scholarly journals The effects of e-cigarette vapor exposure on the transcriptome and virulence of Streptococcus pneumoniae

2019 ◽  
Author(s):  
Kamal Bagale ◽  
Santosh Paudel ◽  
Hayden Cagle ◽  
Erin Sigel ◽  
Ritwij Kulkarni
Keyword(s):  
Streptococcus Pneumoniae ◽  
Stress Response ◽  
Mouse Model ◽  
Pathogenic Bacteria ◽  
Sugar Transport ◽  
Dependent Manner ◽  
Altered Expression ◽  
Acute Pneumonia

AbstractThe effects of e-cigarette vapor (EV) exposure on the physiology of respiratory microflora are not fully defined. We analyzed the effects of exposure to vapor from nicotine-containing and nicotine-free e-liquid formulations on virulence and transcriptome of Streptococcus pneumoniae strain TIGR4, a pathogen that asymptomatically colonizes human nasopharyngeal mucosa. TIGR4 was pre-exposed for 2h to nicotine-containing EV extract (EVE+NIC), nicotine-free EV extract (EVE−NIC), cigarette smoke extract (CSE), or nutrient-rich TS broth (control). The differences in the treatment and control TIGR4 were explored using transcriptome sequencing, in vitro virulence assays, and in vivo mouse model of acute pneumonia. The analysis of RNASeq profiles revealed modest changes in the expression of 14 genes involved in sugar transport and metabolism in EVE−NIC pre-exposed TIGR4 compared to the control. While, EVE+NIC or CSE exposure altered expression of 264 and 982 genes, respectively, most of which were involved in metabolism and stress response. Infection in a mouse model of acute pneumonia with control TIGR4 or with TIGR4 pre-exposed to EVE+NIC, EVE−NIC, or CSE did not show significant differences in disease parameters, such as bacterial organ burden and respiratory cytokine response. Interestingly, TIGR4 exposed to CSE or EVE+NIC (but not EVE−NIC) exhibited moderate induction of biofilm formation. However, none of the treatment groups showed significant alterations in pneumococcal hydrophobicity or epithelial cell adherence. In summary, our study reports that exposure to EV significantly alters the S. pneumoniae transcriptome in a nicotine-dependent manner without affecting pneumococcal virulence.ImportanceWith the increasing popularity of e-cigarettes amongst cigarette smoking and non-smoking adults and children, and the recent reports of vaping related lung illnesses and deaths, further analysis of the adverse health effects of e-cigarette vapor (EV) exposure is warranted. Since pathogenic bacteria such as Streptococcus pneumoniae can colonize the human nasopharynx as commensals, they may be affected by the exposure to bioactive chemicals in EV. Hence in this study we examined the effects of EV exposure on the physiology of S. pneumoniae strain TIGR4. In order to differentiate between the effects of nicotine and non-nicotine components, we specifically compared RNASeq profiles and virulence of TIGR4 exposed to vapor from nicotine-containing and nicotine-free e-liquid formulations. We observed that nicotine-containing EV augmented TIGR4 biofilms and altered expression of TIGR4 genes predominantly involved in metabolism and stress response. However, neither nicotine-containing nor nicotine-free EV affected TIGR4 virulence in a mouse model.

scholarly journals Electronic Cigarette (E-Cigarette) Vapor Exposure Alters the Streptococcus pneumoniae Transcriptome in a Nicotine-Dependent Manner without Affecting Pneumococcal Virulence

2019 ◽  
Vol 86 (3) ◽  
Author(s):  
Kamal Bagale ◽  
Santosh Paudel ◽  
Hayden Cagle ◽  
Erin Sigel ◽  
Ritwij Kulkarni
Keyword(s):  
Streptococcus Pneumoniae ◽  
Stress Response ◽  
Mouse Model ◽  
Pathogenic Bacteria ◽  
Electronic Cigarette ◽  
Dependent Manner ◽  
Rna Seq ◽  
Altered Expression ◽  
Content Type ◽  
Acute Pneumonia

ABSTRACT The effects of electronic cigarette (e-cigarette) vapor (EV) exposure on the physiology of respiratory microflora are not fully defined. We analyzed the effects of exposure to vapor from nicotine-containing and nicotine-free e-liquid formulations on the virulence and transcriptome of Streptococcus pneumoniae strain TIGR4, a pathogen that asymptomatically colonizes the human nasopharyngeal mucosa. TIGR4 was preexposed for 2 h to nicotine-containing EV extract (EVE+NIC), nicotine-free EV extract (EVE−NIC), cigarette smoke extract (CSE), or nutrient-rich tryptic soy (TS) broth (control). The differences between the treatment and control strains were explored using transcriptome sequencing (RNA sequencing [RNA-Seq]), in vitro virulence assays, and an in vivo mouse model of acute pneumonia. The analysis of RNA-Seq profiles revealed modest changes in the expression of 14 genes involved in sugar transport and metabolism in EVE−NIC-preexposed TIGR4 compared to the control, while EVE+NIC or CSE exposure altered expression of 264 and 982 genes, respectively, most of which were involved in metabolism and stress response. Infection in a mouse model of acute pneumonia with control TIGR4 or with TIGR4 preexposed to EVE+NIC, EVE−NIC, or CSE did not show significant differences in disease parameters, such as bacterial organ burden and respiratory cytokine response. Interestingly, TIGR4 exposed to CSE or EVE+NIC (but not EVE−NIC) exhibited moderate induction of biofilm formation. However, none of the treatment groups showed significant alterations in pneumococcal hydrophobicity or epithelial cell adherence. In summary, our study reports that exposure to EV significantly alters the S. pneumoniae transcriptome in a nicotine-dependent manner without affecting pneumococcal virulence. IMPORTANCE With the increasing popularity of e-cigarettes among cigarette smoking and nonsmoking adults and children and the recent reports of vaping-related lung illness and deaths, further analysis of the adverse health effects of e-cigarette vapor (EV) exposure is warranted. Since pathogenic bacteria such as Streptococcus pneumoniae can colonize the human nasopharynx as commensals, they may be affected by exposure to bioactive chemicals in EV. Hence, in this study we examined the effects of EV exposure on the physiology of S. pneumoniae strain TIGR4. In order to differentiate between the effects of nicotine and nonnicotine components, we specifically compared the RNA-Seq profiles and virulence of TIGR4 exposed to vapor from nicotine-containing and nicotine-free e-liquid formulations. We observed that nicotine-containing EV augmented TIGR4 biofilms and altered expression of TIGR4 genes predominantly involved in metabolism and stress response. However, neither nicotine-containing nor nicotine-free EV affected TIGR4 virulence in a mouse model.

scholarly journals Murine monoclonal antibodies against RBD of SARS-CoV-2 neutralize authentic wild type SARS-CoV-2 as well as B.1.1.7 and B.1.351 viruses and protect in vivo in a mouse model in a neutralization dependent manner

2021 ◽  
Author(s):  
Fatima Amanat ◽  
Shirin Strohmeier ◽  
Wen-Hsin Lee ◽  
Sandhya Bangaru ◽  
Andrew B Ward ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Severe Acute Respiratory Syndrome ◽  
Mouse Model ◽  
Neutralizing Antibodies ◽  
Dependent Manner ◽  
Receptor Binding Domain ◽  
Wild Type ◽  
Murine Monoclonal Antibodies

After first emerging in December 2019 in China, severe acute respiratory syndrome 2 (SARS-CoV-2) has since caused a pandemic leading to millions of infections and deaths worldwide. Vaccines have been developed and authorized but supply of these vaccines is currently limited. With new variants of the virus now emerging and spreading globally, it is essential to develop therapeutics that are broadly protective and bind conserved epitopes in the receptor binding domain (RBD) or the whole spike of SARS-CoV-2. In this study, we have generated mouse monoclonal antibodies (mAbs) against different epitopes on the RBD and assessed binding and neutralization against authentic SARS-CoV-2. We have demonstrated that antibodies with neutralizing activity, but not non-neutralizing antibodies, lower viral titers in the lungs when administered in a prophylactic setting in vivo in a mouse challenge model. In addition, most of the mAbs cross-neutralize the B.1.351 as well as the B.1.1.7 variants in vitro.

scholarly journals Immunomodulatory Effects of Adipose Tissue-Derived Stem Cells on Concanavalin A-Induced Acute Liver Injury in Mice

Cell Medicine ◽  
2017 ◽  
Vol 9 (1-2) ◽  
pp. 21-33 ◽  
Author(s):  
Yasuma Yoshizumi ◽  
Hiroshi Yukawa ◽  
Ryoji Iwaki ◽  
Sanae Fujinaka ◽  
Ayano Kanou ◽  
...  
Keyword(s):  
Stem Cells ◽  
Adipose Tissue ◽  
Mouse Model ◽  
Cell Therapy ◽  
Concanavalin A ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Immunomodulatory Effects

Cell therapy with adipose tissue-derived stem cells (ASCs) is expected to be a candidate for the treatment of fulminant hepatic failure (FHF), which is caused by excessive immune responses. In order to evaluate the therapeutic effects of ASCs on FHF, the in vitro and in vivo immunomodulatory effects of ASCs were examined in detail in the mouse model. The in vitro effects of ASCs were examined by assessing their influence on the proliferation of lymphomononuclear cells (LMCs) stimulated with three kinds of mitogens: phorbol 12-myristate 13-acetate (PMA) plus ionomycin, concanavalin A (ConA), and lipopolysaccharide (LPS). The proliferation of LMCs was efficiently suppressed in a dose-dependent manner by ASCs in the cases of PMA plus ionomycin stimulation and ConA stimulation, but not in the case of LPS stimulation. The in vivo effects of transplanted ASCs were examined in the murine FHF model induced by ConA administration. The ALT levels and histological inflammatory changes in the ConA-administered mice were apparently relieved by the transplantation of ASCs. The analysis of mRNA expression patterns in the livers indicated that the expressions of the cytokines such as Il-6, Il-10, Ifn-γ, and Tnf-α, and the cell surface markers such as Cd3γ, Cd4, Cd8α, Cd11b, and Cd11c were downregulated in the ASC-transplanted mice. The immunomodulatory and therapeutic effects of ASCs were confirmed in the mouse model both in vitro and in vivo. These suggest that the cell therapy with ASCs is beneficial for the treatment of FHF.

scholarly journals WrpA Is an Atypical Flavodoxin Family Protein under Regulatory Control of the Brucella abortus General Stress Response System

2016 ◽  
Vol 198 (8) ◽  
pp. 1281-1293 ◽  
Author(s):  
Julien Herrou ◽  
Daniel M. Czyż ◽  
Jonathan W. Willett ◽  
Hye-Sook Kim ◽  
Gekleng Chhor ◽  
...  
Keyword(s):  
Stress Response ◽  
Mouse Model ◽  
Binding Site ◽  
Brucella Abortus ◽  
Chronic Infection ◽  
Content Type ◽  
General Stress Response ◽  
General Stress

ABSTRACTThe general stress response (GSR) system of the intracellular pathogenBrucella abortuscontrols the transcription of approximately 100 genes in response to a range of stress cues. The core genetic regulatory components of the GSR are required forB. abortussurvival under nonoptimal growth conditionsin vitroand for maintenance of chronic infection in anin vivomouse model. The functions of the majority of the genes in the GSR transcriptional regulon remain undefined.bab1_1070is among the most highly regulated genes in this regulon: its transcription is activated 20- to 30-fold by the GSR system under oxidative conditionsin vitro. We have solved crystal structures of Bab1_1070 and demonstrate that it forms a homotetrameric complex that resembles those of WrbA-type NADH:quinone oxidoreductases, which are members of the flavodoxin protein family. However,B. abortusWrbA-relatedprotein (WrpA) does not bind flavin cofactors with a high affinity and does not function as an NADH:quinone oxidoreductasein vitro. Soaking crystals with flavin mononucleotide (FMN) revealed a likely low-affinity binding site adjacent to the canonical WrbA flavin binding site. Deletion ofwrpA(ΔwrpA) does not compromise cell survival under acute oxidative stressin vitroor attenuate infection in cell-based or mouse models. However, a ΔwrpAstrain does elicit increased splenomegaly in a mouse model, suggesting that WrpA modulatesB. abortusinteraction with its mammalian host. Despite high structural homology with canonical WrbA proteins, we propose thatB. abortusWrpA represents a functionally distinct member of the diverse flavodoxin family.IMPORTANCEBrucella abortusis an etiological agent of brucellosis, which is among the most common zoonotic diseases worldwide. The general stress response (GSR) regulatory system ofB. abortuscontrols the transcription of approximately 100 genes and is required for maintenance of chronic infection in a murine model; the majority of GSR-regulated genes remain uncharacterized. We presentin vitroandin vivofunctional and structural analyses of WrpA, whose expression is strongly induced by GSR under oxidative conditions. Though WrpA is structurally related to NADH:quinone oxidoreductases, it does not bind redox cofactors in solution, nor does it exhibit oxidoreductase activityin vitro. However, WrpA does affect spleen inflammation in a murine infection model. Our data provide evidence that WrpA forms a new functional class of WrbA/flavodoxin family proteins.

scholarly journals MyoviridaePhagePDXKills EnteroaggregativeEscherichia coliwithout Human Microbiome Dysbiosis

2018 ◽  
Author(s):  
Leah C. S. Cepko ◽  
Eliotte E. Garling ◽  
Madeline J. Dinsdale ◽  
William P. Scott ◽  
Loralee Bandy ◽  
...  
Keyword(s):  
Mouse Model ◽  
Human Microbiome ◽  
Diversity Indices ◽  
Fecal Bacteria ◽  
Dependent Manner ◽  
Anaerobic Culture ◽  
Human Microbiota ◽  
Α Diversity

AbstractPurposeTo identify therapeutic a bacteriophage that kills diarrheagenic enteroaggregativeEscherichia coli(EAEC) while leaving the human microbiome intact.MethodologyPhages from wastewater in Portland, OR, were screened for bacteriolytic activity using an overlay assay, and isolated in a sequential procedure to enrich for the recognition of core bacterial antigens. Electron microscopy and genome sequencing were performed to classify the isolated phage, and the host range was determined by spot tests and plaque assays. One-step growth curves and time-kill assays were conducted to characterize the life cycle of the phage, and to interrogate the multiplicity of infection (MOI) necessary for killing. A mouse model of infection was used to determine whether the phage could be used therapeutically against EAECin vivo. Anaerobic culture in the presence of human fecal bacteria determined whether the phage could kill EAECin vitro, and to assess whether the microbiome had been altered.ResultsThe isolated phage, termedEscherichia virus PDX, is a member of the strictly lyticMyoviridaefamily of viruses. PhagePDXkilled EAEC isolate EN1E-0227, a case-associated isolate from a child in rural Tennessee, in a dose-dependent manner, and also formed plaques on case-associated clinical EAEC isolates from Columbian children suffering from diarrhea. A single dose ofPDX, at a MOI of 100, one day post infection, reduced the population of recovered EAEC isolate EN1E-0227 bacteria in fecal pellets in a mouse model of colonization, over a five-day period. PhagePDXalso killed EAEC EN1E-0227 when cultured anaerobicallyin vitroin the presence of human fecal bacteria. While the addition of EAEC EN1E-0227 reduced the α-diversity of the human microbiota, that of the cultures with either feces alone, feces with EAEC andPDX, or with just thePDXphage were not different statistically, as measured by Chao1 and Shannon diversity indices. Additionally, β-diversity and differential abundance analyses show that conditions withPDXadded were not different from feces alone, but all groups were significantly different from feces + EAEC.ConclusionsThe strictly bacteriolytic,Myoviridae Escherichia virus PDXkilled EAEC isolate EN1E-0227 bacteria bothin vivoandin vitro, while simultaneously not altering the diversity of normal human microbiota in anaerobic culture. Thus, thePDXphage could be part of an effective therapeutic intervention for children in developing countries who suffer from acute, or persistent EAEC-mediated diarrhea, and to help reduce the serious effects of environmental enteropathy. Because the emerging pathogen EAEC is now the second leading cause of traveler’s diarrhea,PDXcould also provide therapeutic relief for these individuals, particularly in light of the growing crisis of antibiotic resistances.

A Novel BRD4 Inhibitor CA2 Suppresses MM Cell Proliferation in an Orthotopic Myeloma Mouse Model

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4722-4722
Author(s):  
Natsuki Imayoshi ◽  
Makoto Yoshioka ◽  
Susumu Nakata ◽  
Jay Chauhan ◽  
Yoko Kado ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Mouse Model ◽  
Medical Center ◽  
Subcutaneous Administration ◽  
Vehicle Control ◽  
Care Organization ◽  
Dependent Manner ◽  
Proliferation In Vitro

Abstract We previously demonstrated that a novel BRD4 inhibitor, CA2, suppressed MM cell proliferation in vitro in the 57th ASH Annual Meeting. In the present study, we investigated the inhibitory mechanisms of CA2 on myeloma cell proliferation in vitro and in vivo. CA2 has a unique quinolinone core and inhibited the proliferation of MM cell lines, MM.1s, AMO-1, NCI-H929, OPM-2, and IM-9. As expected, CA2 suppressed the expression of c-MYC mRNA and protein in a dose- and time-dependent manner. In addition, CA2 decreased BRD4 binding to c-MYC promoters and c-MYC enhancers in a ChIP assay using IM-9 cells and an anti-BRD4 antibody (Figure 1). Flow cytometric Annexin-V/propodium iodide staining analyses demonstrated that CA2 induced apoptosis in MM cells. Taken together, CA2 may induce apoptosis in MM cells by inhibiting the binding of BRD4 to promoters and enhancers of c-MYC gene thereby suppressing c-MYC expression. We next performed a pharmacokinetic study of CA2 in mice. Half-lives (t1/2) of CA2 following oral administration (50 mg/kg) and subcutaneous administration (10 mg/kg) were approximately 61 min and 139.8 min, respectively. The values of area under the curve for oral and subcutaneous administrations were 179,000 hr·ng/mL and 31,000 hr·ng/mL, respectively. Lastly, we assessed the in vivo effects of CA2 using orthotopic MM mouse model, where IM-9Luc cells were inoculated intravenously through the tail veins of SCID mice that had received 2 Gy of irradiation. The mice in the vehicle-control arm died of MM by approximately 30 days after transplantation; IM-9Luc cells engrafted in many organs including bone marrow of spine and femurs, kidneys, ovaries, and stomach. CA2 treatment consisting of 20 cycles of 100 mg/kg/day b.i.d. oral dosing resulted in reduced luminescence intensity of IM-9Luc cells and prolonged the survival of mice (mean 33.7 days) compared to the vehicle-control arm (mean 27.1 days; p=0.029, Figure 2). In conclusion, CA2 is a BRD4 inhibitor with a novel structure and could be a promising molecular targeting-agent against MM. Disclosures Yoshioka: ConverGene LLC: Employment. Kado:Japan Community Health Care Organization Kyoto Kuramaguchi Medical Center: Employment. Strovel:ConverGene LLC: Employment.

scholarly journals Immunization with Pneumococcal Surface Protein K of Nonencapsulated Streptococcus pneumoniae Provides Protection in a Mouse Model of Colonization

2015 ◽  
Vol 22 (11) ◽  
pp. 1146-1153 ◽  
Author(s):  
Lance E. Keller ◽  
Xiao Luo ◽  
Justin A. Thornton ◽  
Keun-Seok Seo ◽  
Bo Youn Moon ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Mouse Model ◽  
Pneumococcal Disease ◽  
Surface Protein ◽  
Antibody Isotype ◽  
Content Type ◽  
Mouse Immunization ◽  
Escape Mutants

ABSTRACTCurrent vaccinations are effective against encapsulated strains ofStreptococcus pneumoniae, but they do not protect against nonencapsulatedStreptococcus pneumoniae(NESp), which is increasing in colonization and incidence of pneumococcal disease. Vaccination with pneumococcal proteins has been assessed for its ability to protect against pneumococcal disease, but several of these proteins are not expressed by NESp. Pneumococcal surface protein K (PspK), an NESp virulence factor, has not been assessed for immunogenic potential or host modulatory effects. Mammalian cytokine expression was determined in anin vivomouse model and in anin vitrocell culture system. Systemic and mucosal mouse immunization studies were performed to determine the immunogenic potential of PspK. Murine serum and saliva were collected to quantitate specific antibody isotype responses and the ability of antibody and various proteins to inhibit epithelial cell adhesion. Host cytokine response was not reduced by PspK. NESp was able to colonize the mouse nasopharynx as effectively as encapsulated pneumococci. Systemic and mucosal immunization provided protection from colonization by PspK-positive (PspK+) NESp. Anti-PspK antibodies were recovered from immunized mice and significantly reduced the ability of NESp to adhere to human epithelial cells. A protein-based pneumococcal vaccine is needed to provide broad protection against encapsulated and nonencapsulated pneumococci in an era of increasing antibiotic resistance and vaccine escape mutants. We demonstrate that PspK may serve as an NESp target for next-generation pneumococcal vaccines. Immunization with PspK protected against pneumococcal colonization, which is requisite for pneumococcal disease.

scholarly journals Protection of Antigen-Primed Effector T Cells From Glucocorticoid-Induced Apoptosis in Cell Culture and in a Mouse Model of Multiple Sclerosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Jasmina Bier ◽  
Sebastian M. Steiger ◽  
Holger M. Reichardt ◽  
Fred Lühder
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
Mouse Model ◽  
Clinical Symptoms ◽  
Effector T Cells ◽  
Differential Sensitivity ◽  
Dependent Manner ◽  
Induced Apoptosis

Induction of T cell apoptosis constitutes a major mechanism by which therapeutically administered glucocorticoids (GCs) suppress inflammation and associated clinical symptoms, for instance in multiple sclerosis (MS) patients suffering from an acute relapse. The sensitivity of T cells to GC action depends on their maturation and activation status, but the precise effect of antigen-priming in a pathological setting has not been explored. Here we used transgenic and congenic mouse models to compare GC-induced apoptosis between naïve and antigen-specific effector T cells from mice immunized with a myelin peptide. Antigen-primed effector T cells were protected from the pro-apoptotic activity of the synthetic GC dexamethasone in a dose-dependent manner, which resulted in their accumulation relative to naïve T cells in vitro and in vivo. Notably, the differential sensitivity of T cells to GC-induced apoptosis correlated with their expression level of the anti-apoptotic proteins Bcl-2 and Bcl-XL and a loss of the mitochondrial membrane potential. Moreover, accumulation of antigen-primed effector T cells following GC treatment in vitro resulted in an aggravated disease course in an adoptive transfer mouse model of MS in vivo, highlighting the clinical relevance of the observed phenomenon. Collectively, our data indicate that antigen-priming influences the T cells’ sensitivity to therapeutically applied GCs in the context of inflammatory diseases.

scholarly journals Demonstration of N , N -Dimethyldithiocarbamate as a Copper-Dependent Antibiotic against Multiple Upper Respiratory Tract Pathogens

2021 ◽  
Author(s):  
Sanjay V. Menghani ◽  
Angela Rivera ◽  
Miranda Neubert ◽  
James R. Hagerty ◽  
Lourdes Lewis ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Streptococcus Pneumoniae ◽  
Respiratory Tract ◽  
Dependent Manner ◽  
Upper Respiratory Tract ◽  
Coccidioides Posadasii ◽  
Content Type ◽  
Upper Respiratory

With the rise of antibiotic resistance, approaches that add new antimicrobials to the current repertoire are vital. Here, we investigate putative and known copper ionophores in an attempt to intoxicate bacteria and use ionophore/copper synergy, and we ultimately find success with N , N -dimethyldithiocarbamate (DMDC). We show that DMDC has in vitro efficacy in a copper-dependent manner and kills pathogens across three different kingdoms, Streptococcus pneumoniae ( Sr. pneumoniae ), Coccidioides posadasii , and Schistosoma mansoni , and in vivo efficacy against Sr . pneumoniae .

scholarly journals 3BDO Inhibits proliferation, epithelial–mesenchymal transition (EMT) and stemness via suppressing survivin in human glioblastoma cells

2021 ◽  
Author(s):  
zhaotao wang ◽  
yongping Li ◽  
minyi liu ◽  
danmin chen ◽  
yunxiang ji ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Mouse Model ◽  
Molecular Mechanisms ◽  
Migration And Invasion ◽  
Therapeutic Effects ◽  
Dependent Manner ◽  
Mesenchymal Transition ◽  
Xenograft Mouse Model

Abstract BackgroundGlioblastoma (GBM) is a tumor of the central nervous system carries an extremely poor prognosis. Unfortunately, it also is the most frequently encountered tumor in this region. These tumors arise from glioblastoma stem cells (GSCs), which are glioma cells that are known to possess high degrees of stemness. GBM invades through the process of EMT, which features loss of cell differentiation and polarity. Survivin is a type of apoptotic inhibitor that has been characterized in several malignancies such as glioma. Normal tissues rarely express survivin. On the other hand, 3-benzyl-5-((2-nitrophenoxy) methyl) dihydrofuran-2(3H)-one (3BDO) represents an autophagy inhibitor and activates the mTOR pathway. It has been reported that 3BDO shows anti-cancer activities in lung carcinoma. However, the effects of 3BDO on GBM reminds unknown. Therefore, the purpose of this study was to explore the role and molecular mechanisms that 3BDO mediates in GBM.MethodCCK-8 experiments and clone formation assay were performed to detect the cell proliferation. Transwell assay was conducted to examined cell migration and invasion. Western blotting and immunofluorescence staining was used to analyze protein expression levels. Xenograft mouse model was used to evaluate the effect of 3BDO in vivo.ResultsWe found that 3BDO inhibited U87 and U251 cell proliferation in a dose-dependent manner. Additonally, 3BDO decreased the sphere formation and stemness markers (sox2, nestin and CD133) in GSCs. 3BDO also inhibited migration, invasion and suppressed EMT markers (N-cadherin, vimentin and snail) in GBM cells. Moreover, we found that 3BDO downregulated survivin expression of survivin both in GBM cells (U87, U251) and GSCs. Furthermore, overexpression of survivin reduced the therapeutic effects of 3BDO on GBM cell EMT, invasion, migration and proliferation, as well as decreased stemness in GSCs. Finally, we demonstrated that 3BDO inhibited tumor growth in a tumor xenograft mouse model constructed using U87 cells. Similar to the in vitro findings, 3BDO diminished suvivin expression, stemness and levels of EMT makers in vivo.Conclusionsour results demonstrated that 3BDO repressed GBM via downregulating survivin-mediated stemness and EMT both in vitro and in vivo.

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