In Vitro Effects of Candida albicans and Aspergillus fumigatus on Dendritic Cells and the Role of Beta Glucan in this Effect

Although endometriosis is a benign disease characterized by the presence of endometrial tissues outside the uterus, ectopic endometrial cells can exhibit malignant biological behaviors. Retinol-binding protein4 (RBP4) is a novel adipocyte-derived cytokine, which has important roles in regulating insulin sensitivity and energy metabolism. RBP4 is a potent modulator of gene transcription, and acts by directly controlling cell growth, invasiveness, proliferation and differentiation. Here, we evaluated the possible role of RBP4 in the pathogenesis of endometriosis. We compared the levels of RBP4 in the tissues and peritoneal fluid (PF) of women with and without endometriosis and evaluated the in vitro effects of RBP4 on the viability, invasiveness, and proliferation of endometrial stromal cells (ESCs). RBP4 levels were significantly higher in the PF of the women in the endometriosis group than in the controls. RBP4 immunoreactivity was significantly higher in the ovarian endometriomas of women with advanced stage endometriosis than those of controls. In vitro treatment with human recombinant-RBP4 significantly increased the viability, bromodeoxyuridine expression, and invasiveness of ESCs. Transfection with RBP4 siRNA significantly reduced ESC viability and invasiveness. These findings suggest that RBP4 partakes in the pathogenesis of endometriosis by increasing the viability, proliferation and invasion of endometrial cells.

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Protein Synthesis in Human Leukocytes and Lymphocytes: 1. Effect of Steroids and Sterols

Blood ◽

10.1182/blood.v34.3.348.348 ◽

1969 ◽

Vol 34 (3) ◽

pp. 348-356 ◽

Cited By ~ 18

Author(s):

SEYMOUR WERTHAMER ◽

CARL HICKS ◽

LEONARD AMARAL

Keyword(s):

Protein Synthesis ◽

Steroid Hormones ◽

Plasma Protein ◽

Human Lymphocytes ◽

Amino Acid Incorporation ◽

Binding Factor ◽

In Vitro Effects ◽

Physiologic Role

Abstract The in vitro effects of sterols, cholesterol and 3-methyl cholanthrene and steroids, cortisol, prednisolone and testosterone on protein synthesis in separate popultions of human lymphocytes and leukocytes has been investigated. It has been shown that all agents used result in the inhibition of protein synthesis under these conditions. It has also been shown that the inhibitory mechanism of the steroid hormones requires the presence of plasma, presumably as a protein binding factor in order to achieve its effect. The sterol, cholesterol and 3-methyl cholanthrene, in the absence of plasma, still inhibit amino acid incorporation. However, in the case of cholesterol, the magnitude of inhibition is lower than that observed in the presence of plasma, perhaps indicating a partial plasma dependence. The results presented therefore support the hypothesis that the inhibition of lymphocyte protein synthesis by steroid hormones occurs only when the steroid is bound to a plasma protein. The physiologic role of the plasma protein-cortisol complex and its relation to the condition of lymphopenia in man is discussed.

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In Vivo Role of Dendritic Cells in a Murine Model of Pulmonary Cryptococcosis

Infection and Immunity ◽

10.1128/iai.00317-06 ◽

2006 ◽

Vol 74 (7) ◽

pp. 3817-3824 ◽

Cited By ~ 58

Author(s):

Karen L. Wozniak ◽

Jatin M. Vyas ◽

Stuart M. Levitz

Keyword(s):

T Cells ◽

Dendritic Cells ◽

T Cell Activation ◽

Cell Activation ◽

Ex Vivo ◽

Interleukin 2 ◽

Mouse Lung

ABSTRACT Dendritic cells (DC) have been shown to phagocytose and kill Cryptococcus neoformans in vitro and are believed to be important for inducing protective immunity against this organism. Exposure to C. neoformans occurs mainly by inhalation, and in this study we examined the in vivo interactions of C. neoformans with DC in the lung. Fluorescently labeled live C. neoformans and heat-killed C. neoformans were administered intranasally to C57BL/6 mice. At specific times postinoculation, mice were sacrificed, and lungs were removed. Single-cell suspensions of lung cells were prepared, stained, and analyzed by microscopy and flow cytometry. Within 2 h postinoculation, fluorescently labeled C. neoformans had been internalized by DC, macrophages, and neutrophils in the mouse lung. Additionally, lung DC from mice infected for 7 days showed increased expression of the maturation markers CD80, CD86, and major histocompatibility complex class II. Finally, ex vivo incubation of lung DC from infected mice with Cryptococcus-specific T cells resulted in increased interleukin-2 production compared to the production by DC from naïve mice, suggesting that there was antigen-specific T-cell activation. This study demonstrated that DC in the lung are capable of phagocytosing Cryptococcus in vivo and presenting antigen to C. neoformans-specific T cells ex vivo, suggesting that these cells have roles in innate and adaptive pulmonary defenses against cryptococcosis.

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Systematic characterization and genetic interaction analysis of adhesins in Candida albicans virulence

10.1101/2020.10.22.350991 ◽

2020 ◽

Author(s):

Sierra Rosiana ◽

Liyang Zhang ◽

Grace H. Kim ◽

Alexey V. Revtovich ◽

Arjun Sukumaran ◽

...

Keyword(s):

Candida Albicans ◽

Genetic Interaction ◽

Interaction Analysis ◽

Genetic Interactions ◽

Fungal Virulence ◽

C Elegans ◽

Adhesin Proteins ◽

Insight Into

AbstractCandida albicans is a microbial fungus that exists as a commensal member of the human microbiome and an opportunistic pathogen. Cell surface-associated adhesin proteins play a crucial role in C. albicans’ ability to undergo cellular morphogenesis, develop robust biofilms, colonize, and cause infection in a host. However, a comprehensive analysis of the role and relationships between these adhesins has not been explored. We previously established a CRISPR-based platform for efficient generation of single- and double-gene deletions in C. albicans, which was used to construct a library of 144 mutants, comprising 12 unique adhesin genes deleted singly, or in every possible combination of double deletions. Here, we exploit this adhesin mutant library to explore the role of adhesin proteins in C. albicans virulence. We perform a comprehensive, high-throughput screen of this library, using Caenorhabditis elegans as a simplified model host system, which identified mutants critical for virulence and significant genetic interactions. We perform follow-up analysis to assess the ability of high- and low-virulence strains to undergo cellular morphogenesis and form biofilms in vitro, as well as to colonize the C. elegans host. We further perform genetic interaction analysis to identify novel significant negative genetic interactions between adhesin mutants, whereby combinatorial perturbation of these genes significantly impairs virulence, more than expected based on virulence of the single mutant constituent strains. Together, this yields important new insight into the role of adhesins, singly and in combinations, in mediating diverse facets of virulence of this critical fungal pathogen.SummaryCandida albicans is a human fungal pathogen and cause of life-threatening systemic infections. Cell surface-associated adhesins play a central role in this pathogen’s ability to establish infection. Here, we provide a comprehensive analysis of adhesin factors, and their role in fungal virulence. Exploiting a high-throughput workflow, we screened an adhesin mutant library using C. elegans as a simple model host, and identified mutants and genetic interactions involved in virulence. We found that adhesin mutants are impaired in in vitro pathogenicity, irrespective of their virulence. Together, this work provides new insight into the role of adhesin factors in mediating fungal virulence.

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Inhibition of O-GlcNAc Transferase Alters the Differentiation and Maturation Process of Human Monocyte Derived Dendritic Cells

Cells ◽

10.3390/cells10123312 ◽

2021 ◽

Vol 10 (12) ◽

pp. 3312

Author(s):

Matjaž Weiss ◽

Marko Anderluh ◽

Martina Gobec

Keyword(s):

Dendritic Cells ◽

Posttranslational Modification ◽

Human Monocyte ◽

T Cell Differentiation ◽

Maturation Process ◽

Hla Dr ◽

Glcnac Transferase ◽

And Function

The O-GlcNAcylation is a posttranslational modification of proteins regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase. These enzymes regulate the development, proliferation and function of cells, including the immune cells. Herein, we focused on the role of O-GlcNAcylation in human monocyte derived dendritic cells (moDCs). Our study suggests that inhibition of OGT modulates AKT and MEK/ERK pathways in moDCs. Changes were also observed in the expression levels of relevant surface markers, where reduced expression of CD80 and DC-SIGN, and increased expression of CD14, CD86 and HLA-DR occurred. We also noticed decreased IL-10 and increased IL-6 production, along with diminished endocytotic capacity of the cells, indicating that inhibition of O-GlcNAcylation hampers the transition of monocytes into immature DCs. Furthermore, the inhibition of OGT altered the maturation process of immature moDCs, since a CD14medDC-SIGNlowHLA-DRmedCD80lowCD86high profile was noticed when OGT inhibitor, OSMI-1, was present. To evaluate DCs ability to influence T cell differentiation and polarization, we co-cultured these cells. Surprisingly, the observed phenotypic changes of mature moDCs generated in the presence of OSMI-1 led to an increased proliferation of allogeneic T cells, while their polarization was not affected. Taken together, we confirm that shifting the O-GlcNAcylation status due to OGT inhibition alters the differentiation and function of moDCs in in vitro conditions.

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Insights in the molecular mechanisms of an azole stress adapted laboratory-generated Aspergillus fumigatus strain

Medical Mycology ◽

10.1093/mmy/myaa118 ◽

2021 ◽

Author(s):

Marion Aruanno ◽

Samantha Gozel ◽

Isabelle Mouyna ◽

Josie E Parker ◽

Daniel Bachmann ◽

...

Keyword(s):

Transcription Factor ◽

Aspergillus Fumigatus ◽

Molecular Mechanisms ◽

Drug Transporters ◽

Major Facilitator Superfamily ◽

Ergosterol Biosynthesis ◽

Azole Resistance ◽

Drug Transporter

Abstract Aspergillus fumigatus is the main cause of invasive aspergillosis, for which azole drugs are the first-line therapy. Emergence of pan-azole resistance among A. fumigatus is concerning and has been mainly attributed to mutations in the target gene (cyp51A). However, azole resistance may also result from other mutations (hmg1, hapE) or other adaptive mechanisms. We performed microevolution experiment exposing an A. fumigatus azole-susceptible strain (Ku80) to sub-minimal inhibitory concentration of voriconazole to analyze emergence of azole resistance. We obtained a strain with pan-azole resistance (Ku80R), which was partially reversible after drug relief, and without mutations in cyp51A, hmg1, and hapE. Transcriptomic analyses revealed overexpression of the transcription factor asg1, several ATP-binding cassette (ABC) and major facilitator superfamily transporters and genes of the ergosterol biosynthesis pathway in Ku80R. Sterol analysis showed a significant decrease of the ergosterol mass under voriconazole exposure in Ku80, but not in Ku80R. However, the proportion of the sterol compounds was similar between both strains. To further assess the role of transporters, we used the ABC transporter inhibitor milbemycine oxime (MLB). MLB inhibited transporter activity in both Ku80 and Ku80R and demonstrated some potentiating effect on azole activity. Criteria for synergism were reached for MLB and posaconazole against Ku80. Finally, deletion of asg1 revealed some role of this transcription factor in controlling drug transporter expression, but had no impact on azole susceptibility. This work provides further insight in mechanisms of azole stress adaptation and suggests that drug transporters inhibition may represent a novel therapeutic target. Lay Summary A pan-azole-resistant strain was generated in vitro, in which drug transporter overexpression was a major trait. Analyses suggested a role of the transporter inhibitor milbemycin oxime in inhibiting drug transporters and potentiating azole activity.

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