m bovis bcg pasteur 1173p2  (Pasteur Institute)

Journal: PLOS One

Article Title: Leveraging a synthetic biology approach to enhance BCG-mediated expansion of Vγ9Vδ2 T cells

doi: 10.1371/journal.pone.0343925

Figure Lengend Snippet: A. The MEP pathway of isoprenoid biosynthesis. Shown in green are end products and shunts; shown in red is HMBPP, the activator of Vγ9Vδ2 T cells. Genes encoding enzymatic steps are as follows: dxs , DOXP synthase; dxr , DOXP reductoisomerase; ispD , MEP cytidylyltransferase; ispE , CDP-ME kinase; ispF , MEcPP synthase; gcpE , HMBPP synthase; ispH , HMBPP reductase; idi , isopentenyl diphosphate isomerase. B. Silencing of MEP genes is lethal in BCG. Targeting of dxr and ispH by inducible CRISPR interference was lethal in BCG. Target genes, including folC and a dCas9 only plasmid as controls, were transcriptionally repressed by the addition ATc (B) . CFU without (Kan25) and with (Kan25-ATc200) induction were counted and analyzed via two-way ANOVA followed by Sidak’s multiple comparisons (B; *p ≤ 0.05, ****p < 0.0001). Targeting of folC, dxr , or ispH caused a significant decrease in viability, while the dCas9-only control experienced notably less death (C; *p ≤ 0.05, **p ≤ 0.01, ***p < 0.001, one sample t test). Shown is mean ± SD. N = 3 biological replicates. C. Synteny analysis reveals naturally occurring gene pair biases to leverage in our synthetic platform. Several gene pairs are frequently found in proximity to each other across mycobacteria: dxs1 + ispH ; dxr + ispG/gcpE ; ispD + ispF ; and occasionally ispH + ispG/gcpE . Synteny analysis was performed on 353 mycobacterial genomes using each gene as a query to generate this 7x7 matrix of co-occurrence.

Article Snippet: Mycobacterium tuberculosis variant bovis BCG Pasteur (ATCC 35734) was routinely grown in Middlebrook 7H9 liquid medium or 7H10 agar (Difco) supplemented with 10% OADC (oleic acid-albumin-dextrose-catalase) and 0.05% Tween80.

Techniques: CRISPR, Plasmid Preparation, Control

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: The expression of RBMX2 in EBL cells infected by ( A ) M. bovis and ( B ) M. bovis Bacillus Calmette–Guérin (BCG) was analyzed by real-time quantitative polymerase chain reaction (RT-qPCR). Data were represented by fold expression relative to uninfected cells. ( C–E ) The expression of RBMX2 mRNA in ( E ) BoMac cells, ( F ) A549 cells, and ( G ) bovine lung alveolar primary cells infected by M. bovis was analyzed via RT-qPCR. Data were represented by fold expression relative to uninfected cells. ( F ) The expression of RBMX2 in EBL cells infected by M. bovis was analyzed by WB. Data were represented by fold expression relative to uninfected cells. ( G ) Detection of the ability of different RBMX2 knockout site monoclonal EBL cells against M. bovis infection by CCK-8 assay. Data were represented by the absorbance value relative to WT EBL cells after M. bovis infection. ( H ) Detection of the ability of RBMX2 slicing H1299 cells against M. bovis infection by CCK-8 assay. Data were represented by the absorbance value relative to Sh-NC H1299 cells after M. bovis infection. One- and two-way ANOVA were used to determine the statistical significance of differences between different groups. Ns presents no significance; *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 1—source data 1. Original western blots for panel F, indicating the relevant bands. Figure 1—source data 2. Original files for western blot analysis displayed in panel F.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Expressing, Infection, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Knock-Out, CCK-8 Assay, Western Blot

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) The different knockout sites of RBMX2 EBL cells were observed by sequencing compared to the bovine RBMX2 sequence or WT EBL cells. ( B ) Observation of the effect of RBMX2 knockout on cell cycle of EBL cells by flow cytometry assay. Data are represented as the G0/G1 and S phase relative to WT EBL cells. ( C ) The change in cell number of RBMX2 knockout and silence following 96 and 120 hr of M. bovis infection was observed via crystal violet assay in EBL cells and H1299 cells, respectively. Data were represented as the cell number relative to WT EBL cells and H1299-ShNC cells after M. bovis infection. Data were representative of at least three independent experiments.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Knock-Out, Sequencing, Flow Cytometry, Infection, Crystal Violet Assay

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) The heatmap illustrates some genes that had been all enriched in RBMX2 knockout and WT EBL cells after M. bovis infection in 0 (2 hpi post-gentamicin, recorded as 0 hpi), 24, and 48 hpi. Red represents upregulated genes, and blue represents downregulated genes. Each group represented three independent samples. ( B ) Gene Ontology (GO) analysis of all enriched genes in 0 (2 hpi post-gentamicin, recorded as 0 hpi), 24, and 48 hpi. Data were represented as all enriched pathways in RBMX2 knockout EBL cells relative to WT EBL cells after M. bovis infection thrice. ( C ) Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of all enriched genes in 0 (2 hpi post-gentamicin, recorded as 0 hpi), 24, and 48 hpi. Data were represented as all enriched pathways in RBMX2 knockout EBL cells relative to WT EBL cells after M. bovis infection in three times. ( D ) GO analysis of all enriched proteins in 48 hpi. Data were represented as all enriched pathways in RBMX2 knockout EBL cells relative to WT EBL cells after M. bovis infection. ( E ) KEGG analysis of all enriched genes in 48 hpi. Data were represented as all enriched pathways in RBMX2 knockout EBL cells relative to WT EBL cells after M. bovis infection (MOI 20) in three times. ( F ) Identification of the expression of related genes mRNA enriched by real-time quantitative polymerase chain reaction (RT-qPCR). Data were represented as the fold expression in RBMX2 knockout EBL cells relative to WT EBL cells after M. bovis infection. Two-way ANOVA was used to determine the statistical significance of differences between different groups. Ns presents no significance; **p < 0.01, and ***p < 0.001 indicate statistically significant differences.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Knock-Out, Infection, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: Gene Ontology (GO) analysis of enriched genes of ( A ) 0, ( B ) 24, and ( C ) 48 hr after M. bovis infection, respectively. Data were in RBMX2 knockout EBL cells relative to WT EBL cells with M. bovis infection. The changes of these pathways from cell junction-related pathways in 0 hpi to cell proliferation and differentiation-related pathways in 48 hpi. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of enriched genes of ( D ) 0, ( E ) 24 and ( F ) 48 hpi after M. bovis infection, respectively. Data were in RBMX2 knockout EBL cells relative to WT EBL cells with M. bovis infection. The changes of these pathways from inflammation-related pathways in 0 hpi to cancer-related pathways in 48 hpi.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Infection, Knock-Out

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A, B ) A volcano map illustrating the transcriptional enrichment genes after WT EBL cells with M. bovis infection. Data were relative to WT EBL cells without M. bovis infection. ( C ) Identification of the expression of related genes mRNA enriched by real-time quantitative polymerase chain reaction (RT-qPCR). Data were represented as the fold expression in 24/48 hpi relative to 0 hpi (2 hpi post-gentamicin, recorded as 0 hpi).

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Infection, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis was conducted to identify the ( A ) downregulated and ( B ) upregulated pathways among the enriched genes after M. bovis infection of WT EBL cells. Data were relative to WT EBL cells without M. bovis infection. The expression of epithelial cells tight junction-related ( C ) mRNAs ( TJP1 , CLDN-5 , CLDN-7 , and OCLN ) and ( D ) proteins (ZO-1, CLDN-5, and OCLN) were assessed after M. bovis infection of WT EBL cells via real-time quantitative polymerase chain reaction (RT-qPCR) and WB. Data were relative to WT EBL cells without M. bovis infection. ( E ) Cell adhesion ratio was evaluated via cell adhesion assay after WT EBL cells were infected with M. bovis using high-content imaging. Data were relative to WT EBL cells without M. bovis infection. Scale bar: 20 μm. ( F, G ) The expression of epithelial tight junction-related ( F ) mRNAs ( TJP1 , CLDN-5 , and OCLN ) and ( G ) proteins (ZO-1, CLDN-5, and OCLN) in RBMX2 knockout EBL cells after M. bovis infection through real-time quantitative polymerase chain reaction (RT-qPCR) and WB. Data were relative to WT EBL cells with M. bovis infection. ( H ) Cell adhesion assay was conducted to assess the cell adhesion ratio of RBMX2 knockout EBL cells after infection with M. bovis . Data were relative to WT EBL cells with M. bovis infection. Scale bar: 20 μm. ( I ) Expression of inflammatory factors-related factors ( IL-6 , IL-1β , and TNF ) was assessed after RBMX2 knockout EBL cells infected by M. bovis . Data were relative to WT EBL cells with M. bovis infection. T -test and two-way ANOVA were used to determine the statistical significance of differences between different groups. Ns presents no significance; *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 3—source data 1. Original western blots for panel D, indicating the relevant bands. Figure 3—source data 2. Original files for western blot analysis displayed in panel D. Figure 3—source data 3. Original western blots for panel G, indicating the relevant bands. Figure 3—source data 4. Original files for western blot analysis displayed in panel G.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Infection, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Cell Adhesion Assay, Imaging, Knock-Out, Western Blot

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A, B ) Activation of the MAPK pathway-related protein and p65 protein was activated after RBMX2 knockout and WT EBL cells infected by M. bovis via WB. Data were relative to WT EBL cells with M. bovis infection. ( C, D ) Expression of tight junction-related proteins (ZO-1, CLDN-5, and OCLN) was assessed in RBMX2 knockout EBL cells treated with three p38/p65/JNK pathways activators after M. bovis infection via WB. Data were relative to RBMX2 knockout EBL cells untreated activators with M. bovis infection. ( E, F ) Evaluate the impact of three p38/p65/JNK pathways activators on the ratio of intercellular adhesion via cell adhesion assay. Data were relative to RBMX2 knockout EBL cells untreated activators with M. bovis infection. Scale bar: 20 μm. ( G, H ) Evaluate the silencing efficiency of small interfering RNA (siRNA) on p65 protein expression and its impact on the expression of ZO-1, CLDN-5, and OCLN proteins through WB. Data were relative to siRNA-NC in WT EBL cells with M. bovis infection. ( I ) The effect of p65 silencing on the invasive ability of M. bovis in WT EBL cells. Data were relative to siRNA-NC in WT EBL cells with M. bovis infection. ( H ) The effect of RBMX2 on the nuclear translocation of p65 protein after M. bovis infection using WB. β-Actin presents cytosol and Lamin A/C presents nucleus. Data were relative to RBMX2 knockout EBL cells after M. bovis infection. ( K ) The effect of RBMX2 on the nuclear translocation of p65 protein after BCG infection using high-content real-time imaging. Using the pCMV-EGFP-p65 plasmid, transfect RBMX2 knockout and WT EBL cells. The nucleus is stained with blue fluorescence. Data were relative to WT EBL cells without BCG infection. One- and two-way ANOVA were used to determine the statistical significance of differences between different groups. Ns presents no significance; *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 4—source data 1. Original western blots for panel A, indicating the relevant bands. Figure 4—source data 2. Original files for western blot analysis displayed in panel A. Figure 4—source data 3. Original western blots for panel C, indicating the relevant bands. Figure 4—source data 4. Original files for western blot analysis displayed in panel C. Figure 4—source data 5. Original western blots for panel G, indicating the relevant bands. Figure 4—source data 6. Original files for western blot analysis displayed in panel G. Figure 4—source data 7. Original western blots for panel J, indicating the relevant bands. Figure 4—source data 8. Original files for western blot analysis displayed in panel J.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Activation Assay, Knock-Out, Infection, Expressing, Cell Adhesion Assay, Small Interfering RNA, Translocation Assay, Imaging, Plasmid Preparation, Staining, Fluorescence, Western Blot

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A–C ) The impact of M. bovis on the adhesion, invasion, and intracellular survival of RBMX2 knockout and WT EBL cells through plate counting. Data were relative to WT EBL cells after M. bovis infection. ( D–F ) The impact of M. bovis on the adhesion, invasion, and intracellular survival of Sh-NC and Sh-RBMX2 H1299 cells through plate counting. Data were relative to H1299 ShNC cells after M. bovis infection. One- and two-way ANOVA were used to determine the statistical significance of differences between different groups. Ns presents no significance; *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Knock-Out, Infection

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A, B ) The impact of M. bovis on the adhesion and invasion of RBMX2 knockout EBL cells following treatment with related-pathway activators, verified by plate counting. Data were relative to RBMX2 knockout EBL cells untreated by activators. ( C, D ) The impact of M. bovis Bacillus Calmette–Guérin (BCG) and M. smegmatis on the adhesion of RBMX2 knockout EBL cells through plate counting assay. Data were relative to WT EBL cells after BCG and M. smegmatis infection. ( E, F ) The impact of M. bovis BCG and M. smegmatis on the invasion of RBMX2 knockout EBL cells through plate counting assay. Data were relative to WT EBL cells after BCG and M. smegmatis infection. ( G, H ) The impact of Salmonella and E. coli on the adhesion of RBMX2 knockout EBL cells through plate counting assay. Data were relative to WT EBL cells after Salmonella and E. coli infection. ( I, J ) The impact of Salmonella and E. coli on the invasion of RBMX2 knockout EBL cells through plate counting assay. Data were relative to WT EBL cells after Salmonella and E. coli infection. One- and two-way ANOVA were used to determine the statistical significance of differences between different groups. Ns presents no significance; *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Knock-Out, Infection

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) A comparative analysis of the functional domains of the RBMX2 protein across ten different species.( B ) Analyzing the expression patterns of RBMX2 in pan-cancer using TIMER2.0 cancer database. ( C ) The expression of RBMX2 in different lung cancer cells and normal lung epithelial cells via real-time quantitative polymerase chain reaction (RT-qPCR). Data were relative to normal lung epithelial cells (BEAS-2B). ( D, E ) The expression of RBMX2 in lung cancer clinical tissues via IF. RBMX2 is stained with yellow fluorescence, and the nucleus is stained with blue fluorescence. Data were relative to pericancerous lung tissues. Scale bar: 5000 μm. ( F ) The expression of RBMX2 and p65 in lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC) clinical tissues via IF. RBMX2 is stained with red fluorescence, p65 is stained with green fluorescence, and the nucleus is stained with blue fluorescence. Data were relative to normal lung tissues. Scale bar: 100 μm.( G ) Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of differential metabolite enrichment pathways in RBMX2 knockout EBL cells compared to WT EBL cells after M. bovis infection. ( H ) Dynamic distribution map of top 20 differential metabolites in RBMX2 knockout EBL cells compared to WT EBL cells after M. bovis infection. *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Functional Assay, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Staining, Fluorescence, Knock-Out, Infection

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) WB detection of changes in mesenchymal cell markers ( MMP-9 and N-cadherin ) after infection of EBL cells with different infection ratios (10, 20, and 50) of M. bovis . Data were relative to EBL cells without infection of M. bovis . ( B ) WB detection of the ability of M. bovis infection with macrophages (BoMac cells) to induce epithelial (EBL cells) mesenchymal transition. Data were relative to the addition of phosphate-buffered saline (PBS) in the upper chamber of the coculture model. ( C ) Staining the skeleton of EBL cells in coculture model after M. bovis infection using ghost pen cyclic peptides. The cytoskeleton is labeled with red fluorescence, and the nucleus is stained with blue fluorescence. Scale bar: 20 μm. ( D ) Epithelial–mesenchymal transition (EMT)-related mRNAs ( MMP-9 , N-cadherin , and E-cadherin ) expression was verified in coculture model EBL cells after M. bovis infection through real-time quantitative polymerase chain reaction (RT-qPCR). Data were relative to coculture model EBL cells without M. bovis infection. ( E ) The detection of EMT-related mRNAs ( MMP-9 , N-cadherin , and E-cadherin ) of RBMX2 knockout EBL cells after M. bovis-infected BoMac cells via RT-qPCR. Data were relative to WT EBL cells after M. bovis-infected BoMac cells. Two-way ANOVA was used to determine the statistical significance of differences between different groups. Ns presents no significance; **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 6—figure supplement 1—source data 1. Original western blots for panel A, indicating the relevant bands. Figure 6—figure supplement 1—source data 2. Original files for western blot analysis displayed in panel A. Figure 6—figure supplement 1—source data 3. Original western blots for panel B, indicating the relevant bands. Figure 6—figure supplement 1—source data 4. Original files for western blot analysis displayed in panel B.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Infection, Saline, Staining, Labeling, Fluorescence, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Knock-Out, Western Blot

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) A pattern diagram illustrated M. bovis -infected BoMac cells inducing EMT of EBL cells coculture model, drawing by BioRender. ( B ) Detection of IL-6 and TNF expression levels in EBL cells and BoMac cells infected with M. bovis using real-time quantitative polymerase chain reaction (RT-qPCR). Data were relative to BoMac cells without infection of M. bovis . ( C ) Detection of RBMX2 expression levels in a coculture model EBL cells after M. bovis infection using RT-qPCR. Data were relative to BoMac cells without infection of M. bovis . ( D ) Observation of morphological changes in EBL cells infected with M. bovis under electron microscopy. ( E ) EMT-related proteins ( MMP-9 , N-cadherin , and E-cadherin ) expression was verified in coculture model EBL cells after M. bovis infection through WB. Data were relative to coculture model EBL cells without M. bovis infection. ( F, G ) The impact of coculture model EBL cells after M. bovis infection on migration and invasion capacity was detected using Transwell assay. Data were relative to coculture model EBL cells without M. bovis infection. ( H ) The detection of epithelial–mesenchymal transition (EMT)-related proteins ( MMP-9 , N-cadherin , and E-cadherin ) of RBMX2 knockout EBL cells after M. bovis-infected BoMac cells via WB. Data were relative to WT EBL cells after M. bovis-infected BoMac cells. ( I ) The change in the migratory and invasive capabilities of RBMX2 knockout EBL cells after M. bovis-infected BoMac cells was assessed via Transwell assay. Data were relative to WT EBL cells after M. bovis-infected BoMac cells. ( J, K ) Validate the changes in migration abilities of RBMX2 knockout EBL cells after M. bovis-infected BoMac cells through wound-healing assay. Data were relative to WT EBL cells after M. bovis-infected BoMac cells. T -test and two-way ANOVA were used to determine the statistical significance of differences between different groups. Ns presents no significance; *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 6—source data 1. Original western blots for panel E, indicating the relevant bands. Figure 6—source data 2. Original files for western blot analysis displayed in panel E. Figure 6—source data 3. Original western blots for panel H, indicating the relevant bands. Figure 6—source data 4. Original files for western blot analysis displayed in panel H.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Infection, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Electron Microscopy, Migration, Transwell Assay, Knock-Out, Wound Healing Assay, Western Blot

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) EMT-related protein expression was verified in H1299 cells after M. bovis infection through WB. Data were relative to H1299 Sh-Con cells. ( B ) The change in the migratory and invasive capabilities of H1299 cells was assessed via Transwell assay. Data were relative to H1299 Sh-Con cells. Scale bar: 100 μm. ( C ) Activation of the MAPK pathway-related protein and p65 protein was activated after RBMX2 knockout and WT EBL cells infected by M. bovis in this coculture model via WB. Data were relative to WT EBL cells with M. bovis infection. Two-way ANOVA was used to determine the statistical significance of differences between different groups. *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 6—figure supplement 2—source data 1. Original western blots for panel A, indicating the relevant bands. Figure 6—figure supplement 2—source data 2. Original files for western blot analysis displayed in panel A. Figure 6—figure supplement 2—source data 3. Original western blots for panel C, indicating the relevant bands. Figure 6—figure supplement 2—source data 4. Original files for western blot analysis displayed in panel C.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Expressing, Infection, Transwell Assay, Activation Assay, Knock-Out, Western Blot

Journal: eLife

Article Title: RBMX2 links Mycobacterium bovis infection to epithelial–mesenchymal transition and lung cancer progression

doi: 10.7554/eLife.107132

Figure Lengend Snippet: ( A ) Evaluate the impact of pathway activations on expression of EMT-associated proteins ( MMP-9 , N-cadherin , and E-cadherin ) in RBMX2 knockout EBL cells after M. bovis-infected BoMac cells. Data were relative to RBMX2 knockout EBL cells untreated activators. ( B, C ) Evaluate the impact of pathway activations on the migratory and invasive capabilities of RBMX2 knockout EBL cells after M. bovis-infected BoMac cells via Transwell assay. Data were relative to RBMX2 knockout EBL cells untreated activators. ( D, E ) Evaluate the impact of pathway activations on the migratory capabilities of RBMX2 knockout EBL cells after M. bovis-infected BoMac cells via wound-healing assay. Data were relative to RBMX2 knockout EBL cells untreated activators. ( F, G ) The impact of p65 silencing on the expression of MMP-9 protein in WT EBL cells after M. bovis infection was assessed using WB. Data were relative to siRNA-NC in WT EBL cells with M. bovis infection. ( H ) Predicting the binding ability of RBMX2 promoter and p65 using molecular docking dynamics. ( I ) Verification of RBMX2 promoter region and p65 interaction using dual luciferase reporter system. ( I ) Using p65 antibody to precipitate p65 protein in EBL cells, and verification of RBMX2 promoter region and p65 interaction using ChIP-PCR assay. ( K ) Predicting potential binding sites for p65 and RBMX2 via protein docking. ( L ) Verification of MMP-9 promoter region and p65 interaction using dual luciferase reporter system. ( M ) Verification of MMP-9 promoter region and p65 interaction using ChIP-PCR. ( N ) Predicting potential binding sites for p65 and MMP-9 via protein docking. Two-way ANOVA was used to determine the statistical significance of differences between different groups. *p < 0.05, **p < 0.01, and ***p < 0.001 indicate statistically significant differences. Data were representative of at least three independent experiments. Figure 7—source data 1. Original western blots for panel A, indicating the relevant bands. Figure 7—source data 2. Original files for western blot analysis displayed in panel A. Figure 7—source data 3. Original western blots for panel F, indicating the relevant bands. Figure 7—source data 4. Original files for western blot analysis displayed in panel F.

Article Snippet: M. smegmatis mc ( ) 155 ( NC_008596.1 ) and M. bovis BCG-Pasteur (ATCC:35734) were generously provided by Professor Luiz Bermudez from Oregon State University.

Techniques: Expressing, Knock-Out, Infection, Transwell Assay, Wound Healing Assay, Binding Assay, Luciferase, Western Blot

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Timeline and interventions of the study. Eight groups of BALB/c mice were subcutaneously immunized twice at 21-day interval with 100 µL containing 10–15 µg of the TcTPE plus 10 4 CFU of wtBCG or BCG∆BCG1419c strain in 100 μL each to evaluate the effectiveness of the two BCG strains as adjuvants. All procedures were performed in duplicate, and data showed in the subsequent figures are representative of the two independent experiments. SC = subcutaneous, IP = intraperitoneal, BT = blood trypomastigotes, d = days, dpi = day post-infection, TcTPE = total protein extract from Trypanosoma cruzi .

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Infection

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Body weight of BCG-immunized or not immunized mice and infected or not with T. cruzi . Values show the mean with S.D. for each group and are representative of two independent experiments with equivalent results. The values were grouped into four stages (see timeline for further reference, ): baseline, from −10 d to 0 d; booster vaccination, from 1 d to 42 d; acute stage, from 2nd to 48th dpi and euthanasia, from 48th to 50th dpi. Values from each time were analyzed through multiple comparisons among groups by two-way ANOVA test followed by Dunnett’s post hoc test, and significant difference is shown (*) when p ≤ 0.05 compared to the group I (healthy mice), (**) when comparing the groups immunized with wtBCG or BCG∆BCG1419c as adjuvants without infection vs. their infected counterpart groups, and (***) when both adjuvants were compared between them. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Body temperature of mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Values were analyzed with the one-way ANOVA statistical test followed by the Tukey post hoc test at each time point (basal, booster vaccination, acute stage, and euthanasia), and a significant difference was demonstrated (*, p ≤ 0.05) when comparing against the group I (healthy mice), and (**, p ≤ 0.05) when comparing the groups immunized with wtBCG or BCG∆BCG1419c as adjuvants without infection (groups III and V, respectively) vs. their infected counterpart groups (groups IV and VI, respectively). The dotted lines show the normal temperature range in mice. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Physical condition record of mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Values show the mean with the S.D. of each group, which were analyzed with the nonparametric statistical test of Kruskal–Wallis followed by the Dunn post hoc test, and a significant difference was demonstrated (*, p ≤ 0.05) when comparing against the group I (healthy mice) and (**, p ≤ 0.05) when comparing against the group II (not vaccinated and infected mice). Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Humoral immune response in mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Values represent the mean ± S.D. of the optical densities (O.D.) of total IgG ( a ), IgG1 ( b ), and IgG2a ( c ) by group. Statistical analysis was performed using the one-way ANOVA test followed by the Tukey post hoc test. The results were considered significant when (*) p ≤ 0.05 when comparing against the II group infected mice, (**) p ≤ 0.05 when comparing the vaccinated group against its infected pair, and (***) p ≤ 0.05 when comparing the immunized/infected groups between them. Post-imm = post-immunization. The black line in ( a ) shows the cut-off value in the ELISA. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Formulation, Infection, Enzyme-linked Immunosorbent Assay, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Serum cytokines concentration of the Th1 immune profile in mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi. Values represent the mean ± S.D. of the cytokine levels (pg/mL) by group: TNF-α ( a ), IFN-γ ( b ), IL-1β ( c ), IL-2 ( d ), IL-12 ( e ), and IL-18 ( f ). The one-way ANOVA statistical test followed by the Tukey post hoc test was used to determine the existence of a significant difference (* p ≤ 0.05) with respect to the baseline concentration (preimmune sera pool of all mice or group II at post-imm/pre-infection time), (** p ≤ 0.05) of the euthanasia-time groups with respect to their corresponding ones in the post-vaccination/pre-infection time and (*** p ≤ 0.05) when the infected groups (vaccinated or not) were compared between them. Post-imm = post-immunization. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Concentration Assay, Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Serum cytokines concentration of the Th2 immune profile in mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi. Values represent the mean ± S.D. of the cytokine levels (pg/mL) by group: IL-4 ( a ), IL-6 ( b ), and IL-10 ( c ). One-way ANOVA statistical test followed by Tukey’s post hoc test was used to determine significant difference (* p ≤ 0.05) with respect to the baseline concentration (preimmune sera pool of all mice or group II at post-imm/pre-infection time) and (** p ≤ 0.05) when comparing the infected groups (vaccinated or not) among them. Post-imm = post-immunization. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Concentration Assay, Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Histopathological findings in tissue samples of heart ( a ) and skeletal muscle, ( b ) from mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Representative micrographs of each experimental group are shown. Green arrows indicate amastigotes nests. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected. Magnification: ×250 or ×400.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Scores of tissue damage degree from mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi. Values represent the mean ± S.D. of the heart ( a ) or skeletal muscle, ( b ) damage degree scores from each mouse per group at 30th dpi (acute phase of ChD). Kruskal–Wallis test was used for determining significant differences when (*) p ≤ 0.05 when comparing all groups with the I group healthy mice. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Groups of five 6-to 8-week-old, 20.8 ± 0.5 g on average, BALB/c mice were classified as follows and as shown in : (I) not-vaccinated, not-infected mice; (II) not-vaccinated, T. cruzi -infected mice; (III) total protein extract from Trypanosoma cruzi (TcTPE)-vaccinated mice and using the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, not infected; (IV) TcTPE-vaccinated mice plus the wild-type M. bovis BCG Pasteur ATCC 35734 strain as an adjuvant, and T. cruzi -infected; (V) TcTPE-vaccinated mice and using the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, not infected; (VI) TcTPE-vaccinated mice plus the M. bovis BCG∆BCG1419c mutant strain as an adjuvant, and T. cruzi -infected; (VII) TcTPE-vaccinated mice without any adjuvant, not infected; (VIII) TcTPE-vaccinated mice without any adjuvant, and T. cruzi -infected.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Timeline and interventions of the study. Eight groups of BALB/c mice were subcutaneously immunized twice at 21-day interval with 100 µL containing 10–15 µg of the TcTPE plus 10 4 CFU of wtBCG or BCG∆BCG1419c strain in 100 μL each to evaluate the effectiveness of the two BCG strains as adjuvants. All procedures were performed in duplicate, and data showed in the subsequent figures are representative of the two independent experiments. SC = subcutaneous, IP = intraperitoneal, BT = blood trypomastigotes, d = days, dpi = day post-infection, TcTPE = total protein extract from Trypanosoma cruzi .

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Infection

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Body weight of BCG-immunized or not immunized mice and infected or not with T. cruzi . Values show the mean with S.D. for each group and are representative of two independent experiments with equivalent results. The values were grouped into four stages (see timeline for further reference, ): baseline, from −10 d to 0 d; booster vaccination, from 1 d to 42 d; acute stage, from 2nd to 48th dpi and euthanasia, from 48th to 50th dpi. Values from each time were analyzed through multiple comparisons among groups by two-way ANOVA test followed by Dunnett’s post hoc test, and significant difference is shown (*) when p ≤ 0.05 compared to the group I (healthy mice), (**) when comparing the groups immunized with wtBCG or BCG∆BCG1419c as adjuvants without infection vs. their infected counterpart groups, and (***) when both adjuvants were compared between them. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Body temperature of mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Values were analyzed with the one-way ANOVA statistical test followed by the Tukey post hoc test at each time point (basal, booster vaccination, acute stage, and euthanasia), and a significant difference was demonstrated (*, p ≤ 0.05) when comparing against the group I (healthy mice), and (**, p ≤ 0.05) when comparing the groups immunized with wtBCG or BCG∆BCG1419c as adjuvants without infection (groups III and V, respectively) vs. their infected counterpart groups (groups IV and VI, respectively). The dotted lines show the normal temperature range in mice. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Physical condition record of mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Values show the mean with the S.D. of each group, which were analyzed with the nonparametric statistical test of Kruskal–Wallis followed by the Dunn post hoc test, and a significant difference was demonstrated (*, p ≤ 0.05) when comparing against the group I (healthy mice) and (**, p ≤ 0.05) when comparing against the group II (not vaccinated and infected mice). Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Humoral immune response in mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Values represent the mean ± S.D. of the optical densities (O.D.) of total IgG ( a ), IgG1 ( b ), and IgG2a ( c ) by group. Statistical analysis was performed using the one-way ANOVA test followed by the Tukey post hoc test. The results were considered significant when (*) p ≤ 0.05 when comparing against the II group infected mice, (**) p ≤ 0.05 when comparing the vaccinated group against its infected pair, and (***) p ≤ 0.05 when comparing the immunized/infected groups between them. Post-imm = post-immunization. The black line in ( a ) shows the cut-off value in the ELISA. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Formulation, Infection, Enzyme-linked Immunosorbent Assay, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Serum cytokines concentration of the Th1 immune profile in mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi. Values represent the mean ± S.D. of the cytokine levels (pg/mL) by group: TNF-α ( a ), IFN-γ ( b ), IL-1β ( c ), IL-2 ( d ), IL-12 ( e ), and IL-18 ( f ). The one-way ANOVA statistical test followed by the Tukey post hoc test was used to determine the existence of a significant difference (* p ≤ 0.05) with respect to the baseline concentration (preimmune sera pool of all mice or group II at post-imm/pre-infection time), (** p ≤ 0.05) of the euthanasia-time groups with respect to their corresponding ones in the post-vaccination/pre-infection time and (*** p ≤ 0.05) when the infected groups (vaccinated or not) were compared between them. Post-imm = post-immunization. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Concentration Assay, Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Serum cytokines concentration of the Th2 immune profile in mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi. Values represent the mean ± S.D. of the cytokine levels (pg/mL) by group: IL-4 ( a ), IL-6 ( b ), and IL-10 ( c ). One-way ANOVA statistical test followed by Tukey’s post hoc test was used to determine significant difference (* p ≤ 0.05) with respect to the baseline concentration (preimmune sera pool of all mice or group II at post-imm/pre-infection time) and (** p ≤ 0.05) when comparing the infected groups (vaccinated or not) among them. Post-imm = post-immunization. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Concentration Assay, Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Histopathological findings in tissue samples of heart ( a ) and skeletal muscle, ( b ) from mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi . Representative micrographs of each experimental group are shown. Green arrows indicate amastigotes nests. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected. Magnification: ×250 or ×400.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Formulation, Infection, Adjuvant

Journal: Microorganisms

Article Title: Vaccination with a Trypanosoma cruzi Protein Extract Plus BCG∆BCG1419c Promotes a Balanced Th1/Th2 Immune Profile That Improves Control of Acute Chagas Disease in BALB/c Mice

doi: 10.3390/microorganisms13112447

Figure Lengend Snippet: Scores of tissue damage degree from mice immunized with the vaccine formulation of TcTPE using M. bovis BCG strains as adjuvants and infected with T. cruzi. Values represent the mean ± S.D. of the heart ( a ) or skeletal muscle, ( b ) damage degree scores from each mouse per group at 30th dpi (acute phase of ChD). Kruskal–Wallis test was used for determining significant differences when (*) p ≤ 0.05 when comparing all groups with the I group healthy mice. Group I = not vaccinated/not infected; group II = not vaccinated/infected; group III = TcTPE-vaccinated + wtBCG as adjuvant/not infected; group IV = TcTPE-vaccinated + wtBCG as adjuvant/infected; group V = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/not infected; group VI = TcTPE-vaccinated + BCG∆BCG1419c as adjuvant/infected; group VII = TcTPE-vaccinated/not infected; group VIII = TcTPE-vaccinated/infected.

Article Snippet: Other authors observed that only male BALB/c mice vaccinated with M. bovis Danish 1331 BCG strain (BCG SSI) showed a significant weight loss of 20% [ ]; however, in the present study females BALB/c showed such effect with M. bovis Pasteur ATCC 35734 BCG strain.

Techniques: Formulation, Infection, Adjuvant