Candida albicans BIOFILM ON ARTIFICIAL SURFACE IN CONTACT WITH INNATE IMMUNE CELLS
Arce Miranda JE, Baronetti J, Sotomayor CE*, Albesa I, Paraje MG #
Departamento de Farmacia, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba
*Departamento de Bioquímica Clínica, CIBICI, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba.
Haya de la Torre y Medina Allende, Córdoba (Argentina), CP: 5000.
Introduction
It has been documented that at least 65% of all microbial infections are related to formation of biofilms. The formation of these organizations by Candida primarily begin with adherence and colonization of a biotic host or an artificial surface as for example medical devices in current use, including stents, shunts, prostheses (voice box, heart valves, dentures, etc.), implants (lens, breast, etc.), and various types of catheters (1). We previously developed a model of Candida biofilms attached to polystyrene microtiter plates, where the oxidative metabolism under different culture conditions was assayed (2,3). In the present study, the effect of macrophages (Mø), a crucial innate immune cells, on the production of reactive oxygen species (ROS), the liberation of nitric oxide (NO), and the activity of superoxide dismutase (SOD) in these cultures was evaluated.
Materials and methods
Candida albicans was incubated in a 96-well polypropylene microtiter plate to form mature biofilms. Then, mouse Mø cell line (RAW) or medium alone, was added to the plate for 1h, centrifuged and supernatants were collected. The biofilms-forming ability was measured by determination of the adhesion to polystyrene microtiter plates (4). The Biofilms Biomass Unit (BBU) was arbitrarily defined with 0.1 OD595 equal to 1 BBU (5). In the supernatant, the extracellular ROS (eROS) production was detected by the reduction of nitro blue tetrazolium (NBT) to nitroblue diformazan, the NO production was evaluated as nitrite by a microplate assay method using the Griess reagent, and total SOD activity was assessed photochemically based on the inhibition of NBT reduction (6).
Results
The addition of Mø to the cultures results in a decrease in the biomass of mature biofilms with respect to control cultures (medium alone). Besides, we were not able to detect the production of eROS and NO by Candida biofilms subjected to Mø. On the other hand, the SOD activity of bifofilms co-cultured with Mø was higher than that found in control cultures.
Conclusions
Our results show that Mø are able to reduce the biomass of mature biofilms from a polymeric surface. Furthermore, the presence of these cells also induces an antioxidative profile in our model, indicated by the high SOD activity found in this system. In this way, this antioxidant power could be the reason why the production of eROS and NO was not detectable. Therefore, in this in vitro model, Mø can interact with biofilms already formed, diminishing their biomass and altering its oxidative balance, however, futures studies are necessary to explain the mechanism by which these cells induced the biofilms detachment in our system.
Acknowledgments
The authors wish to thank Bioq. Paula Icely. This work was supported by the following Grants: FONCyT, CONICET and SECyT.
Reference
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# Paraje MG. Tel +54 351 4334163, fax +54 351 4334127; e-mail:paraje@fcq.unc.edu.ar
