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Stimulatory effect of Echinacea purpurea extract on the trafficking activity of mouse dendritic cells: revealed by genomic and proteomic analyses.

Yin SY, Wang WH, Wang BX, Aravindaram K, Hwang PI, Wu HM, Yang NS - BMC Genomics (2010)

Bottom Line: The transcriptomic and proteomic effects of this phytoextract on mouse bone marrow-derived dendritic cells (BMDCs) were analyzed using primary cultures.Treatment of BMDCs with [BF/S+L/Ep] did not significantly influence the phenotypic maturation activity of dendritic cells (DCs).Moreover, the signaling networks and molecules highlighted here are potential targets for nutritional or clinical application of Echinacea or other candidate medicinal plants.

View Article: PubMed Central - HTML - PubMed

Affiliation: Agricultural Biotechnology Research Center, Academia Sinica, Taipei, Taiwan.

ABSTRACT

Background: Several Echinacea species have been used as nutraceuticals or botanical drugs for "immunostimulation", but scientific evidence supporting their therapeutic use is still controversial. In this study, a phytocompound mixture extracted from the butanol fraction (BF) of a stem and leaf (S+L) extract of E. purpurea ([BF/S+L/Ep]) containing stringently defined bioactive phytocompounds was obtained using standardized and published procedures. The transcriptomic and proteomic effects of this phytoextract on mouse bone marrow-derived dendritic cells (BMDCs) were analyzed using primary cultures.

Results: Treatment of BMDCs with [BF/S+L/Ep] did not significantly influence the phenotypic maturation activity of dendritic cells (DCs). Affymetrix DNA microarray and bioinformatics analyses of genes differentially expressed in DCs treated with [BF/S+L/Ep] for 4 or 12 h revealed that the majority of responsive genes were related to cell adhesion or motility (Cdh10, Itga6, Cdh1, Gja1 and Mmp8), or were chemokines (Cxcl2, Cxcl7) or signaling molecules (Nrxn1, Pkce and Acss1). TRANSPATH database analyses of gene expression and related signaling pathways in treated-DCs predicted the JNK, PP2C-α, AKT, ERK1/2 or MAPKAPK pathways as the putative targets of [BF/S+L/Ep]. In parallel, proteomic analysis showed that the expressions of metabolic-, cytoskeleton- or NF-κB signaling-related proteins were regulated by treatment with [BF/S+L/Ep]. In vitro flow cytometry analysis of chemotaxis-related receptors and in vivo cell trafficking assay further showed that DCs treated with [BF/S+L/Ep] were able to migrate more effectively to peripheral lymph node and spleen tissues than DCs treated as control groups.

Conclusion: Results from this study suggest that [BF/S+L/Ep] modulates DC mobility and related cellular physiology in the mouse immune system. Moreover, the signaling networks and molecules highlighted here are potential targets for nutritional or clinical application of Echinacea or other candidate medicinal plants.

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Proteomics analyses of E. purpurea phytocompound effects on mouse bone-marrow derived immature dendritic cells (iBMDCs). Effect of Echinacea purpurea extract on differential protein expression in iBMDCs was evaluated by 2D gel electrophoretic analysis. 500 μg of total cell proteins from various treatments of iBMDCs were applied to pH 5-8 IPG strips. After isofocusing and 2D gel separation, SYPRO Ruby was used to stain proteins. Images on gel were analyzed using PD-Quest software. Forty-two differentially-expressed proteins were trypsin-digested in situ, and 23 protein spots were identified by MALDI-TOF MS, with their locations in gel assigned a number. These protein ID numbers are listed in Table 4. A, Up-regulated and B, down-regulated protein spots are reported on a representative two-dimensional gel corresponding to protein expression profile of iBMDCs. The expression ratios are shown in bold print. Gels shown here are representative of thee independent experiments.
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Figure 4: Proteomics analyses of E. purpurea phytocompound effects on mouse bone-marrow derived immature dendritic cells (iBMDCs). Effect of Echinacea purpurea extract on differential protein expression in iBMDCs was evaluated by 2D gel electrophoretic analysis. 500 μg of total cell proteins from various treatments of iBMDCs were applied to pH 5-8 IPG strips. After isofocusing and 2D gel separation, SYPRO Ruby was used to stain proteins. Images on gel were analyzed using PD-Quest software. Forty-two differentially-expressed proteins were trypsin-digested in situ, and 23 protein spots were identified by MALDI-TOF MS, with their locations in gel assigned a number. These protein ID numbers are listed in Table 4. A, Up-regulated and B, down-regulated protein spots are reported on a representative two-dimensional gel corresponding to protein expression profile of iBMDCs. The expression ratios are shown in bold print. Gels shown here are representative of thee independent experiments.

Mentions: Using 2D gel electrophoresis, we were able to routinely obtain representative, high resolution, and highly reproducible 2D protein profiles of mouse DCs as putative proteomic maps (data not shown). Treatment of DCs with [BF/S+L/Ep] at 75 μg/mL resulted in significant changes in expression of some proteins in comparison to the solvent-treated mouse DC samples. Differentially-expressed proteins were then identified by MALDI-TOF-MS and in some cases subsequently analyzed with tandem MS/MS, after manual excision of these protein spots from gels. A total of 23 different known protein species were isolated and characterized by analysis with MALDI-TOF-MS peptide mass fingerprinting (PMF). Table 4 lists the changes in protein expression in DCs treated with [BF/S+L/Ep] in vitro for 12 h, as compared to solvent (vehicle) treatment (0.1% DMSO). Expression of metabolism, cytoskeleton or NF-κB-signaling related proteins were most affected by [BF/S+L/Ep]. 2D gel electrophoretic analyses showed increased levels of a group of specific proteins, such as annexin A4 and peroxiredoxin (Figure 4A). In contrast, the levels of some other proteins, such as macrophage capping protein, were reduced (Figure 4B). These proteins, which function in cytoskeleton organization, did not show a detectable change in mRNA levels after [BF/S+L/Ep] treatment.


Stimulatory effect of Echinacea purpurea extract on the trafficking activity of mouse dendritic cells: revealed by genomic and proteomic analyses.

Yin SY, Wang WH, Wang BX, Aravindaram K, Hwang PI, Wu HM, Yang NS - BMC Genomics (2010)

Proteomics analyses of E. purpurea phytocompound effects on mouse bone-marrow derived immature dendritic cells (iBMDCs). Effect of Echinacea purpurea extract on differential protein expression in iBMDCs was evaluated by 2D gel electrophoretic analysis. 500 μg of total cell proteins from various treatments of iBMDCs were applied to pH 5-8 IPG strips. After isofocusing and 2D gel separation, SYPRO Ruby was used to stain proteins. Images on gel were analyzed using PD-Quest software. Forty-two differentially-expressed proteins were trypsin-digested in situ, and 23 protein spots were identified by MALDI-TOF MS, with their locations in gel assigned a number. These protein ID numbers are listed in Table 4. A, Up-regulated and B, down-regulated protein spots are reported on a representative two-dimensional gel corresponding to protein expression profile of iBMDCs. The expression ratios are shown in bold print. Gels shown here are representative of thee independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3091753&req=5

Figure 4: Proteomics analyses of E. purpurea phytocompound effects on mouse bone-marrow derived immature dendritic cells (iBMDCs). Effect of Echinacea purpurea extract on differential protein expression in iBMDCs was evaluated by 2D gel electrophoretic analysis. 500 μg of total cell proteins from various treatments of iBMDCs were applied to pH 5-8 IPG strips. After isofocusing and 2D gel separation, SYPRO Ruby was used to stain proteins. Images on gel were analyzed using PD-Quest software. Forty-two differentially-expressed proteins were trypsin-digested in situ, and 23 protein spots were identified by MALDI-TOF MS, with their locations in gel assigned a number. These protein ID numbers are listed in Table 4. A, Up-regulated and B, down-regulated protein spots are reported on a representative two-dimensional gel corresponding to protein expression profile of iBMDCs. The expression ratios are shown in bold print. Gels shown here are representative of thee independent experiments.
Mentions: Using 2D gel electrophoresis, we were able to routinely obtain representative, high resolution, and highly reproducible 2D protein profiles of mouse DCs as putative proteomic maps (data not shown). Treatment of DCs with [BF/S+L/Ep] at 75 μg/mL resulted in significant changes in expression of some proteins in comparison to the solvent-treated mouse DC samples. Differentially-expressed proteins were then identified by MALDI-TOF-MS and in some cases subsequently analyzed with tandem MS/MS, after manual excision of these protein spots from gels. A total of 23 different known protein species were isolated and characterized by analysis with MALDI-TOF-MS peptide mass fingerprinting (PMF). Table 4 lists the changes in protein expression in DCs treated with [BF/S+L/Ep] in vitro for 12 h, as compared to solvent (vehicle) treatment (0.1% DMSO). Expression of metabolism, cytoskeleton or NF-κB-signaling related proteins were most affected by [BF/S+L/Ep]. 2D gel electrophoretic analyses showed increased levels of a group of specific proteins, such as annexin A4 and peroxiredoxin (Figure 4A). In contrast, the levels of some other proteins, such as macrophage capping protein, were reduced (Figure 4B). These proteins, which function in cytoskeleton organization, did not show a detectable change in mRNA levels after [BF/S+L/Ep] treatment.

Bottom Line: The transcriptomic and proteomic effects of this phytoextract on mouse bone marrow-derived dendritic cells (BMDCs) were analyzed using primary cultures.Treatment of BMDCs with [BF/S+L/Ep] did not significantly influence the phenotypic maturation activity of dendritic cells (DCs).Moreover, the signaling networks and molecules highlighted here are potential targets for nutritional or clinical application of Echinacea or other candidate medicinal plants.

View Article: PubMed Central - HTML - PubMed

Affiliation: Agricultural Biotechnology Research Center, Academia Sinica, Taipei, Taiwan.

ABSTRACT

Background: Several Echinacea species have been used as nutraceuticals or botanical drugs for "immunostimulation", but scientific evidence supporting their therapeutic use is still controversial. In this study, a phytocompound mixture extracted from the butanol fraction (BF) of a stem and leaf (S+L) extract of E. purpurea ([BF/S+L/Ep]) containing stringently defined bioactive phytocompounds was obtained using standardized and published procedures. The transcriptomic and proteomic effects of this phytoextract on mouse bone marrow-derived dendritic cells (BMDCs) were analyzed using primary cultures.

Results: Treatment of BMDCs with [BF/S+L/Ep] did not significantly influence the phenotypic maturation activity of dendritic cells (DCs). Affymetrix DNA microarray and bioinformatics analyses of genes differentially expressed in DCs treated with [BF/S+L/Ep] for 4 or 12 h revealed that the majority of responsive genes were related to cell adhesion or motility (Cdh10, Itga6, Cdh1, Gja1 and Mmp8), or were chemokines (Cxcl2, Cxcl7) or signaling molecules (Nrxn1, Pkce and Acss1). TRANSPATH database analyses of gene expression and related signaling pathways in treated-DCs predicted the JNK, PP2C-α, AKT, ERK1/2 or MAPKAPK pathways as the putative targets of [BF/S+L/Ep]. In parallel, proteomic analysis showed that the expressions of metabolic-, cytoskeleton- or NF-κB signaling-related proteins were regulated by treatment with [BF/S+L/Ep]. In vitro flow cytometry analysis of chemotaxis-related receptors and in vivo cell trafficking assay further showed that DCs treated with [BF/S+L/Ep] were able to migrate more effectively to peripheral lymph node and spleen tissues than DCs treated as control groups.

Conclusion: Results from this study suggest that [BF/S+L/Ep] modulates DC mobility and related cellular physiology in the mouse immune system. Moreover, the signaling networks and molecules highlighted here are potential targets for nutritional or clinical application of Echinacea or other candidate medicinal plants.

Show MeSH
Related in: MedlinePlus