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Leishmania infection modulates beta-1 integrin activation and alters the kinetics of monocyte spreading over fibronectin.

Figueira CP, Carvalhal DG, Almeida RA, Hermida Md, Touchard D, Robert P, Pierres A, Bongrand P, dos-Santos WL - Sci Rep (2015)

Bottom Line: Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue.This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody.These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis.

View Article: PubMed Central - PubMed

Affiliation: Fundação Oswaldo Cruz-Bahia, Centro de Pesquisas Gonçalo Moniz, Brazilian Ministry of Health, Salvador, Brazil.

ABSTRACT
Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue. In this work, we studied the early stages of bond formation between VLA4 and fibronectin, measured the kinetics of membrane alignment and the monocyte cytoplasm spreading area over a fibronectin-coated surface, and studied the expression of high affinity integrin epitope in uninfected and Leishmania-infected human monocytes. Our results show that the initial VLA4-mediated interaction of Leishmania-infected monocyte with a fibronectin-coated surface is preserved, however, the later stage, leukocyte spreading over the substrate is abrogated in Leishmania-infected cells. The median of spreading area was 72 [55-89] μm(2) for uninfected and 41 [34-51] μm(2) for Leishmania-infected monocyte. This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody. After the initial contact with the fibronectrin-coated surface, uninfected monocyte quickly spread the cytoplasm at a 15 μm(2) s(-1) ratio whilst Leishmania-infected monocytes only made small contacts at a 5.5 μm(2) s(-1) ratio. The expression of high affinity epitope by VLA4 (from 39 ± 21% to 14 ± 3%); and LFA1 (from 37 ± 32% to 18 ± 16%) molecules was reduced in Leishmania-infected monocytes. These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis.

No MeSH data available.


Related in: MedlinePlus

Spreading of human monocyte cytoplasm on fibronectin after Leishmania infection.Peripheral blood monocytes were cultured with medium alone (A,C,D,F) or with medium containing Leishmania (B) or 3 μM latex beads (E) for 18 h. The cells were then allowed to adhere for 1 h to fibronectin-coated coverslips: (A) – Uninfected (control) monocytes; (B) – monocytes cultured with Leishmania; (C) – uninfected cells treated with 0.5 mM EDTA; (D) – uninfected cells treated with 0.5 mM MnCl2; (E) – uninfected cells cultured with latex beads; (F) – uninfected cells treated with anti-VLA4 antibody. The graph shows the area over which the cytoplasm of the monocytes had spread after the various treatments. The data are representative of six (G) and two (H,I) different experiments (Kruskal-Wallis test).
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f2: Spreading of human monocyte cytoplasm on fibronectin after Leishmania infection.Peripheral blood monocytes were cultured with medium alone (A,C,D,F) or with medium containing Leishmania (B) or 3 μM latex beads (E) for 18 h. The cells were then allowed to adhere for 1 h to fibronectin-coated coverslips: (A) – Uninfected (control) monocytes; (B) – monocytes cultured with Leishmania; (C) – uninfected cells treated with 0.5 mM EDTA; (D) – uninfected cells treated with 0.5 mM MnCl2; (E) – uninfected cells cultured with latex beads; (F) – uninfected cells treated with anti-VLA4 antibody. The graph shows the area over which the cytoplasm of the monocytes had spread after the various treatments. The data are representative of six (G) and two (H,I) different experiments (Kruskal-Wallis test).

Mentions: Because we did not observe changes in the initial step (rolling) of infected monocyte adherence to fibronectin, we used SEM to compare the cytoplasmic spreading of uninfected and Leishmania-infected human monocytes on fibronectin. Most of the monocytes cultured with medium alone displayed a flattened cell phenotype with extensive cytoplasmic spreading and irregular edges (Fig. 2A). Uninfected monocytes cultured with medium alone or cultured with 3 μm latex beads showed similar phenotypes (Fig. 2E), as did uninfected monocytes treated with Mn++ just before the adhesion assay (used as a control for high-affinity adhesion, Fig. 2D). On the other hand, most of the monocytes cultured with Leishmania had a rounded morphology with low levels of cytoplasmic spreading (Fig. 2B), which was similar to the morphology observed when the monocytes were treated with EDTA before the adhesion assay (used as a negative control for cytoplasmic spreading, Fig. 2C). The spread area (μm2) of the monocyte cytoplasm, 72 [55–89] (median [lower and upper quartiles]), was larger for the monocytes cultured with medium alone than for the monocytes cultured with Leishmania (49 [43–57]; Mann-Whitney test, P < 0.0001, Fig. 2G).


Leishmania infection modulates beta-1 integrin activation and alters the kinetics of monocyte spreading over fibronectin.

Figueira CP, Carvalhal DG, Almeida RA, Hermida Md, Touchard D, Robert P, Pierres A, Bongrand P, dos-Santos WL - Sci Rep (2015)

Spreading of human monocyte cytoplasm on fibronectin after Leishmania infection.Peripheral blood monocytes were cultured with medium alone (A,C,D,F) or with medium containing Leishmania (B) or 3 μM latex beads (E) for 18 h. The cells were then allowed to adhere for 1 h to fibronectin-coated coverslips: (A) – Uninfected (control) monocytes; (B) – monocytes cultured with Leishmania; (C) – uninfected cells treated with 0.5 mM EDTA; (D) – uninfected cells treated with 0.5 mM MnCl2; (E) – uninfected cells cultured with latex beads; (F) – uninfected cells treated with anti-VLA4 antibody. The graph shows the area over which the cytoplasm of the monocytes had spread after the various treatments. The data are representative of six (G) and two (H,I) different experiments (Kruskal-Wallis test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4528201&req=5

f2: Spreading of human monocyte cytoplasm on fibronectin after Leishmania infection.Peripheral blood monocytes were cultured with medium alone (A,C,D,F) or with medium containing Leishmania (B) or 3 μM latex beads (E) for 18 h. The cells were then allowed to adhere for 1 h to fibronectin-coated coverslips: (A) – Uninfected (control) monocytes; (B) – monocytes cultured with Leishmania; (C) – uninfected cells treated with 0.5 mM EDTA; (D) – uninfected cells treated with 0.5 mM MnCl2; (E) – uninfected cells cultured with latex beads; (F) – uninfected cells treated with anti-VLA4 antibody. The graph shows the area over which the cytoplasm of the monocytes had spread after the various treatments. The data are representative of six (G) and two (H,I) different experiments (Kruskal-Wallis test).
Mentions: Because we did not observe changes in the initial step (rolling) of infected monocyte adherence to fibronectin, we used SEM to compare the cytoplasmic spreading of uninfected and Leishmania-infected human monocytes on fibronectin. Most of the monocytes cultured with medium alone displayed a flattened cell phenotype with extensive cytoplasmic spreading and irregular edges (Fig. 2A). Uninfected monocytes cultured with medium alone or cultured with 3 μm latex beads showed similar phenotypes (Fig. 2E), as did uninfected monocytes treated with Mn++ just before the adhesion assay (used as a control for high-affinity adhesion, Fig. 2D). On the other hand, most of the monocytes cultured with Leishmania had a rounded morphology with low levels of cytoplasmic spreading (Fig. 2B), which was similar to the morphology observed when the monocytes were treated with EDTA before the adhesion assay (used as a negative control for cytoplasmic spreading, Fig. 2C). The spread area (μm2) of the monocyte cytoplasm, 72 [55–89] (median [lower and upper quartiles]), was larger for the monocytes cultured with medium alone than for the monocytes cultured with Leishmania (49 [43–57]; Mann-Whitney test, P < 0.0001, Fig. 2G).

Bottom Line: Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue.This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody.These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis.

View Article: PubMed Central - PubMed

Affiliation: Fundação Oswaldo Cruz-Bahia, Centro de Pesquisas Gonçalo Moniz, Brazilian Ministry of Health, Salvador, Brazil.

ABSTRACT
Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue. In this work, we studied the early stages of bond formation between VLA4 and fibronectin, measured the kinetics of membrane alignment and the monocyte cytoplasm spreading area over a fibronectin-coated surface, and studied the expression of high affinity integrin epitope in uninfected and Leishmania-infected human monocytes. Our results show that the initial VLA4-mediated interaction of Leishmania-infected monocyte with a fibronectin-coated surface is preserved, however, the later stage, leukocyte spreading over the substrate is abrogated in Leishmania-infected cells. The median of spreading area was 72 [55-89] μm(2) for uninfected and 41 [34-51] μm(2) for Leishmania-infected monocyte. This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody. After the initial contact with the fibronectrin-coated surface, uninfected monocyte quickly spread the cytoplasm at a 15 μm(2) s(-1) ratio whilst Leishmania-infected monocytes only made small contacts at a 5.5 μm(2) s(-1) ratio. The expression of high affinity epitope by VLA4 (from 39 ± 21% to 14 ± 3%); and LFA1 (from 37 ± 32% to 18 ± 16%) molecules was reduced in Leishmania-infected monocytes. These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis.

No MeSH data available.


Related in: MedlinePlus