Endocytosis of<i>Candida albicans</i>by vascular endothelial cells is associated with tyrosine phosphorylation of specific host cell proteins

Belanger, Paul H.; Johnston, Douglas A.; Fratti, Rutilio A.; Zhang, Mason Z.; Filler, Scott G.

doi:10.1046/j.1462-5822.2002.00232.x

Cited by 32 publications

(26 citation statements)

References 40 publications

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“…This organism is similar to A fumigatus in that both live and killed hyphae are endocytosed by endothelial cells and this process can be inhibited by cytochalasin D. 14,16,19 However, a major difference between the 2 organisms is that A fumigatus conidia are endocytosed by endothelial cells more avidly than hyphae, whereas C albicans hyphae are endocytosed more avidly than blastoconidia. 15 There are also significant differences in endothelial cell injury caused by A fumigatus and C albicans.…”

Section: Discussionmentioning

confidence: 99%

“…[14][15][16][17] Briefly, endothelial cells grown on glass coverslips were infected with 6 ϫ 10 4 conidia or germ tubes of A fumigatus H237 in tissue culture medium for 45 minutes. In some experiments, the microfilament inhibitor cytochalasin D was added to the wells just before the organisms to achieve a final concentration of 70 nM.…”

Section: Endocytosis Assaymentioning

confidence: 99%

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Interactions of Aspergillus fumigatus with endothelial cells: internalization, injury, and stimulation of tissue factor activity

Lópes-Bezerra

Filler

2004

Blood

114

112

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Invasive aspergillosis causes significant mortality among patients with hematologic malignancies. This infection is characterized by vascular invasion and thrombosis. To study the pathogenesis of invasive aspergillosis, we investigated the interactions of Aspergillus fumigatus conidia and hyphae with endothelial cells in vitro. We found that both forms of the organism induced endothelial cell microfilament rearrangement and subsequent endocytosis. Conidia were endocytosed 2-fold more avidly than hyphae, and endocytosis was independent of fungal viability. Endocytosed conidia and hyphae caused progressive endothelial cell injury after 4 hours of infection. Live conidia induced more endothelial cell injury than did live hyphae. However, endothelial cell injury caused by conidia was dependent on fungal viability, whereas injury caused by hyphae was not, indicating that conidia and hyphae injure endothelial cells by different mechanisms. Neither live nor killed conidia increased tissue factor activity of endothelial cells. In contrast, both live and killed hyphae stimulated significant endothelial cell tissue factor activity, as well as the expression of tissue factor antigen on the endothelial cell surface. These results suggest that angioinvasion and thrombosis caused by A fumigatus hyphae in vivo may be due in part to endothelial cell invasion, induction of injury, and stimulation of tissue factor activity. ( IntroductionInvasive aspergillosis has become one of the leading causes of death among bone marrow transplantation and leukemia patients. Even with current therapy, the mortality rate of invasive aspergillosis is 80% to 90% in immunocompromised patients. 1 Aspergillus fumigatus is responsible for 90% of invasive Aspergillus infections. 1 This ubiquitous mould releases numerous conidia into the atmosphere, which are small enough (2 to 3 m in diameter) to reach the pulmonary alveoli after they are inhaled. 2 Once the conidia reach the alveoli, they swell and germinate, producing hyphae that invade the pulmonary parenchyma. These hyphae have a marked tropism for blood vessels. A key finding in invasive aspergillosis is angioinvasion, which subsequently leads to thrombosis and tissue infarction. [3][4][5] In immunocompromised hosts, invasion of the pulmonary vasculature can result in widespread hematogenous dissemination to organs such the brain, kidneys, heart, and eyes. 1,3 During angioinvasion, A fumigatus hyphae interact with vascular endothelium. We hypothesize that this interaction plays an important role in the pathogenesis of invasive aspergillosis. Once the hyphae have entered the bloodstream, they must adhere to and penetrate the endothelial cell lining of the blood vessels to invade the deep tissues of the target organs.In addition, the prominent thrombosis at sites of A fumigatus angioinvasion suggests that the organism stimulates endothelial cells to become prothrombotic. A major mechanism by which endothelial cells can promote intravascular thrombus formation is by expressing tissue factor, also k...

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Section: Discussionmentioning

confidence: 99%

Section: Endocytosis Assaymentioning

confidence: 99%

Interactions of Aspergillus fumigatus with endothelial cells: internalization, injury, and stimulation of tissue factor activity

Lópes-Bezerra

Filler

2004

Blood

114

112

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“…All of these strains are clinical isolates and have been shown previously to be endocytosed by and cause injury to vascular endothelial cells in vitro (9,11,13). The tpk2⌬/tpk2⌬ mutant and a TPK2-reconstituted strain (tpk2⌬/tpk2⌬::TPK2), which were constructed from strain CAI-4 (14), were kindly provided by Joachim Ernst (Heinrich-Heine-Universität, Dü sseldorf, Germany).…”

Section: Methodsmentioning

confidence: 99%

“…The endocytosis of C. albicans hyphae by endothelial cells is independent of fungal viability and requires functional endothelial cell microfilaments and microtubules (8,9). Moreover, the endocytic process is governed in part by the tyrosine phosphorylation of endothelial cell proteins (11). Importantly, the endothelial cell receptor(s) that are bound by C. albicans and that mediate the endocytosis of the organism have not been identified previously.…”

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confidence: 99%

N-cadherin Mediates Endocytosis of Candida albicans by Endothelial Cells

Phan

Fratti

Prasadarao

et al. 2005

Journal of Biological Chemistry

105

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Candida albicans is the most common cause of fungal bloodstream infections. To invade the deep tissues, blood-borne organisms must cross the endothelial cell lining of the vasculature. We have found previously that C. albicans hyphae, but not blastospores, invade endothelial cells in vitro by inducing their own endocytosis. Therefore, we set out to identify the endothelial cell receptor that mediates the endocytosis of C. albicans. We determined that endocytosis of C. albicans was not mediated by bridging molecules in the serum and that it was partially dependent on the presence of extracellular calcium. Using an affinity purification procedure, we discovered that endothelial cell N-cadherin bound to C. albicans hyphae but not blastospores. N-cadherin also co-localized with C. albicans hyphae that were being endocytosed by endothelial cells. Chinese hamster ovary (CHO) cells expressing human N-cadherin endocytosed significantly more C. albicans hyphae than did CHO cells expressing either human VE-cadherin or no human cadherins. The expression of N-cadherin by the CHO cells resulted in enhanced endocytosis of hyphae, but not blastospores, indicating the selectivity of the N-cadherin-mediated endocytosis. Down-regulation of endothelial cell N-cadherin expression with small interfering RNA significantly inhibited the endocytosis of C. albicans hyphae. Therefore, a novel function of N-cadherin is that it serves as an endothelial cell receptor, which mediates the endocytosis of C. albicans.

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“…albicans damages human vascular endothelial cells in vivo and in vitro (5,10,17,25). Maximal endothelial cell damage (ECD) occurs in vitro when C. albicans adheres to and invades the endothelial cells and then secretes lytic enzymes (2,10,11,13,16). Moreover, some C. albicans mutants with filamentation defects cause significantly less ECD than the wild-type parent strain (24).…”

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confidence: 99%

Relationship between Candida albicans Virulence during Experimental Hematogenously Disseminated Infection and Endothelial Cell Damage In Vitro

et al. 2004

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Candida albicans must penetrate the endothelial cell lining of the vasculature to invade the deep tissues during a hematogenously disseminated infection. We compared 27 C. albicans mutants with their wild-type parent for their capacity to damage endothelial cells in vitro and cause a lethal infection in mice following tail vein inoculation. Of 10 mutants with significantly impaired capacity to damage endothelial cells, all had attenuated virulence. Therefore, the endothelial cell damage assay can be used as a screen to identify some virulence factors relevant to hematogenously disseminated candidiasis.During the initiation of hematogenously disseminated candidiasis, blood-borne organisms must adhere to and penetrate the endothelial cell lining of the blood vessels to invade the deep tissues. One mechanism by which Candida albicans can penetrate the vascular endothelium is by damaging and eventually killing the endothelial cells. Damaged endothelial cells detach from the basement membrane, leaving gaps through which the organism can invade. Also, the exposed basement membrane can be avidly bound by additional organisms (18).C. albicans damages human vascular endothelial cells in vivo and in vitro (5,10,17,25). Maximal endothelial cell damage (ECD) occurs in vitro when C. albicans adheres to and invades the endothelial cells and then secretes lytic enzymes (2,10,11,13,16). Moreover, some C. albicans mutants with filamentation defects cause significantly less ECD than the wild-type parent strain (24). These filamentation mutants also have attenuated virulence in various experimental models of infection (reviewed in reference 23). We hypothesized that the in vitro assay for C. albicans-induced ECD (ECD assay) can serve as a model for certain aspects of host-pathogen interactions in vivo, such as the ability of the organism to adhere to, invade, and injure host cells. This hypothesis predicts that some mutants with virulence defects will be defective in the ECD assay. The goal of the present study was to investigate this prediction.C. albicans strains. The genotypes and sources of the C. albicans strains used here are listed in Tables 1 and 2. Each strain was grown overnight in yeast nitrogen base broth (Difco, Detroit, Mich.) supplemented with 2% glucose (wt/vol) at 20°C on a rotating drum. The blastospores were harvested by centrifugation, washed with phosphate-buffered saline, enumerated with a hemacytometer, and suspended in RPMI 1640 medium (Irvine Scientific, Santa Ana, Calif.).ECD assay. We used our standard 51 Cr release ECD assay to determine the abilities of mutants of C. albicans to damage endothelial cells in vitro (24). The ECD assay was performed in 96-well tissue culture plates (Corning Inc., Acton, Mass.) with endothelial cells isolated from human umbilical cord veins, as described previously (24). The inoculum was 4 ϫ 10 4 organisms per well, and the organisms were incubated with the endothelial cells for 3 h in 5% CO 2 at 37°C. At the end of the incubation period, the wells were examined with an inv...

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Endocytosis ofCandida albicansby vascular endothelial cells is associated with tyrosine phosphorylation of specific host cell proteins

Cited by 32 publications

References 40 publications

Interactions of Aspergillus fumigatus with endothelial cells: internalization, injury, and stimulation of tissue factor activity

Interactions of Aspergillus fumigatus with endothelial cells: internalization, injury, and stimulation of tissue factor activity

N-cadherin Mediates Endocytosis of Candida albicans by Endothelial Cells

Relationship between Candida albicans Virulence during Experimental Hematogenously Disseminated Infection and Endothelial Cell Damage In Vitro

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