scholarly journals Migration of Dendritic Cells Facilitates Systemic Dissemination of Burkholderia pseudomallei

2014 ◽  
Vol 82 (10) ◽  
pp. 4233-4240 ◽  
Author(s):  
Natasha L. Williams ◽  
Jodie L. Morris ◽  
Catherine M. Rush ◽  
Natkunam Ketheesan
Keyword(s):  
Dendritic Cells ◽  
Burkholderia Pseudomallei ◽  
Content Type ◽  
Systemic Dissemination ◽  
Systemic Spread ◽  
Rapid Dissemination ◽  
Migration Capacity ◽  
Secondary Lymphoid Organs

ABSTRACTBurkholderia pseudomallei, the etiological agent for melioidosis, is an important cause of community-acquired sepsis in northern Australia and northeast Thailand. Due to the rapid dissemination of disease in acute melioidosis, we hypothesized that dendritic cells (DC) could act as a vehicle for dissemination ofB. pseudomallei. Therefore, this study investigated the effect ofB. pseudomalleiinfection on DC migration capacity and whether migration of DC enabled transportation ofB. pseudomalleifrom the site of infection.B. pseudomalleistimulated significantly increased migration of bone marrow-derived DC (BMDC), bothin vitroandin vivo, compared to uninfected BMDC. Furthermore, migration of BMDC enabled significantly increasedin vitrotrafficking ofB. pseudomalleiandin vivodissemination ofB. pseudomalleito secondary lymphoid organs and lungs of C57BL/6 mice. DC within the footpad infection site of C57BL/6 mice also internalizedB. pseudomalleiand facilitated dissemination. Although DC have previously been shown to kill intracellularB. pseudomalleiin vitro, the findings of this study demonstrate thatB. pseudomallei-infected DC facilitate the systemic spread of this pathogen.

scholarly journals Characterization of New Virulence Factors Involved in the Intracellular Growth and Survival of Burkholderia pseudomallei

2015 ◽  
Vol 84 (3) ◽  
pp. 701-710 ◽  
Author(s):  
Madeleine G. Moule ◽  
Natasha Spink ◽  
Sam Willcocks ◽  
Jiali Lim ◽  
José Afonso Guerra-Assunção ◽  
...  
Keyword(s):  
Burkholderia Pseudomallei ◽  
Insertion Site ◽  
Wild Type ◽  
Content Type ◽  
Growth And Survival ◽  
New Genes ◽  
Insertion Sites

Burkholderia pseudomallei, the causative agent of melioidosis, has complex and poorly understood extracellular and intracellular lifestyles. We used transposon-directed insertion site sequencing (TraDIS) to retrospectively analyze a transposon library that had previously been screened through a BALB/c mouse model to identify genes important for growth and survivalin vivo. This allowed us to identify the insertion sites and phenotypes of negatively selected mutants that were previously overlooked due to technical constraints. All 23 unique genes identified in the original screen were confirmed by TraDIS, and an additional 105 mutants with various degrees of attenuationin vivowere identified. Five of the newly identified genes were chosen for further characterization, and clean, unmarkedbpsl2248,tex,rpiR,bpsl1728, andbpss1528deletion mutants were constructed from the wild-type strain K96243. Each of these mutants was testedin vitroandin vivoto confirm their attenuated phenotypes and investigate the nature of the attenuation. Our results confirm that we have identified new genes important toin vivovirulence with roles in different stages ofB. pseudomalleipathogenesis, including extracellular and intracellular survival. Of particular interest, deletion of the transcription accessory protein Tex was shown to be highly attenuating, and thetexmutant was capable of providing protective immunity against challenge with wild-typeB. pseudomallei, suggesting that the genes identified in our TraDIS screen have the potential to be investigated as live vaccine candidates.

scholarly journals Enpp2 Expression by Dendritic Cells Is a Key Regulator in Migration

Biomedicines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1727
Author(s):  
Jun-Ho Lee ◽  
So-Yeon Choi ◽  
Soo-Yeoun Park ◽  
Nam-Chul Jung ◽  
Kyung-Eun Noh ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Lymph Nodes ◽  
Cancer Vaccines ◽  
Biological Effects ◽  
Migratory Capacity ◽  
Migration Capacity ◽  
Antigen Presenting ◽  
Si Rna ◽  
Secondary Lymphoid Organs

Enpp2 is an enzyme that catalyzes the conversion of lysophosphatidylcholine (LPC) to lysophosphatidic acid (LPA), which exhibits a wide variety of biological functions. Here, we examined the biological effects of Enpp2 on dendritic cells (DCs), which are specialized antigen-presenting cells (APCs) characterized by their ability to migrate into secondary lymphoid organs and activate naïve T-cells. DCs were generated from bone marrow progenitors obtained from C57BL/6 mice. Enpp2 levels in DCs were regulated using small interfering (si)RNA or recombinant Enpp2. Expression of Enpp2 in LPS-stimulated mature (m)DCs was high, however, knocking down Enpp2 inhibited mDC function. In addition, the migratory capacity of mDCs increased after treatment with rmEnpp2; this phenomenon was mediated via the RhoA-mediated signaling pathway. Enpp2-treated mDCs showed a markedly increased capacity to migrate to lymph nodes in vivo. These findings strongly suggest that Enpp2 is necessary for mDC migration capacity, thereby increasing our understanding of DC biology. We postulate that regulating Enpp2 improves DC migration to lymph nodes, thus improving the effectiveness of cancer vaccines based on DC.

scholarly journals Role of RelA and SpoT in Burkholderia pseudomallei Virulence and Immunity

2012 ◽  
Vol 80 (9) ◽  
pp. 3247-3255 ◽  
Author(s):  
Claudia M. Müller ◽  
Laura Conejero ◽  
Natasha Spink ◽  
Matthew E. Wand ◽  
Gregory J. Bancroft ◽  
...  
Keyword(s):  
Burkholderia Pseudomallei ◽  
Galleria Mellonella ◽  
Bacterial Species ◽  
Virulence Gene ◽  
Live Attenuated Vaccine ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Control And Prevention

ABSTRACTBurkholderia pseudomalleiis a Gram-negative soil bacterium and the causative agent of melioidosis, a disease of humans and animals. It is also listed as a category B bioterrorism threat agent by the U.S. Centers for Disease Control and Prevention, and there is currently no melioidosis vaccine available. Small modified nucleotides such as the hyperphosphorylated guanosine molecules ppGpp and pppGpp play an important role as signaling molecules in prokaryotes. They mediate a global stress response under starvation conditions and have been implicated in the regulation of virulence and survival factors in many bacterial species. In this study, we created arelA spoTdouble mutant inB. pseudomalleistrain K96243, which lacks (p)ppGpp-synthesizing enzymes, and investigated its phenotypein vitroandin vivo. TheB. pseudomalleiΔrelAΔspoTmutant displayed a defect in stationary-phase survival and intracellular replication in murine macrophages. Moreover, the mutant was attenuated in theGalleria mellonellainsect model and in both acute and chronic mouse models of melioidosis. Vaccination of mice with the ΔrelAΔspoTmutant resulted in partial protection against infection with wild-typeB. pseudomallei. In summary, (p)ppGpp signaling appears to represent an essential component of the regulatory network governing virulence gene expression and stress adaptation inB. pseudomallei, and the ΔrelAΔspoTmutant may be a promising live-attenuated vaccine candidate.

scholarly journals Proteomic Analysis of the Burkholderia pseudomallei Type II Secretome Reveals Hydrolytic Enzymes, Novel Proteins, and the Deubiquitinase TssM

2014 ◽  
Vol 82 (8) ◽  
pp. 3214-3226 ◽  
Author(s):  
Mary N. Burtnick ◽  
Paul J. Brett ◽  
David DeShazer
Keyword(s):  
Burkholderia Pseudomallei ◽  
Signal Sequence ◽  
Hydrolytic Enzymes ◽  
Opportunistic Pathogen ◽  
Etiologic Agent ◽  
Type Ii ◽  
Secretion Systems ◽  
Content Type

ABSTRACTBurkholderia pseudomallei, the etiologic agent of melioidosis, is an opportunistic pathogen that harbors a wide array of secretion systems, including a type II secretion system (T2SS), three type III secretion systems (T3SS), and six type VI secretion systems (T6SS). The proteins exported by these systems provideB. pseudomalleiwith a growth advantagein vitroandin vivo, but relatively little is known about the full repertoire of exoproducts associated with each system. In this study, we constructed deletion mutations ingspDandgspE, T2SS genes encoding an outer membrane secretin and a cytoplasmic ATPase, respectively. The secretion profiles ofB. pseudomalleiMSHR668 and its T2SS mutants were noticeably different when analyzed by SDS-PAGE. We utilized liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify proteins present in the supernatants ofB. pseudomalleiMSHR668 andB. pseudomalleiΔgspDgrown in rich and minimal media. The MSHR668 supernatants contained 48 proteins that were either absent or substantially reduced in the supernatants of ΔgspDstrains. Many of these proteins were putative hydrolytic enzymes, including 12 proteases, two phospholipases, and a chitinase. Biochemical assays validated the LC-MS/MS results and demonstrated that the export of protease, phospholipase C, and chitinase activities is T2SS dependent. Previous studies had failed to identify the mechanism of secretion of TssM, a deubiquitinase that plays an integral role in regulating the innate immune response. Here we present evidence that TssM harbors an atypical signal sequence and that its secretion is mediated by the T2SS. This study provides the first in-depth characterization of theB. pseudomalleiT2SS secretome.

scholarly journals In VitroandIn VivoStudies of Monoclonal Antibodies with Prominent Bactericidal Activity against Burkholderia pseudomallei and Burkholderia mallei

2011 ◽  
Vol 18 (5) ◽  
pp. 825-834 ◽  
Author(s):  
Shimin Zhang ◽  
Shaw-Huey Feng ◽  
Bingjie Li ◽  
Hyung-Yong Kim ◽  
Joe Rodriguez ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Bactericidal Activity ◽  
Burkholderia Pseudomallei ◽  
Protective Efficacy ◽  
Lethal Dose ◽  
Burkholderia Mallei ◽  
Content Type ◽  
Bactericidal Activities

ABSTRACTOur laboratory has developed more than a hundred mouse monoclonal antibodies (MAbs) againstBurkholderia pseudomalleiandBurkholderia mallei. These antibodies have been categorized into different groups based on their specificities and the biochemical natures of their target antigens. The current study first examined the bactericidal activities of a number of these MAbs by anin vitroopsonic assay. Then, thein vivoprotective efficacy of selected MAbs was evaluated using BALB/c mice challenged intranasally with a lethal dose of the bacteria. The opsonic assay using dimethyl sulfoxide-treated human HL-60 cells as phagocytes revealed that 19 out of 47 tested MAbs (40%) have prominent bactericidal activities againstB. pseudomalleiand/orB. mallei. Interestingly, all MAbs with strong opsonic activities are those with specificity against either the capsular polysaccharides (PS) or the lipopolysaccharides (LPS) of the bacteria. On the other hand, none of the MAbs reacting to bacterial proteins or glycoproteins showed prominent bactericidal activity. Further study revealed that the antigenic epitopes on either the capsular PS or LPS molecules were readily available for binding in intact bacteria, while the epitopes on proteins/glycoproteins were less accessible to the MAbs. Ourin vivostudy showed that four MAbs reactive to either the capsular PS or LPS were highly effective in protecting mice against lethal bacterial challenge. The result is compatible with that of ourin vitrostudy. The MAbs with the highest protective efficacy are those reactive to either the capsular PS or LPS of theBurkholderiabacteria.

scholarly journals Exploring Aztreonam in Combination with Ceftazidime-Avibactam or Meropenem-Vaborbactam as Potential Treatments for Metallo- and Serine-β-Lactamase-Producing Enterobacteriaceae

2019 ◽  
Vol 63 (12) ◽  
Author(s):  
M. Biagi ◽  
T. Wu ◽  
M. Lee ◽  
S. Patel ◽  
D. Butler ◽  
...  
Keyword(s):  
Treatment Options ◽  
Triple Combination ◽  
Content Type ◽  
Similar Activity ◽  
Rapid Dissemination ◽  
Resistant Strains ◽  
Number Of Treatment ◽  
Time Kill

ABSTRACT Metallo-β-lactamase (MBL)-producing Enterobacteriaceae, particularly those that coharbor serine β-lactamases, are a serious emerging public health threat given their rapid dissemination and the limited number of treatment options. Preclinical and anecdotal clinical data support the use of aztreonam in combination with ceftazidime-avibactam against these pathogens, but other aztreonam-based combinations have not been explored. The objective of this study was to evaluate the in vitro activity and compare synergy between aztreonam in combination with ceftazidime-avibactam and meropenem-vaborbactam against serine and MBL-producing Enterobacteriaceae via time-kill analyses. Eight clinical Enterobacteriaceae strains (4 Escherichia coli and 4 Klebsiella pneumoniae) coproducing NDM and at least one serine β-lactamase were used for all experiments. Drugs were tested alone, in dual β-lactam combinations, and in triple-drug combinations against all strains. All strains were resistant to ceftazidime-avibactam and meropenem-vaborbactam and 7/8 (87.5%) strains were resistant to aztreonam. Aztreonam combined with ceftazidime-avibactam was synergistic against all 7 aztreonam-resistant strains. Aztreonam combined with meropenem-vaborbactam was synergistic against all aztreonam-resistant strains with the exception of an OXA-232-producing K. pneumoniae strain. Neither triple combination was synergistic against the aztreonam-susceptible strain. These data suggest that aztreonam plus meropenem-vaborbactam has similar activity to aztreonam plus ceftazidime-avibactam against Enterobacteriaceae producing NDM and other non-OXA-48-like serine β-lactamases. Confirmation of these findings in future in vitro and in vivo models is warranted.

scholarly journals Tumor cytotoxicity and immunogenicity of a novel V-jet neon plasma source compared to the kINPen

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Lea Miebach ◽  
Eric Freund ◽  
Stefan Horn ◽  
Felix Niessner ◽  
Sanjeev Kumar Sagwal ◽  
...  
Keyword(s):  
Cell Death ◽  
Dendritic Cells ◽  
Cancer Cells ◽  
Treatment Time ◽  
Plasma Source ◽  
In Vivo Model ◽  
Antitumor Effects ◽  
Plasma Device

AbstractRecent research indicated the potential of cold physical plasma in cancer therapy. The plethora of plasma-derived reactive oxygen and nitrogen species (ROS/RNS) mediate diverse antitumor effects after eliciting oxidative stress in cancer cells. We aimed at exploiting this principle using a newly designed dual-jet neon plasma source (Vjet) to treat colorectal cancer cells. A treatment time-dependent ROS/RNS generation induced oxidation, growth retardation, and cell death within 3D tumor spheroids were found. In TUM-CAM, a semi in vivo model, the Vjet markedly reduced vascularized tumors' growth, but an increase of tumor cell immunogenicity or uptake by dendritic cells was not observed. By comparison, the argon-driven single jet kINPen, known to mediate anticancer effects in vitro, in vivo, and in patients, generated less ROS/RNS and terminal cell death in spheroids. In the TUM-CAM model, however, the kINPen was equivalently effective and induced a stronger expression of immunogenic cancer cell death (ICD) markers, leading to increased phagocytosis of kINPen but not Vjet plasma-treated tumor cells by dendritic cells. Moreover, the Vjet was characterized according to the requirements of the DIN-SPEC 91315. Our results highlight the plasma device-specific action on cancer cells for evaluating optimal discharges for plasma cancer treatment.

scholarly journals The Antitumor Activity of Sodium Selenite Alone and in Combination with Gemcitabine in Pancreatic Cancer: An In Vitro and In Vivo Study

Cancers ◽  
2021 ◽  
Vol 13 (13) ◽  
pp. 3169
Author(s):  
Kevin Doello ◽  
Cristina Mesas ◽  
Francisco Quiñonero ◽  
Gloria Perazzoli ◽  
Laura Cabeza ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Sodium Selenite ◽  
Molecular Mechanisms ◽  
Combined Therapy ◽  
Significant Inhibition ◽  
In Vivo Studies ◽  
C57bl Mice ◽  
Migration Capacity

Sodium selenite acts by depleting enzymes that protect against cellular oxidative stress. To determine its effect alone or in combination with gemcitabine (GMZ) in pancreatic cancer, we used PANC-1 and Pan02 cell lines and C57BL mice bearing a Pan02-generated tumor. Our results demonstrated a significant inhibition of pancreatic cancer cell viability with the use of sodium selenite alone and a synergistic effect when associated with GMZ. The molecular mechanisms of the antitumor effect of sodium selenite alone involved apoptosis-inducing factor (AIF) and the expression of phospho-p38 in the combined therapy. In addition, sodium selenite alone and in association with GMZ significantly decreased the migration capacity and colony-forming ability, reduced tumor activity in multicellular tumor spheroids (MTS) and decreased sphere formation of cancer stem cells. In vivo studies demonstrated that combined therapy not only inhibited tumor growth (65%) compared to the untreated group but also relative to sodium selenite or GMZ used as monotherapy (up to 40%), increasing mice survival. These results were supported by the analysis of C57BL/6 albino mice bearing a Pan02-generated tumor, using the IVIS system. In conclusion, our results showed that sodium selenite is a potential agent for the improvement in the treatment of pancreatic cancer and should be considered for future human clinical trials.

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