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Neutrophil paralysis in Plasmodium vivax malaria.

Leoratti FM, Trevelin SC, Cunha FQ, Rocha BC, Costa PA, Gravina HD, Tada MS, Pereira DB, Golenbock DT, Antonelli LR, Gazzinelli RT - PLoS Negl Trop Dis (2012)

Bottom Line: The activation of innate immune responses by Plasmodium vivax results in activation of effector cells and an excessive production of pro-inflammatory cytokines that may culminate in deleterious effects.Blood samples were collected from P. vivax-infected patients at admission (day 0) and 30-45 days after treatment with chloroquine and primaquine.While monocytes were found to be the main source of cytokines in response to TLR ligands, neutrophils showed enhanced phagocytic activity and superoxide production.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Imunopatologia, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brazil.

ABSTRACT

Background: The activation of innate immune responses by Plasmodium vivax results in activation of effector cells and an excessive production of pro-inflammatory cytokines that may culminate in deleterious effects. Here, we examined the activation and function of neutrophils during acute episodes of malaria.

Materials and methods: Blood samples were collected from P. vivax-infected patients at admission (day 0) and 30-45 days after treatment with chloroquine and primaquine. Expression of activation markers and cytokine levels produced by highly purified monocytes and neutrophils were measured by the Cytometric Bead Assay. Phagocytic activity, superoxide production, chemotaxis and the presence of G protein-coupled receptor (GRK2) were also evaluated in neutrophils from malaria patients.

Principal findings: Both monocytes and neutrophils from P. vivax-infected patients were highly activated. While monocytes were found to be the main source of cytokines in response to TLR ligands, neutrophils showed enhanced phagocytic activity and superoxide production. Interestingly, neutrophils from the malaria patients expressed high levels of GRK2, low levels of CXCR2, and displayed impaired chemotaxis towards IL-8 (CXCL8).

Conclusion: Activated neutrophils from malaria patients are a poor source of pro-inflammatory cytokines and display reduced chemotactic activity, suggesting a possible mechanism for an enhanced susceptibility to secondary bacterial infection during malaria.

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Related in: MedlinePlus

TLR agonists induce production of high IL-1β, IL-6 and TNF-α levels by monocytes from P. vivax-infected subjects.Purified monocytes (A) or neutrophils (B) from P. vivax-infected subjects before (closed circles; n = 13) and 30–45 days after treatment (open circles; n = 13) were cultured for 48 hours in the absence or presence of LPS or Pam. Levels of IL-1β, IL-6, IL-10, TNF-α, and IL-8 (CXCL8) were measured in supernatant of monocyte (A) and neutrophil (B) cultures. Levels of cytokines were measured employing the Cytometric Bead Array (CBA). Significant differences are indicated p-values using paired t test or Wilcoxon signed rank test when a normality test failed.
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pntd-0001710-g003: TLR agonists induce production of high IL-1β, IL-6 and TNF-α levels by monocytes from P. vivax-infected subjects.Purified monocytes (A) or neutrophils (B) from P. vivax-infected subjects before (closed circles; n = 13) and 30–45 days after treatment (open circles; n = 13) were cultured for 48 hours in the absence or presence of LPS or Pam. Levels of IL-1β, IL-6, IL-10, TNF-α, and IL-8 (CXCL8) were measured in supernatant of monocyte (A) and neutrophil (B) cultures. Levels of cytokines were measured employing the Cytometric Bead Array (CBA). Significant differences are indicated p-values using paired t test or Wilcoxon signed rank test when a normality test failed.

Mentions: The expression of TLR2 was lower on both monocytes (8 out of 11 subjects) and neutrophils (10 out of 11 subjects) from malaria patients, whereas expression of TLR4 was only affected on neutrophils (8 out of 12) from infected individuals (Figures 2A and 2B). After parasitological cure, the surface expression of TLR returned to levels comparable to cells from HD (monocytes: TLR2 (median: 256; IQR: 241–276) and TLR4 (median: 11; IQR: 9–17; neutrophils: TLR2 (median 46; IQR: 36–62) and TLR4 (median: 3.5; IQR: 2.9–6.4)). Two millions per ml (2×105/well) of highly purified monocytes or neutrophils were cultured with or without TLR agonists and cytokines measured in the tissue culture supernatants (Figure 3A and B). Cell preparations reached over 98% of purity, as shown in the representative dot plots presented in Figure 3. Except for IL-8, neutrophils were over all a poor source of cytokines. No detectable levels of IL-1β, IL-6, IL-10 and TNF-α were found in culture supernatants of neutrophils stimulated or not with LPS or Pam (Figure 3B), even when neutrophils were used at a concentration of 1×107/ml (1×106/well). On the other hand, upon stimulation with LPS or Pam, high levels of IL-1β, IL-6, and TNF-α were produced by monocytes from malaria patients. (Figure 3A). Interestingly, the levels of LPS-induced IL-10 were lower in culture supernatants of monocytes derived from patients with ongoing P. vivax infection. After parasitological cure the cytokine production by monocytes returned to levels comparable to those produced by monocytes from HD (median: 1692; IQR: 1200–5265). IL-8 (CXCL8) was the only cytokine produced in high levels by neutrophils. Despite the down-modulation of TLR2 and TLR4, stimulation with TLR agonists increased the levels of IL-8 produced by neutrophils (Figure 3B) from malaria patients before (medium versus LPS, p = 0.0010 or Pam, p = 0.0024) and after treatment (medium vs LPS, p = 0.0002; or Pam, p = 0.0017). IL-8 was also produced by monocytes and the levels of cytokine did not differ between patients, before and after treatment (Figure 3A).


Neutrophil paralysis in Plasmodium vivax malaria.

Leoratti FM, Trevelin SC, Cunha FQ, Rocha BC, Costa PA, Gravina HD, Tada MS, Pereira DB, Golenbock DT, Antonelli LR, Gazzinelli RT - PLoS Negl Trop Dis (2012)

TLR agonists induce production of high IL-1β, IL-6 and TNF-α levels by monocytes from P. vivax-infected subjects.Purified monocytes (A) or neutrophils (B) from P. vivax-infected subjects before (closed circles; n = 13) and 30–45 days after treatment (open circles; n = 13) were cultured for 48 hours in the absence or presence of LPS or Pam. Levels of IL-1β, IL-6, IL-10, TNF-α, and IL-8 (CXCL8) were measured in supernatant of monocyte (A) and neutrophil (B) cultures. Levels of cytokines were measured employing the Cytometric Bead Array (CBA). Significant differences are indicated p-values using paired t test or Wilcoxon signed rank test when a normality test failed.
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Related In: Results  -  Collection

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pntd-0001710-g003: TLR agonists induce production of high IL-1β, IL-6 and TNF-α levels by monocytes from P. vivax-infected subjects.Purified monocytes (A) or neutrophils (B) from P. vivax-infected subjects before (closed circles; n = 13) and 30–45 days after treatment (open circles; n = 13) were cultured for 48 hours in the absence or presence of LPS or Pam. Levels of IL-1β, IL-6, IL-10, TNF-α, and IL-8 (CXCL8) were measured in supernatant of monocyte (A) and neutrophil (B) cultures. Levels of cytokines were measured employing the Cytometric Bead Array (CBA). Significant differences are indicated p-values using paired t test or Wilcoxon signed rank test when a normality test failed.
Mentions: The expression of TLR2 was lower on both monocytes (8 out of 11 subjects) and neutrophils (10 out of 11 subjects) from malaria patients, whereas expression of TLR4 was only affected on neutrophils (8 out of 12) from infected individuals (Figures 2A and 2B). After parasitological cure, the surface expression of TLR returned to levels comparable to cells from HD (monocytes: TLR2 (median: 256; IQR: 241–276) and TLR4 (median: 11; IQR: 9–17; neutrophils: TLR2 (median 46; IQR: 36–62) and TLR4 (median: 3.5; IQR: 2.9–6.4)). Two millions per ml (2×105/well) of highly purified monocytes or neutrophils were cultured with or without TLR agonists and cytokines measured in the tissue culture supernatants (Figure 3A and B). Cell preparations reached over 98% of purity, as shown in the representative dot plots presented in Figure 3. Except for IL-8, neutrophils were over all a poor source of cytokines. No detectable levels of IL-1β, IL-6, IL-10 and TNF-α were found in culture supernatants of neutrophils stimulated or not with LPS or Pam (Figure 3B), even when neutrophils were used at a concentration of 1×107/ml (1×106/well). On the other hand, upon stimulation with LPS or Pam, high levels of IL-1β, IL-6, and TNF-α were produced by monocytes from malaria patients. (Figure 3A). Interestingly, the levels of LPS-induced IL-10 were lower in culture supernatants of monocytes derived from patients with ongoing P. vivax infection. After parasitological cure the cytokine production by monocytes returned to levels comparable to those produced by monocytes from HD (median: 1692; IQR: 1200–5265). IL-8 (CXCL8) was the only cytokine produced in high levels by neutrophils. Despite the down-modulation of TLR2 and TLR4, stimulation with TLR agonists increased the levels of IL-8 produced by neutrophils (Figure 3B) from malaria patients before (medium versus LPS, p = 0.0010 or Pam, p = 0.0024) and after treatment (medium vs LPS, p = 0.0002; or Pam, p = 0.0017). IL-8 was also produced by monocytes and the levels of cytokine did not differ between patients, before and after treatment (Figure 3A).

Bottom Line: The activation of innate immune responses by Plasmodium vivax results in activation of effector cells and an excessive production of pro-inflammatory cytokines that may culminate in deleterious effects.Blood samples were collected from P. vivax-infected patients at admission (day 0) and 30-45 days after treatment with chloroquine and primaquine.While monocytes were found to be the main source of cytokines in response to TLR ligands, neutrophils showed enhanced phagocytic activity and superoxide production.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Imunopatologia, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brazil.

ABSTRACT

Background: The activation of innate immune responses by Plasmodium vivax results in activation of effector cells and an excessive production of pro-inflammatory cytokines that may culminate in deleterious effects. Here, we examined the activation and function of neutrophils during acute episodes of malaria.

Materials and methods: Blood samples were collected from P. vivax-infected patients at admission (day 0) and 30-45 days after treatment with chloroquine and primaquine. Expression of activation markers and cytokine levels produced by highly purified monocytes and neutrophils were measured by the Cytometric Bead Assay. Phagocytic activity, superoxide production, chemotaxis and the presence of G protein-coupled receptor (GRK2) were also evaluated in neutrophils from malaria patients.

Principal findings: Both monocytes and neutrophils from P. vivax-infected patients were highly activated. While monocytes were found to be the main source of cytokines in response to TLR ligands, neutrophils showed enhanced phagocytic activity and superoxide production. Interestingly, neutrophils from the malaria patients expressed high levels of GRK2, low levels of CXCR2, and displayed impaired chemotaxis towards IL-8 (CXCL8).

Conclusion: Activated neutrophils from malaria patients are a poor source of pro-inflammatory cytokines and display reduced chemotactic activity, suggesting a possible mechanism for an enhanced susceptibility to secondary bacterial infection during malaria.

Show MeSH
Related in: MedlinePlus