Establishment of an <em>In vitro</em> System to Study Intracellular Behavior of <em>Candida glabrata</em> in Human THP-1 Macrophages

Maruti Nandan Rai; Sapan Borah; Gaurav Bairwa; Sriram Balusu; Neelima Gorityala; Rupinder Kaur

doi:10.3791/50625

Istituzione di un In vitro Sistema per studiare intracellulare Comportamento Candida...

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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

Istituzione di un In vitro Sistema per studiare intracellulare Comportamento Candida glabrata In umani THP-1 macrofagi

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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.

L'attuale articolo descrive un protocollo per stabilire un sistema cellulare modello di coltura in vitro per studiare l'interazione di una facoltativo intracellulare patogeno fungino umano Candida glabrata con i macrofagi umani che saranno un utile strumento per far progredire la nostra conoscenza dei meccanismi di virulenza fungine.

L'obiettivo generale di questa procedura è quello di stabilire un sistema modello di coltura in vitro per studiare il comportamento intracellulare di un patogeno fungino opportunista umano. Canida lab ratta in macrofagi derivati dalla linea cellulare monocitica umana TP one. Per ottenere questo THP, un monocito viene prima trattato con quattro esteri per differenziarli in macrofagi.

Successivamente, i macrofagi THP 1 vengono infettati con cellule GTA di canida due ore dopo l'infezione. Le cellule GTA di canida non fagocitosio vengono rimosse dai lavatori PBS e le cellule e intracellulari vengono recuperate per ulteriori studi mediante osmos di macrofagi infetti. Questo protocollo può essere utilizzato per valutare sia le cellule GTA virali a canide nei macrofagi, sia la risposta di Ross e citochine di maturazione faiz nella THP su macro FIDS in seguito all'infezione da candida GTA.

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