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Triglyceride-Rich Lipoproteins Modulate the Distribution and Extravasation of Ly6C/Gr1(low) Monocytes.

Saja MF, Baudino L, Jackson WD, Cook HT, Malik TH, Fossati-Jimack L, Ruseva M, Pickering MC, Woollard KJ, Botto M - Cell Rep (2015)

Bottom Line: We observed an increased crawling and retention of the Gr1(low) monocytes at the endothelial interface and a marked accumulation of CD68(+) macrophages in several organs.Hypertriglyceridemia was accompanied by an increased expression of tissue, and plasma CCL4 and blood Gr1(low) monocyte depletion involved a pertussis-toxin-sensitive receptor axis.The behavior of these cells in response to dyslipidemia highlights the significant impact that high levels of triglyceride-rich lipoproteins may have on innate immune cells.

View Article: PubMed Central - PubMed

Affiliation: Centre for Complement and Inflammation Research, Division of Immunology and Inflammation, Department of Medicine, Imperial College London, Du Cane Road, London W12 ONN, UK.

No MeSH data available.


Related in: MedlinePlus

Chemokine Expression Triggered by P-407 Treatment(A–D) The effect of P-407 on mRNA expression of chemokine/cytokine was analyzed by RT-PCR. Relative expression of (A) CCL3, (B) CCL4, (C) CCL2, and (D) IL-6 to the housekeeping gene HPRT. Kidney, liver, and heart specimens from mice treated with P-407 were compared with those from PBS-treated animals at each time point.(E) CCL4 and CCL2 mRNA expression relative to HPRT in CD45+F4/80+ sorted tissue macrophages from liver, heart, and kidney after 14 days of P-407 or PBS treatment. Data are pooled from three animals per condition and representative of two independent experiments.(F) Plasma concentrations of CCL4 and CCL2 after 7, 14, and 28 days of P-407 or PBS treatment. Values represent mean ± SE, n = 4 mice per group.(G) Frequencies of Gr1high and Gr1low monocyte subsets in P-407-treated B6 mice injected with PT or PBS. PBS-treated mice were used as controls. Values represent mean ± SE, n = 3 mice per group. Results are representative of three independent experiments. Significant p values are indicated (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; unpaired t test). ND, nondetectable; ns, nonsignificant.(H) Transwell migration assay. Fluorescence-activated cell-sorted Gr1high and Gr1low monocytes were added to a transwell chamber for 2 hr in the presence of recombinant mouse CCL4 (1–1,000 ng/ml) or PBS (0). The number of migrated cells per field was quantified (five fields per sample); pool of n = 3 mice in triplicate. Data are mean ± SE. ∗∗∗p < 0.001 (unpaired t test).See also Figure S6.
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fig6: Chemokine Expression Triggered by P-407 Treatment(A–D) The effect of P-407 on mRNA expression of chemokine/cytokine was analyzed by RT-PCR. Relative expression of (A) CCL3, (B) CCL4, (C) CCL2, and (D) IL-6 to the housekeeping gene HPRT. Kidney, liver, and heart specimens from mice treated with P-407 were compared with those from PBS-treated animals at each time point.(E) CCL4 and CCL2 mRNA expression relative to HPRT in CD45+F4/80+ sorted tissue macrophages from liver, heart, and kidney after 14 days of P-407 or PBS treatment. Data are pooled from three animals per condition and representative of two independent experiments.(F) Plasma concentrations of CCL4 and CCL2 after 7, 14, and 28 days of P-407 or PBS treatment. Values represent mean ± SE, n = 4 mice per group.(G) Frequencies of Gr1high and Gr1low monocyte subsets in P-407-treated B6 mice injected with PT or PBS. PBS-treated mice were used as controls. Values represent mean ± SE, n = 3 mice per group. Results are representative of three independent experiments. Significant p values are indicated (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; unpaired t test). ND, nondetectable; ns, nonsignificant.(H) Transwell migration assay. Fluorescence-activated cell-sorted Gr1high and Gr1low monocytes were added to a transwell chamber for 2 hr in the presence of recombinant mouse CCL4 (1–1,000 ng/ml) or PBS (0). The number of migrated cells per field was quantified (five fields per sample); pool of n = 3 mice in triplicate. Data are mean ± SE. ∗∗∗p < 0.001 (unpaired t test).See also Figure S6.

Mentions: To investigate the mechanism(s) of the extravasation we quantified tissue mRNA expression of a large panel of chemokines/cytokines. As shown in Figure 6, this analysis showed a marked upregulation of CCL4 in the organs from the P-407-treated animals compared to those from the PBS controls. CCL2 and CCL3 also showed some increased expression. Of note, IL-6 mRNA expression showed a very different pattern and was reduced in the liver from P-407-treated animals (Figure 6D), confirming that the P-407 treatment did not elicit an overt inflammatory response. In keeping with the intravital data, the expression of CX3CL1 (fractalkine) was not affected by the P-407 treatment (Figure S6A). Similarly, other chemokines such as CXCL9, CXCL1, and CCL5 were expressed at similar levels in the two experimental groups (Figures S6B–S6D). As macrophages can produce a large amount of CCL4 (Maurer and von Stebut, 2004) and were increased in tissues following P-407 treatment, we hypothesized that they could represent a potential source of the CCL4. We therefore analyzed CCL4 mRNA expression in sorted CD45+F4/80+ tissue macrophages after 2 weeks of PBS or P-407 treatment. We found that macrophages from heart and liver of P-407-treated animals had a more than 2-fold increase in CCL4 expression compared to those from PBS-treated controls (Figure 6E). We observed only a slight increase in CCL2 expression in the liver macrophages from P-407-treated mice (Figure 6E). We then measured CCL2 and CCL4 plasma levels. In keeping with the gene expression data, we detected CCL4 only in the animals treated with P-407 and not in the PBS controls (Figure 6F), demonstrating that a hyper-TGRL environment could induce the production of this chemokine. We also detected an increase in CCL2 levels in the P-407 mice, but the increase reached statistical significance only at day 14 (Figure 6F).


Triglyceride-Rich Lipoproteins Modulate the Distribution and Extravasation of Ly6C/Gr1(low) Monocytes.

Saja MF, Baudino L, Jackson WD, Cook HT, Malik TH, Fossati-Jimack L, Ruseva M, Pickering MC, Woollard KJ, Botto M - Cell Rep (2015)

Chemokine Expression Triggered by P-407 Treatment(A–D) The effect of P-407 on mRNA expression of chemokine/cytokine was analyzed by RT-PCR. Relative expression of (A) CCL3, (B) CCL4, (C) CCL2, and (D) IL-6 to the housekeeping gene HPRT. Kidney, liver, and heart specimens from mice treated with P-407 were compared with those from PBS-treated animals at each time point.(E) CCL4 and CCL2 mRNA expression relative to HPRT in CD45+F4/80+ sorted tissue macrophages from liver, heart, and kidney after 14 days of P-407 or PBS treatment. Data are pooled from three animals per condition and representative of two independent experiments.(F) Plasma concentrations of CCL4 and CCL2 after 7, 14, and 28 days of P-407 or PBS treatment. Values represent mean ± SE, n = 4 mice per group.(G) Frequencies of Gr1high and Gr1low monocyte subsets in P-407-treated B6 mice injected with PT or PBS. PBS-treated mice were used as controls. Values represent mean ± SE, n = 3 mice per group. Results are representative of three independent experiments. Significant p values are indicated (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; unpaired t test). ND, nondetectable; ns, nonsignificant.(H) Transwell migration assay. Fluorescence-activated cell-sorted Gr1high and Gr1low monocytes were added to a transwell chamber for 2 hr in the presence of recombinant mouse CCL4 (1–1,000 ng/ml) or PBS (0). The number of migrated cells per field was quantified (five fields per sample); pool of n = 3 mice in triplicate. Data are mean ± SE. ∗∗∗p < 0.001 (unpaired t test).See also Figure S6.
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fig6: Chemokine Expression Triggered by P-407 Treatment(A–D) The effect of P-407 on mRNA expression of chemokine/cytokine was analyzed by RT-PCR. Relative expression of (A) CCL3, (B) CCL4, (C) CCL2, and (D) IL-6 to the housekeeping gene HPRT. Kidney, liver, and heart specimens from mice treated with P-407 were compared with those from PBS-treated animals at each time point.(E) CCL4 and CCL2 mRNA expression relative to HPRT in CD45+F4/80+ sorted tissue macrophages from liver, heart, and kidney after 14 days of P-407 or PBS treatment. Data are pooled from three animals per condition and representative of two independent experiments.(F) Plasma concentrations of CCL4 and CCL2 after 7, 14, and 28 days of P-407 or PBS treatment. Values represent mean ± SE, n = 4 mice per group.(G) Frequencies of Gr1high and Gr1low monocyte subsets in P-407-treated B6 mice injected with PT or PBS. PBS-treated mice were used as controls. Values represent mean ± SE, n = 3 mice per group. Results are representative of three independent experiments. Significant p values are indicated (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001; unpaired t test). ND, nondetectable; ns, nonsignificant.(H) Transwell migration assay. Fluorescence-activated cell-sorted Gr1high and Gr1low monocytes were added to a transwell chamber for 2 hr in the presence of recombinant mouse CCL4 (1–1,000 ng/ml) or PBS (0). The number of migrated cells per field was quantified (five fields per sample); pool of n = 3 mice in triplicate. Data are mean ± SE. ∗∗∗p < 0.001 (unpaired t test).See also Figure S6.
Mentions: To investigate the mechanism(s) of the extravasation we quantified tissue mRNA expression of a large panel of chemokines/cytokines. As shown in Figure 6, this analysis showed a marked upregulation of CCL4 in the organs from the P-407-treated animals compared to those from the PBS controls. CCL2 and CCL3 also showed some increased expression. Of note, IL-6 mRNA expression showed a very different pattern and was reduced in the liver from P-407-treated animals (Figure 6D), confirming that the P-407 treatment did not elicit an overt inflammatory response. In keeping with the intravital data, the expression of CX3CL1 (fractalkine) was not affected by the P-407 treatment (Figure S6A). Similarly, other chemokines such as CXCL9, CXCL1, and CCL5 were expressed at similar levels in the two experimental groups (Figures S6B–S6D). As macrophages can produce a large amount of CCL4 (Maurer and von Stebut, 2004) and were increased in tissues following P-407 treatment, we hypothesized that they could represent a potential source of the CCL4. We therefore analyzed CCL4 mRNA expression in sorted CD45+F4/80+ tissue macrophages after 2 weeks of PBS or P-407 treatment. We found that macrophages from heart and liver of P-407-treated animals had a more than 2-fold increase in CCL4 expression compared to those from PBS-treated controls (Figure 6E). We observed only a slight increase in CCL2 expression in the liver macrophages from P-407-treated mice (Figure 6E). We then measured CCL2 and CCL4 plasma levels. In keeping with the gene expression data, we detected CCL4 only in the animals treated with P-407 and not in the PBS controls (Figure 6F), demonstrating that a hyper-TGRL environment could induce the production of this chemokine. We also detected an increase in CCL2 levels in the P-407 mice, but the increase reached statistical significance only at day 14 (Figure 6F).

Bottom Line: We observed an increased crawling and retention of the Gr1(low) monocytes at the endothelial interface and a marked accumulation of CD68(+) macrophages in several organs.Hypertriglyceridemia was accompanied by an increased expression of tissue, and plasma CCL4 and blood Gr1(low) monocyte depletion involved a pertussis-toxin-sensitive receptor axis.The behavior of these cells in response to dyslipidemia highlights the significant impact that high levels of triglyceride-rich lipoproteins may have on innate immune cells.

View Article: PubMed Central - PubMed

Affiliation: Centre for Complement and Inflammation Research, Division of Immunology and Inflammation, Department of Medicine, Imperial College London, Du Cane Road, London W12 ONN, UK.

No MeSH data available.


Related in: MedlinePlus