Antimicrobial Activity of Calprotectin Isolated from Human Empyema Fluid Supernatants

doi:10.1006/clin.1995.1127

Clinical Immunology and Immunopathology

Volume 76, Issue 3, September 1995, Pages 285-290

Clinical Immunology ...

https://doi.org/10.1006/clin.1995.1127 Get rights and content

Abstract

Abscess and empyema fluid supernatants have zinc-reversible antimicrobial activity that is thought to be due to calprotectin, a calcium- and zinc-binding protein present within neutrophil cytoplasm. The present studies were undertaken to determine if calprotectin isolated from human empyema fluid supernatants demonstrated similar antimicrobial activity to that of the original specimens. The characteristics of the calprotectin complex on SDS-PAGE and Western blotting with specific antisera were similar in neutrophil lysates and in empyema fluid supernatants. Ion-exchange and size-exclusion chromatography were used to obtain highly purified preparations of calprotectin from empyema fluids, and these preparations demonstrated zinc-reversible anti-Candida albicans activity which was similar to that observed in the original specimens. These findings suggest that calprotectin is responsible for most of the growth-inhibitory activity of empyema fluid supernatants against this organism.

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Cited by (24)

Fungal susceptibility to zinc deprivation
2004, Journal of Laboratory and Clinical Medicine
Citation Excerpt :
In the study reported here, 6 additional pathogenic fungal organisms were tested to determine whether this kind of susceptibility to zinc deprivation might be a more general phenomenon for fungi. As discussed above, abscess fluids have broad-spectrum microbistatic activity because of their content of a neutrophil-derived protein called calprotectin5,8,12; this protein appears to compete with the organisms for the zinc that they need to grow.14 In this study, as in the last one, zinc was the most potent of the metals tested in reversing growth inhibition by TPEN, EDTA, DTPA and abscess fluid for all 7 organisms tested, even though in some cases the stability constants of certain of these chelators were higher for other metals than for zinc (most notably copper and iron).
Calprotectin is a neutrophil-derived antimicrobial protein that competes with microorganisms for zinc. The zinc-specific effect of calprotectin against Candida albicans appears to be related to this organism's marked susceptibility to deprivation of this metal. However, it is not known whether this susceptibility is particular to C albicans or whether it is a characteristic of pathogenic fungi in general. As a means of deciding between these 2 possibilities, we undertook the study reported here to compare the susceptibility to zinc deprivation of 6 other pathogenic fungal species in addition to that of C albicans. We tested the effect of metals in reversing growth inhibition of the 7 fungi against abscess-fluid supernatant (a source of calprotectin) and 3 chemical chelators. Data were expressed as the concentration of metal required to bring about 50% restoration of growth. Zinc was found to be much more potent than the other metals tested in reversing growth inhibition of all the organisms by human abscess fluid and all 3 chemical chelators. Copper and manganese also had some effect. In some cases, chelator stability constants were higher for other metals than for zinc; in particular, although diethylenetriaminopentaacetic acid has a stability constant for iron almost 10¹⁰ times greater than that for zinc, zinc was more effective than iron in reversing growth inhibition by this chelator against all of the organisms. These results suggest that marked susceptibility to zinc deprivation is a general characteristic of pathogenic fungi.
Zinc, Iron, Copper, Selenium, Lactoferrin, and Ferritin in Human Pus
2004, American Journal of the Medical Sciences
Citation Excerpt :
These studies were done with a single clinical isolate of E coli and a single pus specimen and were presented to illustrate qualitative effects of cation replenishment and pH modification. Reduction of the bioavailability of zinc and iron are well known mechanisms of host defense against infection.5–12,17 Concentrations of free zinc or iron in pus should be low because calprotectin and lactoferrin are present in high concentrations in neutrophils, and neutrophils are concentrated in pus.5–9
Restriction of zinc and iron available for microbial growth in tissues are well-recognized host defense mechanisms. The present studies were performed to characterize some constituents of human pus that may affect these important host defenses.
Zinc, iron, copper, calcium, and magnesium in pus were measured using an atomic absorption spectrophotometer; selenium was measured fluorometrically. Ferritin was measured with a fluorometric enzyme immunoassay, and lactoferrin was measured with a radial diffusion assay. The growth of Escherichia coli at 37°C was measured in pus supernate adjusted to pH 5.5 or 7.4, in boiled supernate, or in supernate adjusted with 1.3 mM iron or 0.9 mM zinc singly or together.
Zinc and iron concentrations in pus exceeded normal serum. Calcium and magnesium levels were 2- to 3-fold lower and higher, respectively, than normal serum values. Lactoferrin concentrations of were 880 ± 48 μg/mL and ferritin levels were 20,726 ± 2,667 ng/mL. Growth of an E coli strain was inhibited in pus at pH 5.5 but not at pH 7.4, and growth was enhanced by addition of iron or zinc to E coli suspended in pus at pH 6.7.
To our knowledge, this is the first report of the zinc, iron, copper, selenium, lactoferrin, and ferritin levels of human pus. These studies provide additional insight into host defense mechanisms mediated by the restriction of the bioavailability of zinc and iron in suppurative infection.
A comparison of salivary calprotectin levels in subjects with and without oral candidiasis
2001, Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology, and Endodontics
Objectives: The purpose of this study was to compare salivary calprotectin levels in subjects with oral candidiasis to the levels in healthy age- and sex-matched control subjects and to investigate the relationship of salivary calprotectin to intensity of oral candidal carriage and infection and to predisposing factors for candidiasis. Study Design: Using enzyme-linked immunosorbent assay, we measured calprotectin in unstimulated whole saliva collected from 22 subjects with oral candidiasis and in saliva collected from 22 control subjects. Calprotectin levels in the 2 groups were compared by using the Wilcoxon signed rank test. The relationships between calprotectin levels and Candida counts were assessed by using the Spearman rank correlation test. The nonparametric 1-way analysis of variance test was used to study the relationship between calprotectin levels and presence of non-albicans Candida and individual predisposing factors for candidiasis. The Kruskal-Wallis test was used to evaluate the relationship between calprotectin and predisposing factors in subjects grouped by total number of predisposing factors identified. Results: No relationship was found between calprotectin and individual or total number of predisposing factors. Higher calprotectin levels were found in subjects with candidiasis, and calprotectin concentration positively correlated with intensity of candidal infection. Conclusions: These findings suggest that calprotectin production, or release, or both, may be increased in subjects with candidiasis. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2001;92:62-7)
Serial analysis of gene expression in differentiated cultures of human epidermal keratinocytes
2001, Journal of Investigative Dermatology
Citation Excerpt :
SKALP/elafin is a member of the recently described trappin gene family (Schalkwijk et al. 1999) and is involved in the protection against microbial infection (Simpson et al. 1999). MRPs are calcium-binding proteins, and it has been demonstrated that MRP-8 and MRP-14 can form a heterodimer and have anti-microbial activity in vitro (Brandtzaeg et al. 1995;Santhanagopalan et al. 1995;Loomans et al. 1998). Both proteins are significantly upregulated after TNF-α stimulation.
Keratinocyte gene expression was surveyed more comprehensively than before, by means of serial analysis of gene expression. A total of 25,694 tags derived from expressed mRNA, were analyzed in a model for normal differentiation and in a model where cultured keratinocytes were stimulated for a prolonged period of time with tumor necrosis factor-α, thus mimicking aberrant differentiation in the context of cutaneous inflammation. Serial analysis of gene expression revealed many transcripts derived from unknown genes and a large number of genes that are not known to be expressed in keratinocytes; furthermore, these data provide quantitative information about the relative abundance of transcripts, allowing the identification of differentially expressed genes. A major part of the identified transcripts accounted for genes involved in energy metabolism and protein synthesis. A large proportion of all transcripts (6%) corresponded to genes associated with terminal differentiation and barrier formation. Another highly expressed functional group of genes (2% of all transcripts) corresponded to proteins involved in host protection such as antimicrobial proteins and proteinase inhibitors. Three of these genes were not known to be expressed in keratinocytes, and some were upregulated after prolonged tumor necrosis factor-α exposure. Our data on expressed genes in keratinocytes are consistent with the known function of human epidermis, and provide a first step to generate a transcriptome of human keratinocytes.
Effect of metals on Candida albicans growth in the presence of chemical chelators and human abscess fluid
2001, Journal of Laboratory and Clinical Medicine
Citation Excerpt :
In the present study, growth inhibition by chemical chelators was compared with that caused by abscess fluid to evaluate the patterns of metal reversibility obtained. Abscess fluids have broad-spectrum microbistatic activity because they contain a neutrophil-derived protein called calprotectin.2,5,9 Calprotectin is a calcium- and zinc-binding protein of the S100 class with microbistatic activity that is reversible by the addition of zinc but not by the addition of calcium or other metals.2
Calprotectin is a calcium- and zinc-binding protein that is present in abscess fluid supernatants and appears to inhibit microbial growth through competition for zinc. In the present study, growth inhibition by chemical chelators was compared with that produced by human abscess fluid to determine whether other chelators, perhaps with different metal specificities, would have the same or different patterns of metal reversibility as abscess fluid. Zinc was found to be more potent than the other metals tested in reversing C. albicans growth inhibition by human abscess fluid and three chemical chelators, even though in some cases the stability constants of certain of these chelators were higher for other metals. For example, in the presence of the chelator diethylenetriaminopentaacetic acid, zinc stimulated Candida growth at a 10-fold lower concentration than did iron, even though this chelator has a stability constant for iron that is almost 10¹⁰ higher than that for zinc. These results suggest that the zinc specificity of calprotectin's C. albicans growth inhibition can best be explained by the marked sensitivity of this organism to zinc deprivation rather than by selective binding of this metal by the protein. (J Lab Clin Med 2001;137:284-9)
Antimicrobial and antibiofilm effects of human amniotic/chorionic membrane extract on Streptococcus pneumoniae
2017, Frontiers in Microbiology

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Regular ArticleAntimicrobial Activity of Calprotectin Isolated from Human Empyema Fluid Supernatants

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Regular Article
Antimicrobial Activity of Calprotectin Isolated from Human Empyema Fluid Supernatants