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NK cell activity differs between patients with localized and diffuse cutaneous leishmaniasis infected with Leishmania mexicana: a comparative study of TLRs and cytokines.

Cañeda-Guzmán IC, Salaiza-Suazo N, Fernández-Figueroa EA, Carrada-Figueroa G, Aguirre-García M, Becker I - PLoS ONE (2014)

Bottom Line: The altered protein expression found in NK cells of DCL patients correlated with their down-regulation of IFN-γ gene expression in LPG-stimulated and non-stimulated cells as compared to LCL patients.We conclude that in DCL patients the reduced NK-cell numbers and their diminished activity, evidenced by low TLR expression and low cytokine production, are possibly involved in the severity of the disease.Our results provide new information on the contribution of NK cells in Leishmania infections of the human host.

View Article: PubMed Central - PubMed

Affiliation: Unidad de Investigación en Medicina Experimental, Facultad de Medicina, Universidad Nacional Autónoma de México, Hospital General de México, México, D.F., México.

ABSTRACT
Leishmania mexicana causes localized (LCL) or diffuse cutaneous leishmaniasis (DCL). The cause of dissemination in DCL remains unknown, yet NK cells possibly play a role in activating leishmanicidal mechanisms during innate and adaptive immune responses. We had previously shown that Leishmania lipophosphoglycan (LPG) is a ligand for TLR2, activating human NK cells. We have now analyzed NK cells in LCL and DCL patients. NK numbers and effector mechanisms differed drastically between both groups of patients: DCL patients showed reduced NK cell numbers; diminished IFN-γ and TNF-α production; and lower TLR2, TLR1, and TLR6 expression as compared to LCL patients. The altered protein expression found in NK cells of DCL patients correlated with their down-regulation of IFN-γ gene expression in LPG-stimulated and non-stimulated cells as compared to LCL patients. NK cell response was further analyzed according to gender, age, and disease evolution in LCL patients showing that female patients produced higher IFN-γ levels throughout the disease progression, whereas TLR2 expression diminished in both genders with prolonged disease evolution and age. We furthermore show the activation pathway of LPG binding to TLR2 and demonstrated that TLR2 forms immunocomplexes with TLR1 and TLR6. In addition to the reduced NK cell numbers in peripheral blood, DCL patients also showed reduced NK cell numbers in the lesions. They were randomly scattered within the lesions, showing diminished cytokine production, which contrasts with those of LCL lesions, where NK cells produced IFN-γ and TNF-α and were found within organized granulomas. We conclude that in DCL patients the reduced NK-cell numbers and their diminished activity, evidenced by low TLR expression and low cytokine production, are possibly involved in the severity of the disease. Our results provide new information on the contribution of NK cells in Leishmania infections of the human host.

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TLR1, TLR2 and TLR6 expression on NK-cell subsets (CD56bright and CD56dim) of patients and controls.(A) Representative flow cytometry dot plot (TLR2 vs. CD56) and histogram of TLR2 expression in NK cells. (B) This gate was subsequently analyzed for TLR2 expression on NK-cell subsets. Dot plot (CD16+/CD56+) and histogram for CD56bright (blue box) and CD56dim (green box) NK cells. (C) TLR1 expression. (D) TLR2 expression. (E) TLR6 expression. □ Non-stimulated NK cells; ▪ LPG-stimulated NK cells. Lane 1: CD56dim; lane 2: CD56bright NK cells. Cell surface expression is indicated by MFI. Results are the mean ±SEM. *Significant differences were observed between NK cell subsets of LCL and DCL patients for all 3 TLRs in LPG-stimulated and non-stimulated cells. LCL patients (n = 9), DCL patients (n = 4) and controls (n = 4).
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pone-0112410-g007: TLR1, TLR2 and TLR6 expression on NK-cell subsets (CD56bright and CD56dim) of patients and controls.(A) Representative flow cytometry dot plot (TLR2 vs. CD56) and histogram of TLR2 expression in NK cells. (B) This gate was subsequently analyzed for TLR2 expression on NK-cell subsets. Dot plot (CD16+/CD56+) and histogram for CD56bright (blue box) and CD56dim (green box) NK cells. (C) TLR1 expression. (D) TLR2 expression. (E) TLR6 expression. □ Non-stimulated NK cells; ▪ LPG-stimulated NK cells. Lane 1: CD56dim; lane 2: CD56bright NK cells. Cell surface expression is indicated by MFI. Results are the mean ±SEM. *Significant differences were observed between NK cell subsets of LCL and DCL patients for all 3 TLRs in LPG-stimulated and non-stimulated cells. LCL patients (n = 9), DCL patients (n = 4) and controls (n = 4).

Mentions: Since NK cells are subdivided phenotypically according to their function into CD56dim (cytotoxic) and CD56bright (cytokine producing) cells, we were interested in analyzing TLR1, TLR2 and TLR6 expression in both NK subsets of LCL and DCL patients (Fig. 7A and 7B). We therefore analyzed the expression of these receptors in 9 LCL and 4 DCL patients, before and after stimulation with LPG. Only NK CD56bright cells expressed high levels of TLR1, TLR2 and TLR6, which were significantly higher in LCL patients (4 to 8-fold), as compared to DCL patients. There were also significant differences in TLRs expression between both NK subsets: CD56dim expressed significantly lower levels of TLRs as compared to CD56bright (Fig. 7C, 7D and 7E). However, no significant differences were found between non-stimulated and LPG-stimulated NK cells in both subsets of NK cells of the different groups.


NK cell activity differs between patients with localized and diffuse cutaneous leishmaniasis infected with Leishmania mexicana: a comparative study of TLRs and cytokines.

Cañeda-Guzmán IC, Salaiza-Suazo N, Fernández-Figueroa EA, Carrada-Figueroa G, Aguirre-García M, Becker I - PLoS ONE (2014)

TLR1, TLR2 and TLR6 expression on NK-cell subsets (CD56bright and CD56dim) of patients and controls.(A) Representative flow cytometry dot plot (TLR2 vs. CD56) and histogram of TLR2 expression in NK cells. (B) This gate was subsequently analyzed for TLR2 expression on NK-cell subsets. Dot plot (CD16+/CD56+) and histogram for CD56bright (blue box) and CD56dim (green box) NK cells. (C) TLR1 expression. (D) TLR2 expression. (E) TLR6 expression. □ Non-stimulated NK cells; ▪ LPG-stimulated NK cells. Lane 1: CD56dim; lane 2: CD56bright NK cells. Cell surface expression is indicated by MFI. Results are the mean ±SEM. *Significant differences were observed between NK cell subsets of LCL and DCL patients for all 3 TLRs in LPG-stimulated and non-stimulated cells. LCL patients (n = 9), DCL patients (n = 4) and controls (n = 4).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4232367&req=5

pone-0112410-g007: TLR1, TLR2 and TLR6 expression on NK-cell subsets (CD56bright and CD56dim) of patients and controls.(A) Representative flow cytometry dot plot (TLR2 vs. CD56) and histogram of TLR2 expression in NK cells. (B) This gate was subsequently analyzed for TLR2 expression on NK-cell subsets. Dot plot (CD16+/CD56+) and histogram for CD56bright (blue box) and CD56dim (green box) NK cells. (C) TLR1 expression. (D) TLR2 expression. (E) TLR6 expression. □ Non-stimulated NK cells; ▪ LPG-stimulated NK cells. Lane 1: CD56dim; lane 2: CD56bright NK cells. Cell surface expression is indicated by MFI. Results are the mean ±SEM. *Significant differences were observed between NK cell subsets of LCL and DCL patients for all 3 TLRs in LPG-stimulated and non-stimulated cells. LCL patients (n = 9), DCL patients (n = 4) and controls (n = 4).
Mentions: Since NK cells are subdivided phenotypically according to their function into CD56dim (cytotoxic) and CD56bright (cytokine producing) cells, we were interested in analyzing TLR1, TLR2 and TLR6 expression in both NK subsets of LCL and DCL patients (Fig. 7A and 7B). We therefore analyzed the expression of these receptors in 9 LCL and 4 DCL patients, before and after stimulation with LPG. Only NK CD56bright cells expressed high levels of TLR1, TLR2 and TLR6, which were significantly higher in LCL patients (4 to 8-fold), as compared to DCL patients. There were also significant differences in TLRs expression between both NK subsets: CD56dim expressed significantly lower levels of TLRs as compared to CD56bright (Fig. 7C, 7D and 7E). However, no significant differences were found between non-stimulated and LPG-stimulated NK cells in both subsets of NK cells of the different groups.

Bottom Line: The altered protein expression found in NK cells of DCL patients correlated with their down-regulation of IFN-γ gene expression in LPG-stimulated and non-stimulated cells as compared to LCL patients.We conclude that in DCL patients the reduced NK-cell numbers and their diminished activity, evidenced by low TLR expression and low cytokine production, are possibly involved in the severity of the disease.Our results provide new information on the contribution of NK cells in Leishmania infections of the human host.

View Article: PubMed Central - PubMed

Affiliation: Unidad de Investigación en Medicina Experimental, Facultad de Medicina, Universidad Nacional Autónoma de México, Hospital General de México, México, D.F., México.

ABSTRACT
Leishmania mexicana causes localized (LCL) or diffuse cutaneous leishmaniasis (DCL). The cause of dissemination in DCL remains unknown, yet NK cells possibly play a role in activating leishmanicidal mechanisms during innate and adaptive immune responses. We had previously shown that Leishmania lipophosphoglycan (LPG) is a ligand for TLR2, activating human NK cells. We have now analyzed NK cells in LCL and DCL patients. NK numbers and effector mechanisms differed drastically between both groups of patients: DCL patients showed reduced NK cell numbers; diminished IFN-γ and TNF-α production; and lower TLR2, TLR1, and TLR6 expression as compared to LCL patients. The altered protein expression found in NK cells of DCL patients correlated with their down-regulation of IFN-γ gene expression in LPG-stimulated and non-stimulated cells as compared to LCL patients. NK cell response was further analyzed according to gender, age, and disease evolution in LCL patients showing that female patients produced higher IFN-γ levels throughout the disease progression, whereas TLR2 expression diminished in both genders with prolonged disease evolution and age. We furthermore show the activation pathway of LPG binding to TLR2 and demonstrated that TLR2 forms immunocomplexes with TLR1 and TLR6. In addition to the reduced NK cell numbers in peripheral blood, DCL patients also showed reduced NK cell numbers in the lesions. They were randomly scattered within the lesions, showing diminished cytokine production, which contrasts with those of LCL lesions, where NK cells produced IFN-γ and TNF-α and were found within organized granulomas. We conclude that in DCL patients the reduced NK-cell numbers and their diminished activity, evidenced by low TLR expression and low cytokine production, are possibly involved in the severity of the disease. Our results provide new information on the contribution of NK cells in Leishmania infections of the human host.

Show MeSH
Related in: MedlinePlus