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JoVE Journal
Immunology and Infection
Einrichtung einer In-vitro- System zur intrazelluläre Verhalten der Studie Candida g...
Einrichtung einer In-vitro- System zur intrazelluläre Verhalten der Studie Candida g...
JoVE Journal
Immunology and Infection
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JoVE Journal Immunology and Infection
Establishment of an In vitro System to Study Intracellular Behavior of Candida glabrata in Human THP-1 Macrophages

Einrichtung einer In-vitro- System zur intrazelluläre Verhalten der Studie Candida glabrata Menschen THP-1-Makrophagen

Full Text
12,258 Views
13:47 min
December 10, 2013

DOI: 10.3791/50625-v

Maruti Nandan Rai1, Sapan Borah1, Gaurav Bairwa*1, Sriram Balusu*1,2, Neelima Gorityala1, Rupinder Kaur1

1Laboratory of Fungal Pathogenesis,Centre for DNA Fingerprinting and Diagnostics, Andhra Pradesh, India, 2Current location: VIB Department for Molecular Biomedical Research, UGent,Fiers-Schell-Van Montagu Building, Technologiepark 927, B-9052 Ghent (Zwijnaarde), Belgium

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Overview

This article presents a protocol for establishing an in vitro cell culture model to investigate the interaction between the human fungal pathogen Candida glabrata and human macrophages. This model aims to enhance understanding of fungal virulence mechanisms.

Key Study Components

Area of Science

  • Microbiology
  • Immunology
  • Cell Biology

Background

  • Candida glabrata is a facultative intracellular pathogen.
  • Understanding its interaction with macrophages is crucial for insights into fungal infections.
  • THP-1 cell line is commonly used for macrophage differentiation.
  • In vitro models are essential for studying pathogen-host interactions.

Purpose of Study

  • To develop a reliable in vitro model for studying Candida glabrata.
  • To assess the intracellular behavior of the pathogen in human macrophages.
  • To evaluate macrophage responses to Candida infection.

Methods Used

  • THP-1 monocytes are differentiated into macrophages using PMA treatment.
  • Macrophages are infected with Candida glabrata cells.
  • Non-phagocytosed cells are removed through PBS washes.
  • Intracellular cells are recovered for further analysis.

Main Results

  • The protocol allows for the assessment of Candida glabrata virulence in macrophages.
  • It enables the study of cytokine responses in THP-1 macrophages.
  • Insights into the maturation processes of macrophages upon infection are provided.

Conclusions

  • This in vitro model is a valuable tool for studying fungal pathogenesis.
  • It contributes to the understanding of host-pathogen interactions.
  • Future studies can leverage this model to explore therapeutic interventions.

Frequently Asked Questions

What is the significance of studying Candida glabrata?
Candida glabrata is an opportunistic pathogen that poses a significant threat to immunocompromised individuals, making its study crucial for developing effective treatments.
How are THP-1 cells differentiated into macrophages?
THP-1 cells are treated with PMA to induce differentiation into macrophage-like cells.
What techniques are used to analyze intracellular Candida glabrata?
The protocol includes washing non-phagocytosed cells and recovering intracellular cells for further analysis.
What are the expected outcomes of this study?
The study aims to elucidate the virulence mechanisms of Candida glabrata and the immune response of macrophages.
Can this model be used for other pathogens?
Yes, the model can potentially be adapted to study other intracellular pathogens.
What is the role of cytokines in this study?
Cytokines are key indicators of the immune response and are measured to assess macrophage activation during infection.

Der aktuelle Artikel beschreibt ein Protokoll, um eine In-vitro-Zellkultur-Modellsystem, um die Interaktion einer fakultativ intrazellulären Humanfunguspathogen Candida glabrata mit menschlichen Makrophagen, die ein nützliches Werkzeug, um unser Wissen von Pilz Virulenzmechanismen voranzubringen studieren zu etablieren.

Das übergeordnete Ziel dieses Verfahrens ist es, ein in vitro Kulturmodellsystem zu etablieren, um das intrazelluläre Verhalten eines humanen opportunistischen Pilzerregers zu untersuchen. Canida Laborratta in Makrophagen, die aus der humanen monozytären Zelllinie TP one gewonnen wurden. Um dies zu erreichen, werden die Monozyten zunächst mit vier Estern behandelt, um sie zu Makrophagen zu differenzieren.

Als nächstes werden THP 1-Makrophagen zwei Stunden nach der Infektion mit Canida-GTA-Zellen infiziert. Nicht phagozytierte canida-GTA-Zellen werden mit PBS-Waschmaschinen entfernt und intrazelluläre E-Zellen werden für weitere Studien durch Osmos von infizierten Makrophagen gewonnen. Dieses Protokoll kann verwendet werden, um sowohl die viralen zu Canida-GTA-Zellen in Makrophagen als auch die Faiz-Reifungs-Ross- und Zytokinantwort in THP auf Makro-FIDS nach einer Candida-GTA-Infektion zu bewerten.

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