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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One <t>Way</t> <t>ANOVA</t> with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.
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Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One Way ANOVA with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Infection by the Helminth Parasite Fasciola hepatica Requires Rapid Regulation of Metabolic, Virulence, and Invasive Factors to Adjust to Its Mammalian Host *

doi: 10.1074/mcp.RA117.000445

Figure Lengend Snippet: Analysis of NEJ-specific F. hepatica cathepsin L (FhCL3) and B (FhCLB) cysteine proteases. A , Immunolocalization of FhCL3 in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence (FITC staining) within the NEJ gut. B , Immunolocalization of FhCB in NEJ by CSLM, over a time-course of 48 h, represented by green fluorescence within the NEJ gut (FITC staining). Time-course: (i) at excystment, (ii) 1 h post-excystment, (iii) 6 h post-excystment, (iv) 10 h post-excystment, (v) 24 h post-excystment. All specimens were counter-stained with phalloidin- TRITC to stain muscle tissue (red fluorescence) and provide structure. Scale bars = 20 μ m . C , Relative fold expression of cathepsin L (FhCL3) and B (FhCB1/CB3 and FhCB2) genes over a time-course of 48 h normalized to expression at NEJ excystment relative to a GAPDH reference, with S.E. Statistical analysis was carried out using One Way ANOVA with Tukey's post hoc test ( p < 0.05: *; p < 0.01: **; p < 0.001: ***). D , Graphical representation of gene transcription (i), protein abundance within the somatic proteome (ii) and protein abundance within the secretome (iii). Relative expression/abundance is shown by a blue to red scale, depicting low to high levels of expression/abundance, respectively.

Article Snippet: Statistical analysis was carried out by One Way ANOVA (version 6.00 for Windows, GraphPad Software); p value <0.05 was deemed statistically significant.

Techniques: Fluorescence, Staining, Expressing, Quantitative Proteomics

Proliferation of neoblast-like cells during the first 48 h post excystment. A , Incorporation of 5-ethynyl-2-deoxyuridine (EdU) by the proliferative neoblast-like cells highlighted by green fluorescence; (i) NEJ following 24 h culture with Edu at 4 °C, (ii) NEJ following 24 h culture at 4 °C followed by 24 h culture with Edu at 37 °C, (iii) NEJ following 48 h culture with Edu at 37 °C. Scale bars = 20 μ m . B , Graphical representation of the expression of genes associated with neoblast-like cells in transcripts per million (TPM) across the F. hepatica lifecycle, displayed on a log2 scale. C , Number of neoblast cells identified after (a) NEJ incubated for 48 h with 5-ethynyl-2-deoxyuridine (EdU) at 37 °C, (b) NEJ cultured for 48 h at 37 °C, with the addition of Edu for the last 24 h of culture, (c) NEJ cultured for 48 h, the first 24 h at 4 °C, following by the remaining 24 h culture at 37 °C with the addition of Edu. The differences between these groups were statistically significant ( p < 0.001: ***). D , Relative fold expression of genes associated with neoblast-like cells normalized to expression at 24 h relative to a GAPDH reference, performed in duplicate, with S.E. Statistical analysis was carried out using One Way ANOVA with Tukey's post hoc test ( p < 0.01: **; p < 0.001: ***).

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Infection by the Helminth Parasite Fasciola hepatica Requires Rapid Regulation of Metabolic, Virulence, and Invasive Factors to Adjust to Its Mammalian Host *

doi: 10.1074/mcp.RA117.000445

Figure Lengend Snippet: Proliferation of neoblast-like cells during the first 48 h post excystment. A , Incorporation of 5-ethynyl-2-deoxyuridine (EdU) by the proliferative neoblast-like cells highlighted by green fluorescence; (i) NEJ following 24 h culture with Edu at 4 °C, (ii) NEJ following 24 h culture at 4 °C followed by 24 h culture with Edu at 37 °C, (iii) NEJ following 48 h culture with Edu at 37 °C. Scale bars = 20 μ m . B , Graphical representation of the expression of genes associated with neoblast-like cells in transcripts per million (TPM) across the F. hepatica lifecycle, displayed on a log2 scale. C , Number of neoblast cells identified after (a) NEJ incubated for 48 h with 5-ethynyl-2-deoxyuridine (EdU) at 37 °C, (b) NEJ cultured for 48 h at 37 °C, with the addition of Edu for the last 24 h of culture, (c) NEJ cultured for 48 h, the first 24 h at 4 °C, following by the remaining 24 h culture at 37 °C with the addition of Edu. The differences between these groups were statistically significant ( p < 0.001: ***). D , Relative fold expression of genes associated with neoblast-like cells normalized to expression at 24 h relative to a GAPDH reference, performed in duplicate, with S.E. Statistical analysis was carried out using One Way ANOVA with Tukey's post hoc test ( p < 0.01: **; p < 0.001: ***).

Article Snippet: Statistical analysis was carried out by One Way ANOVA (version 6.00 for Windows, GraphPad Software); p value <0.05 was deemed statistically significant.

Techniques: Fluorescence, Expressing, Incubation, Cell Culture