Members of the Candida parapsilosis Complex and Candida albicans are Differentially Recognized by Human Peripheral Blood Mononuclear Cells

Abstract

The systemic infections caused by members of the Candida parapsilosis complex are currently associated to high morbility and mortality rates, and are considered as relevant as those caused by Candida albicans. Since the fungal cell wall is the first point of contact with the host cells, here we performed a comparison of this organelle in members of the C. parapsilosis complex, and its relevance during interaction with human peripheral blood mononuclear cells (PBMCs). We found that the wall of the C. parapsilosis complex members is similar in composition, but differs to that from C. albicans, with less mannan content and more β-glucan and porosity levels. Furthermore, lectin-based analysis showed increased chitin and β1,3-glucan exposure at the surface of C. parapsilosis sensu lato when compared to C. albicans. Yeast cells of members of the C. parapsilosis complex stimulated more cytokine production by human PBMCs than C. albicans cells; and this significantly changed upon removal of O-linked mannans, indicating this wall component plays a significant role in cytokine stimulation by C. parapsilosis sensu lato. When inner wall components were exposed on the wall surface, C. parapsilosis sensu stricto and C. metapsilosis, but not C. orthopsilosis, stimulated higher cytokine production. Moreover, we found a strong dependency on β1,3-glucan recognition for the members of the C. parapsilosis complex, but not for live C. albicans cells; whereas TLR4 was required for TNFα production by the three members of the complex, and stimulation of IL-6 by C. orthopsilosis. Mannose receptor had a significant role during TNFα and IL-1β stimulation by members of the complex. Finally, we demonstrated that purified N- and O-mannans from either C. parapsilosis sensu lato or C. albicans are capable to block the recognition of these pathogens by human PBMCs. Together; our results suggest that the innate immune recognition of the members of the C. parapsilosis complex is differential of that reported for C. albicans. In addition, we propose that purified cell wall mannans can be used as antagonist to block specific receptors on innate immune cells.

Keywords: Candida albicans host-fungus interaction; Candida parapsilosis; cell wall; cytokine; mononuclear cells.

FIGURE 1

Analysis of β1,3-glucan and chitin exposure at the cell wall surface. Live (L) yeast cells were incubated with IgG Fc-Dectin-1and then with FITC-conjugated IgG to label β1,3-glucan (closed circles). Alternatively, chitin was labeled with WGA-FITC (open circles). The fluorescence associated to each cell is plotted. In addition, the experiment was conducted with cells previously heat-inactivated (HK). Strains used are Candida parapsilosis sensu stricto SZMC 8110, C. orthopsilosis SZMC 1545, C. metapsilosis SZMC 1548, and C. albicans SC5314. One hundred cells were analyzed for each group. ‡P < 0.05, when compared with the live cells; †P < 0.05 when compared to the other groups with live cells.

FIGURE 2

Candida albicans and members of the C. parapsilosis complex differentially stimulate the cytokine production by human PBMCs. Human PBMCs were co-incubated 24 h with live yeast cells, the supernatant collected and used to quantify the cytokine levels. Strains used are C. albicans SC5314, C. parapsilosis sensu stricto SZMC 8110, C. orthopsilosis SZMC 1545, and C. metapsilosis SZMC 1548. ^∗P < 0.05, when compared with the cytokine level stimulated by other species.

FIGURE 3

Cytokine production by human PBMCs stimulated with either live, heat-killed (HK) or β-eliminated yeast cells. Human PBMCs were co-incubated 24 h with either live, HK, live and β-eliminated, or HK and β-eliminated yeast cells, supernatant collected and used to quantify the cytokine levels. Strains used are C. albicans SC5314, C. parapsilosis sensu stricto SZMC 8110, C. orthopsilosis SZMC 1545, and C. metapsilosis SZMC 1548. ^∗P < 0.05, when compared with live cells from the same species.

FIGURE 4

Cytokine production by human PBMCs pre-incubated with laminarin. Human PBMCs were pre-incubated 60 min with 200 μg/mL laminarin before challenged for 24 h with either live or HK yeast cells, supernatant were collected and used to quantify the cytokine levels. Strains used are C. albicans SC5314, C. parapsilosis sensu stricto SZMC 8110, C. orthopsilosis SZMC 1545, and C. metapsilosis SZMC 1548. ^∗P < 0.05, when compared with untreated cells from the same organism.

FIGURE 5

Cytokine production by human PBMCs pre-incubated with anti-TLR4 antibodies. Human PBMCs were pre-incubated 60 min with 10 μg/mL either anti-TLR4 antibodies or IgG₁ before challenged for 24 h with live cells, supernatant were collected and used to quantify the cytokine levels. Strains used are C. albicans SC5314, C. parapsilosis sensu stricto SZMC 8110, C. orthopsilosis SZMC 1545, and C. metapsilosis SZMC 1548. ^∗P < 0.05, when compared with untreated cells from the same organism. NT, non-treated cells.

FIGURE 6

Cytokine production by human PBMCs pre-incubated with anti-MR antibodies. Human PBMCs were pre-incubated 60 min with 10 μg/mL either anti-MR antibodies or IgG₁ before challenged for 24 h with live cells, supernatant were collected and used to quantify the cytokine levels. Strains used are C. albicans SC5314, C. parapsilosis sensu stricto SZMC 8110, C. orthopsilosis SZMC 1545, and C. metapsilosis SZMC 1548. ^∗P < 0.05, when compared with untreated cells from the same organism. NT, non-treated cells.