IngentaConnect Release of calcium from intracellular stores and subsequent uptak...

Authors: Antonella Lupetti¹; Carlo P. J. M. Brouwer²; Heleen E. C. Dogterom-Ballering¹; Sonia Senesi³; Mario Campa³; Jaap T. van Dissel¹; Peter H. Nibbering¹

Source: Journal of Antimicrobial Chemotherapy, Volume 54, Number 3, 1 September 2004 , pp. 603-608(6)

Publisher: Oxford University Press

Abstract:

Objectives: Earlier studies showed that mitochondrial damage is a hallmark of the candidacidal activity of an N-terminal peptide of human lactoferrin, further referred to as hLF(1–11). Since uptake of Ca²⁺ by mitochondria may be essential for their activation, the aim of this study was to define the role of Ca²⁺ in killing of Candida albicans by the hLF(1–11) peptide.

Methods: The effect of compounds interfering with Ca²⁺ homeostasis on the hLF(1–11)-induced candidacidal activity, changes in mitochondrial membrane potential, and reactive oxygen species production were evaluated using a killing assay, rhodamine 123 staining, and 2 prime

-dichlorofluorescein diacetate, respectively. The increase in cellular Ca²⁺ content was measured using ⁴⁵Ca²⁺.

Results: Our results revealed that Ruthenium Red, which inhibits the mitochondrial Ca²⁺-uniporter and the voltage-sensitive Ca²⁺ release from internal stores, blocked (P<0.05) the hLF(1–11)-induced candidacidal activity as well as changes in the membrane potential of mitochondria, and reactive oxygen species production. Oxalate, which precipitates Ca²⁺ in intracellular organelles, decreased (P<0.05) the peptide-induced changes in the membrane potential of mitochondria, reactive oxygen species production, and candidacidal activity. Furthermore, the Ca²⁺ ionophore ionomycin combined with high CaCl<inf>2</inf> concentrations enhanced the hLF(1–11)-induced candidacidal activity. Moreover, hLF(1–11) caused an influx of Ca²⁺ from the extracellular medium into C. albicans reaching a three-fold increase at 2 h, whereas no increase was found in unexposed cells. In agreement, the Ca²⁺-chelator EGTA blocked the peptide-induced candidacidal activity.

Conclusions: Ca²⁺ release from intracellular stores, probably through subsequent mitochondrial Ca²⁺ uptake, is essential for the hLF(1–11)-induced candidacidal activity.

Keywords: lactoferrin peptide; mitochondrial Ca

Document Type: Research article

DOI: 10.1093/jac/dkh385

Affiliations: 1: Department of Infectious Diseases, C5-P, Leiden University Medical Center (LUMC), P.O. Box 9600, 2300 RC Leiden; 2: AM-Pharma, Rumpsterweg, 6, 3981 AK Bunnik, The Netherlands; 3: Dipartimento di Patologia Sperimentale, Biotecnologie Mediche, Infettivologia ed Epidemiologia, Università degli Studi di Pisa, Via S. Zeno, 35–39, 56127 Pisa, Italy

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Release of calcium from intracellular stores and subsequent uptake by mitochondria are essential for the candidacidal activity of an N-terminal peptide of human lactoferrin