Live and let dye: Probes enabling immune cells to announce their success

Associate Professor Ji-Hwan Ryu
Yonsei University College of Medicine

■ Medicine /

Live and let dye: Probes enabling immune cells to announce their success

Yonsei biochemists develop a fluorescent probe capable of signaling immune-cell destruction of invading pathogens

Among the many roles played by the human immune system, capturing and destroying invading bacterial pathogens is among the most important. However, the destruction of these pathogens results in production of molecules that can be harmful to cells and tissues. Moreover, the accumulation of these molecules can potentially result in the initiation of cardiovascular and inflammatory diseases.

Researchers are continually engaged in the discovery of methods that enable the detection of such molecules in order to promote their use as diagnostic indicators of disease progression. A recent paper by Ryu et al. (2016) and published in Nature Protocols described the creation and application of a novel chemical-based fluorescent probe capable of detecting one such molecule, hypochlorous acid (HOCl).

Upon infection, immune cells called neutrophils capture and destroy pathogens in highly acidic compartments. This process results in the generation of large amounts of HOCl that, when not appropriately managed, can lead to cellular dysfunction. Alternatively, HOCl has also been shown to exert anti-cancer activities and promote processes that help our immune system “learn” about how to effectively fight infections. Given its contrasting roles, methods capable of detecting HOCl have become important for helping researchers determine which biological pathways are associated with its positive and negative effects.

Fluorescent probes are a widely used method allowing scientists to visualize interactions between molecules. In such cases, a fluorescent signal occurs whenever one molecule interacts with its target. Using a low-cost, commercially available dye, Ryu et al. (2016) developed a fluorescent probe specific for HOCl and capable of “finding” this molecule in the highly acidic environments of immune cells. In addition, based on the importance of probes binding only to their specific molecular targets, Ryu et al. (2016) showed that their probe “found” its target at rates that exceeded current commercially available HOCl-specific probes.

Testing the probe in vivo
To demonstrate the use of their probe in different biological environments, Ryu et al. (2016) described experiments performed on both fruit flies and mice. The fruit fly experiment was described in a previous study published by their lab [Lee et al. (2013)] and demonstrated the in vivo ability of the probe to signal the presence of increased amounts of HOCl following bacterial infection.

Success in a mammalian model
In order to demonstrate the efficacy of this method in a mammalian system, immune cells were removed from mice and infected with bacteria. After the cells captured and destroyed the bacteria, resulting in production of HOCl, researchers were able to observe fluorescent signals indicating that the probe had effectively “found” the products and reported their presence. Given the importance of showing the ability of the probe to work under physiological conditions, a respiratory infection was initiated in mice, followed by isolation of immune cells from their lungs. Upon administration of the probe to these cells, the researchers observed fluorescent signals confirming the ability of probe to detect HOCl production in response to infection.

Given the involvement of this molecule in both positive and negative aspects of mammalian diseases, the development of affordable methods that allow researchers to identify the differences between these pathways is critical. The protocols described by Chen et al. (2016) offer an opportunity to investigate HOCl, specifically, as a potential anticancer therapeutic, as well as its role in the progression of diseases, including rheumatoid arthritis and atherosclerosis.

Updated in Feb 2017

X. Chen, K.-A. Lee, X. Ren, J.-C. Ryu, G. Kim, J.-H. Ryu, W.-J. Lee, J. Yoon, “Synthesis of a highly HOCl-selective fluorescent probe and its use for imaging HOCl in cells and organisms”, Nature Protocols, doi: 10.1038/nprot.2016.062