Macrophages are an important category of immune cells that patrol our body to find and destroy pathogens, often by swallowing them whole — a process known as phagocytosis. A recent study by researchers at the Indian Association for the Cultivation of Science (IACS), Kolkata, has discovered how the physical properties of macrophages change in response to phagocytosis and how this, in turn, affects their function.
Macrophages are immune cells that play a primary role in our body’s first line of immunity. Since their activity is associated with a number of lifestyle conditions, any factor that affects macrophage function can lead to compromised immunity and a heightened risk of infection.
Macrophages routinely patrol deep tissues to pick up and engulf pathogens and dying cells. Both actions require considerable deformation of these large cells. Recently, Deepak Sinha and his team at Indian Association for the Cultivation of Science (IACS), Kolkata, have discovered a molecular culprit — reactive oxygen species (ROS) — that can hamper the ability of macrophages to deform by causing their cellular regions to stiffen.
How do macrophages undergo such massive deformation? Though it might be tempting to imagine cells as squeezable blobs, their content is actually structured on a framework of skeletal elements — much like a tent supported by poles. These skeletal elements are rigid structures that can be dynamically remodelled when a cell needs to change its shape. They can be disassembled at one part of the cell and quickly reassembled elsewhere to help the canvas (the cell in this case) deform. When, for any reason, these elements fail to reassemble rapidly, it leads to a stiffening of the cell.
These skeletal elements are made up of small proteins called actin monomers which assemble end-to-end to form filaments (the tent poles). When the authors allowed macrophages grown in the laboratory to engulf a fluorescent bead (representing a pathogen) they found that this resulted in a chemical modification of the monomers. This modification, known as glutathionylation, prevented the actin monomers from reassembling into filaments, causing cells to stiffen. This modification is catalysed by a build-up of ROS – a group of oxygen-containing unstable molecules whose levels are known to increase inside macrophages in response to engulfing a pathogen.
“Within a cell, the physical properties are not homogeneous, some pockets can be softer (or gel-like), the others stiffer,” says Deepak Sinha. The researchers were able to measure the deforming ability of the cell by a technique known as particle tracking microrheology which analyses two aspects of matter — viscosity and elasticity.
The cytoplasm — the jelly-like substance inside the cell — falls somewhere in between a solid and a liquid in its behaviour. Viscosity, typically the property of liquids, refers to the amount of resistance a substance offers to flowing freely. Elasticity, usually considered a property of solids, refers to the extent to which a material can recover its original shape after being deformed. The combination of these two properties — viscoelasticity - can help us understand the mechanical behaviour of the cytoplasm.
Particle tracking microrheology involves monitoring and measuring the movement of fluorescent beads to understand how cellular material flows within a sub-region of the cell. Cellular regions with high viscoelasticity can be called “soft” while those with lower viscoelasticity appear “stiff”.
Normal human macrophages have the potential to ingest more than one particle one after the other. Since researchers observed that macrophages stiffen upon ingestion, they wanted to understand if it would hamper ingestion of other particles immediately afterwards. When they fed the macrophages with two differently fluorescing beads, they found out that macrophages that had engulfed one bead could not engulf another. The ROS levels took about 8 hours to reduce to levels before ingestion – suggesting that the macrophage takes about as much time to be ready for engulfing another bead.
While these experiments were conducted on macrophage cell lines growing in culture, whether human macrophages behave similarly within the body (in vivo) is yet to be understood. On this, Sinha says, “An extrapolation of this estimate to macrophages from humans is not possible. However, we plan to conduct relative experiments to see if ageing and lifestyle impact phagocytic ability.”
Avinash Sonawane, a scientist at Indian Institute of Technology (IIT), Indore, not associated with the study, says, “While the research is extensive, there could be an interplay of complex factors that could impact actin turnover, which needs to be further explored.”
The study can be intriguing from the perspective of infectious biology — would a delay in the ingestion of a second particle benefit the pathogen or the host? Also, given that cellular ROS levels can be influenced by a variety of factors, like ageing and alcohol consumption, it would be interesting to investigate if these factors affect the flexibility and function of macrophages.