Effect of matrix metalloprotease inhibitors on the 95 kDa metallopeptidase of Candida albicans.

A 95 kDa metallopeptidase of Candida albicans could be involved in the process of dissemination of the yeast. Matrix metalloproteases (MMPs) are also responsible for collagen breakdown in inflammatory and malignant processes. We tested six compounds on the C. albicans enzyme. Doxycycline, gentamicin, cefalothin, galardin, and elaidic and oleic acids are known for their capacity to inhibit some MMPs. Amongst these agents, only oleic acid was able to markedly inhibit the purified metallopeptidase at very low concentrations. Moreover, this fatty acid inhibited the secretion of the enzyme in the culture medium without altering the yeast viability.

Influence of sub-inhibitory concentrations of conventional antifungals on metabolism of Candida albicans and on its adherence to polystyrene and extracellular matrix proteins.

Five antifungal agents with different mechanisms of action were compared for their ability to affect mitochondrial dehydrogenase activity and adherence capacity of Candida albicans to polystyrene and extracellular matrix proteins. Only amphotericin B inhibited mitochondrial dehydrogenase activity when the culture medium was supplemented with galactose. 5-Fluorocytosine and terbinafine did not affect this activity, whereas itraconazole and fluconazole improved it. Furthermore, in these experimental conditions, the effect of sub-inhibitory concentrations of antifungals on adherence was dependent on the tested antifungal and the adherence surface: amphotericin B inhibited adherence to polystyrene and fibrinogen, but improved adherence to extracellular matrix. For all surfaces tested, when culture medium was supplemented with galactose, fluorocytosine did not affect adherence, and itraconazole, fluconazole and terbinafine inhibited adherence. Our results also confirmed the influence of the carbohydrates: sub-minimum inhibitory concentrations (MIC) of itraconazole increased or did not modify the mitochondrial metabolism of yeasts when the culture medium was supplemented with galactose, but this antifungal always decreased mitochondrial metabolism when the culture medium was supplemented with glucose. These data indicate that antifungals used below their MIC values can have various effects. It is important to distinguish the effects of antifungals on the metabolism of C. albicans from effects on its adherence capacity. The former effects are linked to the viability of the yeast and the latter depends on the colonization of cellular as opposed to inert surfaces.

A Candida albicans metallopeptidase degrades constitutive proteins of extracellular matrix.

Among potential virulence factors of Candida albicans, enzymes seem to play an important role. Many studies concern the secreted aspartic proteinases (saps), and the degradation of some components of the subendothelial extracellular matrix by the isoenzyme sap2 has been proved. Nevertheless, other proteolytic enzymes could be involved in the pathogenicity of the yeast. We studied the degradation of four constitutive proteins of the extracellular matrix: type I and IV collagens, laminin and fibronectin, by a 95-kDa metallopeptidase, localised in the cell wall of C. albicans. Each of these constituents was incubated with the purified enzyme and its degradation products analysed by an electrophoretic method. We observed that type I collagen and fibronectin were totally degraded by the enzyme whereas type IV collagen and laminin were only partially degraded. The C. albicans metallopeptidase may play a role in the degradation of the subendothelial extracellular matrix components. This enzyme could facilitate the migration of the yeast in the tissues after crossing the endothelial layer, allowing the fungal invasion of target organs.

Improved immunodiagnosis of human candidiasis by an enzyme-linked immunosorbent assay using a Candida albicans 52-kilodalton metallopeptidase.

An immunogenic aminopeptidase of Candida albicans was purified by high-performance liquid chromatography. It was then used for the development of an enzyme-linked immunosorbent assay to detect antibodies directed against this antigen in sera from patients with candidiasis. This enzyme specifically cleaves the L-Arg-7-amino-4-methyl-coumarin substrate at pH 7.4 and was detected in the crude extract of different C. albicans isolates. Sera used for this study were obtained from healthy blood donors or from patients with one of the following: systemic candidiasis, aspergillosis, cryptococcosis, toxoplasmosis, or malaria. The statistical analysis demonstrates significant differences between absorbency values obtained with sera from patients with candidiasis and with sera from the other groups (P = 0.000001). Diagnostic parameters show high diagnostic specificity of 97% and a sensitivity of 83% at a cutoff value of 0.425 and suggest the usefulness of this aminopeptidase for the diagnosis of systemic candidiasis.