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Published on: October 6, 2014
Monitoring Circulating Myeloid Cells in Peritonitis with an In Vivo Imaging Flow Cytometer
Sunitha Pulikkot1, Souvik Paul2,3, Alexxus Hall1
1Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA.
Abstract:
Peritonitis is a common and life-threatening inflammatory disease. Myeloid cells are elevated in the peripheral blood and contribute to peritonitis, but their circulating dynamics are not clear. In vivo flow cytometry (IVFC) is a noninvasive technique for monitoring the dynamics of circulating cells in live animals. It has been extensively used to detect circulating tumor cells, but rarely for monitoring immune cells. Here, we describe a method adapting an intravital microscope for IVFC so that we can monitor LysM-EGFP-labeled circulating myeloid cells in a tumor necrosis factor (TNF) α-induced peritonitis mouse model. Using this IVFC method, we quantified the blood flow velocity and cell concentration in circulation. We observed a significant increase in LysM-EGFP+ cells in circulation after TNFα intraperitoneal (i.p.) injection, which reached a plateau in ~20 min. Conventional cytometry analysis showed that most LysM-EGFP+ cells were neutrophils. Increasing blood neutrophils were accompanied by neutrophil recruitment to the peritoneal cavity and neutrophil emigration from the bone marrow. We then monitored neutrophil CD64 expression in vivo and found a significant increase in TNFα-induced peritonitis. We also found that CD18 blockade doubled the circulating neutrophil number in TNFα-induced peritonitis, suggesting that CD18 is critical for neutrophil recruitment in peritonitis. Overall, we demonstrate that IVFC techniques are useful for studying the circulating dynamics of immune cells during inflammatory diseases.
Insights
This study introduces in vivo flow cytometry (IVFC) to track myeloid cells during peritonitis. IVFC reveals increased circulating neutrophils after TNFα injection, aiding understanding of immune cell dynamics in inflammation.
Area of Science:
- Immunology
- Cell Biology
- Medical Imaging
Background:
- Peritonitis is a severe inflammatory condition where myeloid cells play a key role.
- The dynamic behavior of circulating myeloid cells in peritonitis remains poorly understood.
- In vivo flow cytometry (IVFC) is an established technique for monitoring circulating tumor cells but less utilized for immune cells.
Purpose of the Study:
- To adapt and apply in vivo flow cytometry (IVFC) for monitoring circulating myeloid cells in a mouse model of peritonitis.
- To investigate the dynamics of myeloid cells, particularly neutrophils, during tumor necrosis factor (TNF) α-induced peritonitis.
- To explore the role of CD18 in neutrophil recruitment during peritonitis.
Main Methods:
- Adaptation of intravital microscopy for IVFC to monitor LysM-EGFP-labeled myeloid cells.
- Induction of peritonitis using intraperitoneal (i.p.) injection of TNFα in mice.
- Quantification of circulating cell concentration, blood flow velocity, and neutrophil CD64 expression using IVFC and conventional cytometry.
- Assessment of CD18 blockade's effect on circulating neutrophil numbers.
Main Results:
- IVFC successfully monitored circulating LysM-EGFP+ myeloid cells in a TNFα-induced peritonitis model.
- A significant, rapid increase in circulating neutrophils was observed post-TNFα injection, peaking around 20 minutes.
- Neutrophil CD64 expression increased significantly during peritonitis.
- CD18 blockade doubled circulating neutrophils, highlighting its critical role in peritonitis-related recruitment.
Conclusions:
- In vivo flow cytometry (IVFC) is a valuable tool for studying the dynamics of circulating immune cells during inflammatory conditions like peritonitis.
- TNFα induces rapid changes in circulating neutrophil populations and expression of activation markers.
- CD18 is essential for neutrophil recruitment in the context of peritonitis.
