET-1 have been used to assess the effect of potentiated apoptotic signals on cardiac differentiation.The hanging drop method was adopted to generate embryoid bodies(EBs)and induce terminal differentiation of mESCs.The size of the generated EBs was measured in each condition compared to the wild type.At the functional level,the percentage of cardiac differentiation was measured by calculating the number of beating cardiomyocytes in the manipulated mESCs compared to the control.At the molecular level,quantitative reverse transcription-polymerase chain reaction was used to assess the mRNA expression of three cardiac markers:Troponin T,GATA4,and NKX2.5.Additionally,troponin T protein expression was evaluated through immunofluorescence and western blot assays.RESULTS Our findings showed that the upregulation of Bcl-2,Pim-2,and Met-1 genes led to a reduction in the size of the EBs derived from the manipulated mESCs,in comparison with their wild-type counterpart.Additionally,a decrease in the count of beating cardiomyocytes among differentiated cells was observed.Furthermore,the mRNA expression of three cardiac markers-troponin T,GATA4,and NKX2.5-was diminished in mESCs overexpressing the three anti-apoptotic genes compared to the control cell line.Moreover,the overexpression of the anti-apoptotic genes resulted in a reduction in troponin T protein expression.CONCLUSION Our findings revealed that the upregulation of Bcl-2,Pim-2,and Met-1 genes altered cardiac differentiation,providing insight into the intricate interplay between apoptosis and ESC fate determination.

Subcellular distribution of prohibitin 1 in rat liver during liver regeneration and its cellular implication

BACKGROUND The function of prohibitin 1(Phb1)during liver regeneration(LR)remains relatively unexplored.Our previous research identified downregulation of Phb1 in rat liver mitochondria 24 h after 70%partial hepatectomy(PHx),as determined by subcellular proteomic analysis.AIM To investigate the potential role of Phb1 during LR.METHODS We examined changes in Phb1 mRNA and protein levels,subcellular distribution,and abundance in rat liver during LR following 70%PHx.We also evaluated mitochondrial changes and apoptosis using electron microscopy and flow cytometry.RNA-interference-mediated knockdown of Phb1(PHBi)was performed in BRL-3A cells.RESULTS Compared with sham-operation control groups,Phb1 mRNA and protein levels in 70%PHx test groups were downregulated at 24 h,then upregulated at 72 and 168 h.Phb1 was mainly located in mitochondria,showed a reduced abundance at 24 h,significantly increased at 72 h,and almost recovered to normal at 168 h.Phb1 was also present in nuclei,with continuous increase in abundance observed 72 and 168 h after 70%PHx.The altered ultrastructure and reduced mass of mitochondria during LR had almost completely recovered to normal at 168 h.PHBi in BRL-3A cells resulted in increased S-phase entry,a higher number of apoptotic cells,and disruption of mitochondrial membrane potential.CONCLUSION Phb1 may contribute to maintaining mitochondrial stability and could play a role in regulating cell proliferation and apoptosis of rat liver cells during LR.

C-reactive protein to albumin ratio predict responses to programmed cell death-1 inhibitors in hepatocellular carcinoma patients

BACKGROUND Over the years,programmed cell death-1(PD-1)inhibitors have been routinely used for hepatocellular carcinoma(HCC)treatment and yielded improved survival outcomes.Nonetheless,significant heterogeneity surrounds the outcomes of most studies.Therefore,it is critical to search for biomarkers that predict the efficacy of PD-1 inhibitors in patients with HCC.AIM To investigate the role of the C-reactive protein to albumin ratio(CAR)in evaluating the efficacy of PD-1 inhibitors for HCC.METHODS The clinical data of 160 patients with HCC treated with PD-1 inhibitors from January 2018 to November 2022 at the First Affiliated Hospital of Guangxi Medical University were retrospectively analyzed.RESULTS The optimal cut-off value for CAR based on progression-free survival(PFS)was determined to be 1.20 using x-tile software.Cox proportional risk model was used to determine the factors affecting prognosis.Eastern Cooperative Oncology Group performance status[hazard ratio(HR)=1.754,95%confidence interval(95%CI)=1.045-2.944,P=0.033],CAR(HR=2.118,95%CI=1.057-4.243,P=0.034)and tumor number(HR=2.932,95%CI=1.246-6.897,P=0.014)were independent prognostic factors for overall survival.CAR(HR=2.730,95%CI=1.502-4.961,P=0.001),tumor number(HR=1.584,95%CI=1.003-2.500,P=0.048)and neutrophil to lymphocyte ratio(HR=1.120,95%CI=1.022-1.228,P=0.015)were independent prognostic factors for PFS.Two nomograms were constructed based on independent prognostic factors.The C-index index and calibration plots confirmed that the nomogram is a reliable risk prediction tool.The ROC curve and decision curve analysis confirmed that the nomogram has a good predictive effect as well as a net clinical benefit.CONCLUSION Overall,we reveal that the CAR is a potential predictor of short-and long-term prognosis in patients with HCC treated with PD-1 inhibitors.If further verified,CAR-based nomogram may increase the number of markers that predict individualized prognosis.

Camrelizumab,apatinib and hepatic artery infusion chemotherapy combined with microwave ablation for advanced hepatocellular carcinoma

BACKGROUND Hepatic arterial infusion chemotherapy and camrelizumab plus apatinib(TRIPLET protocol)is promising for advanced hepatocellular carcinoma(Ad-HCC).However,the usefulness of microwave ablation(MWA)after TRIPLET is still controversial.AIM To compare the efficacy and safety of TRIPLET alone(T-A)vs TRIPLET-MWA(TM)for Ad-HCC.METHODS From January 2018 to March 2022,217 Ad-HCC patients were retrospectively enrolled.Among them,122 were included in the T-A group,and 95 were included in the T-M group.A propensity score matching(PSM)was applied to balance bias.Overall survival(OS)was compared using the Kaplan-Meier curve with the log-rank test.The overall objective response rate(ORR)and major complications were also assessed.RESULTS After PSM,82 patients were included both the T-A group and the T-M group.The ORR(85.4%)in the T-M group was significantly higher than that(65.9%)in the T-A group(P<0.001).The cumulative 1-,2-,and 3-year OS rates were 98.7%,93.4%,and 82.0%in the T-M group and 85.1%,63.1%,and 55.0%in the T-A group(hazard ratio=0.22;95%confidence interval:0.10-0.49;P<0.001).The incidence of major complications was 4.9%(6/122)in the T-A group and 5.3%(5/95)in the T-M group,which were not significantly different(P=1.000).CONCLUSION T-M can provide better survival outcomes and comparable safety for Ad-HCC than T-A.

3D Collagen Gels:A Promising Platform for Dendritic Cell Culture in Biomaterials Research

The three-dimensional(3D)cell culture system has garnered significant attention in recent years as a means of studying cell behavior and tissue development,as opposed to traditional two-dimensional cultures.These systems can induce specific cell reactions,promote specific tissue functions,and serve as valuable tools for research in tissue engineering,regenerative medicine,and drug discovery.This paper discusses current developments in the field of three-dimensional cell culture and the potential applications of 3D type 1 collagen gels to enhance the growth and maturation of dendritic cells.

Combined TIM-3 and PD-1 blockade restrains hepatocellular carcinoma development by facilitating CD4+ and CD8+T cellmediated antitumor immune responses

BACKGROUND Immune checkpoint inhibitors(ICIs)targeting programmed cell death protein 1(PD-1)and T cell immunoglobulin and mucin domain-containing protein 3(TIM-3)are beneficial to the resumption of anti-tumor immunity response and hold extreme potential as efficient therapies for certain malignancies.However,ICIs with a single target exhibit poor overall response rate in hepatocellular carcinoma(HCC)patients due to the complex pathological mechanisms of HCC.AIM To investigate the effects of combined TIM-3 and PD-1 blockade on tumor development in an HCC mouse model,aiming to identify more effective immunotherapies and provide more treatment options for HCC patients.METHODS The levels of PD-1 and TIM-3 on CD4+and CD8+T cells from tumor tissues,ascites,and matched adjacent tissues from HCC patients were determined with flow cytometry.An HCC xenograft mouse model was established and treated with anti-TIM-3 monoclonal antibody(mAb)and/or anti-PD-1 mAb.Tumor growth in each group was measured.Hematoxylin and eosin staining and immunohistochemical staining were used to evaluate T cell infiltration in tumors.The percentage of CD4+and CD8+T cells in tissue samples from mice was tested with flow cytometry.The percentages of PD-1+CD8+,TIM-3+CD8+,and PD-1+TIM-3+CD8+T cells was accessed by flow cytometry.The levels of the cytokines including tumor necrosis factor alpha(TNF-α),interferon-γ(IFN-γ),interleukin(IL)-6,and IL-10 in tumor tissues were gauged with enzyme-linked immunosorbent assay kits.RESULTS We confirmed that PD-1 and TIM-3 expression was substantially upregulated in CD4+and CD8+T cells isolated from tumor tissues and ascites of HCC patients.TIM-3 mAb and PD-1 mAb treatment both reduced tumor volume and weight,while combined blockade had more substantial anti-tumor effects than individual treatment.Then we showed that combined therapy increased T cell infiltration into tumor tissues,and downregulated PD-1 and TIM-3 expression on CD8+T cells in tumor tissues.Moreover,combined treatment facilitated the production of T cell effector cytokines TNF-α and IFN-γ,and reduced the production of immunosuppressive cytokines IL-10 and IL-6 in tumor tissues.Thus,we implicated that combined blockade could ameliorate T cell exhaustion in HCC mouse model.CONCLUSION Combined TIM-3 and PD-1 blockade restrains HCC development by facilitating CD4+ and CD8+T cell-mediated antitumor immune responses.

Overexpressing NeuroD1 reprograms Müller cells into various types of retinal neurons

The onset of retinal degenerative disease is often associated with neuronal loss. Therefore, how to regenerate new neurons to restore vision is an important issue. NeuroD1 is a neural transcription factor with the ability to reprogram brain astrocytes into neurons in vivo. Here, we demonstrate that in adult mice, NeuroD1 can reprogram Müller cells, the principal glial cell type in the retina, to become retinal neurons. Most strikingly, ectopic expression of NeuroD1 using two different viral vectors converted Müller cells into different cell types. Specifically, AAV7 m8 GFAP681::GFP-ND1 converted Müller cells into inner retinal neurons, including amacrine cells and ganglion cells. In contrast, AAV9 GFAP104::ND1-GFP converted Müller cells into outer retinal neurons such as photoreceptors and horizontal cells, with higher conversion efficiency. Furthermore, we demonstrate that Müller cell conversion induced by AAV9 GFAP104::ND1-GFP displayed clear dose-and time-dependence. These results indicate that Müller cells in adult mice are highly plastic and can be reprogrammed into various subtypes of retinal neurons.

TM9SF1 promotes bladder cancer cell growth and infiltration

BACKGROUND Bladder cancer(BC)is the most common urological tumor.It has a high recur-rence rate,displays tutor heterogeneity,and resists chemotherapy.Furthermore,the long-term survival rate of BC patients has remained unchanged for decades,which seriously affects the quality of patient survival.To improve the survival rate and prognosis of BC patients,it is necessary to explore the molecular mechanisms of BC development and progression and identify targets for treatment and intervention.Transmembrane 9 superfamily member 1(TM9SF1),also known as MP70 and HMP70,is a member of a family of nine transmembrane superfamily proteins,which was first identified in 1997.TM9SF1 can be expressed in BC,but its biological function and mechanism in BC are not clear.AIM To investigate the biological function and mechanism of TM9SF1 in BC.Overexpression of TM9SF1 increased the in vitro proliferation,migration,and invasion of BC cells by promoting the entry of BC cells into the G2/M phase.Silencing of TM9SF1 inhibited in vitro proliferation,migration,and invasion of BC cells and blocked BC cells in the G1 phase.CONCLUSION TM9SF1 may be an oncogene in BC.

PbGIF1 promoting cell-proliferation in pear fruit is transcriptionally activated by Pb RR1

As a cell proliferation regulator involved in wide biological processes in plants,GRF-INTERACTING FACTOR(GIF)controls different tissues development.However,whether GIF participates in fruit development remains unclear.According to transcriptome data,we identified PbGIF1was highly expressed during fruit development in cytokinins induced parthenocarpy pear.In the present study,the biofunction of PbGIF1 was initially verified.Overexpression of PbGIF1 promoted fruit size of transgenic tomato.The size of flesh fruit was not affected by cell expansion but the cell proliferation was promoted by overexpressing Pb GIF1.The accelerated cell proliferation process was also observed in PbGIF1-overexpressed transgenic pear fruit calli.The transcriptional regulation of cytokinins on PbGIF1 was further confirmed by exogenous CPPU treatments in pear fruitlets.To investigate the underlying mechanism,the cytokinins-responded factor,PbRR1,was further focused on.The results of Yeast-one-hybrid assay suggested that PbRR1 can bind to the promoter sequence of PbGIF1.The transcriptional activation of PbRR1 on PbGIF1 was also confirmed by Dual-Luciferase assays.Taken together,the results showed that cytokinins control pear fruit development via the transcriptional activation of PbGIF1 by PbRR1.

Prox1 Suppresses Proliferation and Drug Resistance of Retinoblastoma Cells via Targeting Notch1

Objective Retinoblastoma(RB)is a prevalent type of eye cancer in youngsters.Prospero homeobox 1(Prox1)is a homeobox transcriptional repressor and downstream target of the proneural gene that is relevant in lymphatic,hepatocyte,pancreatic,heart,lens,retinal,and cancer cells.The goal of this study was to investigate the role of Prox1 in RB cell proliferation and drug resistance,as well as to explore the underlying Notch1 mechanism.Methods Human RB cell lines(SO-RB50 and Y79)and a primary human retinal microvascular endothelial cell line(ACBRI-181)were used in this study.The expression of Prox1 and Notch1 mRNA and protein in RB cells was detected using quantitative real time-polymerase chain reaction(RT-qPCR)and Western blotting.Cell proliferation was assessed after Prox1 overexpression using the Cell Counting Kit-8 and the MTS assay.Drug-resistant cell lines(SO-RB50/vincristine)were generated and treated with Prox1 to investigate the role of Prox1 in drug resistance.We employed pcDNA-Notch1 to overexpress Notch1 to confirm the role of Notch1 in the protective function of Prox1.Finally,a xenograft model was constructed to assess the effect of Prox1 on RB in vivo.Results Prox1 was significantly downregulated in RB cells.Overexpression of Prox1 effectively decreased RB cell growth while increasing the sensitivity of drug-resistant cells to vincristine.Notch1 was involved in Prox1’s regulatory effects.Notch1 was identified as a target gene of Prox1,which was found to be upregulated in RB cells and repressed by increased Prox1 expression.When pcDNA-Notch1 was transfected,the effect of Prox1 overexpression on RB was removed.Furthermore,by downregulating Notch1,Prox1 overexpression slowed tumor development and increased vincristine sensitivity in vivo.Conclusion These data show that Prox1 decreased RB cell proliferation and drug resistance by targeting Notch1,implying that Prox1 could be a potential therapeutic target for RB.

TAX1BP1 and FIP200 orchestrate non-canonical autophagy of p62 aggregates for mouse neural stem cell maintenance

Autophagy plays a pivotal role in diverse biological processes,including the maintenance and differentiation of neural stem cells(NSCs).Interestingly,while complete deletion of Fip200 severely impairs NSC maintenance and differentiation,inhibiting canonical autophagy via deletion of core genes,such as Atg5,Atg16l1,and Atg7,or blockade of canonical interactions between FIP200 and ATG13(designated as FIP200-4A mutant or FIP200 KI)does not produce comparable detrimental effects.This highlights the likely critical involvement of the non-canonical functions of FIP200,the mechanisms of which have remained elusive.Here,utilizing genetic mouse models,we demonstrated that FIP200 mediates non-canonical autophagic degradation of p62/sequestome1,primarily via TAX1BP1 in NSCs.Conditional deletion of Tax1bp1 in fip200hGFAP conditional knock-in(cKI)mice led to NSC deficiency,resembling the fip200hGFAP conditional knockout(cKO)mouse phenotype.Notably,reintroducing wild-type TAX1BP1 not only restored the maintenance of NSCs derived from tax1bp1-knockout fip200hGFAP cKI mice but also led to a marked reduction in p62 aggregate accumulation.Conversely,a TAX1BP1 mutant incapable of binding to FIP200 or NBR1/p62 failed to achieve this restoration.Furthermore,conditional deletion of Tax1bp1 in fip200hGFAP cKO mice exacerbated NSC deficiency and p62 aggregate accumulation compared to fip200hGFAP cKO mice.Collectively,these findings illustrate the essential role of the FIP200-TAX1BP1 axis in mediating the non-canonical autophagic degradation of p62 aggregates towards NSC maintenance and function,presenting novel therapeutic targets for neurodegenerative diseases.