Inflammasome Assembly Inside Human Cells Captured for the First Time

For the first time, researchers have captured the formation of a key driver of inflammatory responses inside intact human cells. Pioneering a new system for observing molecular assemblies, researchers discovered that the NLRP3 inflammasome has a dense, gel-like structure that is much less organized than expected.

The findings, published in the journal Science Advances, provide new insights into the body's inflammatory response, which could influence the design of treatments for inflammatory diseases. The observations also help explain why cells tend not to divide while mounting an inflammatory response.

“In structural biology, we use a very popular quote: ‘Seeing is believing’. Nothing can compare with actually seeing the structure,” lead author Dr. Phyllis Wang, research associate at SLAC National Accelerator Laboratory and Stanford University, told Technology Networks. “Using our workflow, we started to systematically visualize how molecular machines involved in inflammation are organized and function in situ. This has broad implications for both basic biology and therapeutic targeting.”

Refining our understanding of inflammation

Inflammation is triggered when cells detect infection or other stress signals. When cells sense danger, they utilize molecular warning systems, such as inflammasomes, to trigger the immune system.

One of the most extensively studied inflammasomes is NLRP3, which has been implicated in the development of common diseases such as Alzheimer’s disease, cardiovascular diseases, and type 2 diabetes. “It is also one of the few inflammasomes that localize to the centrosome, making it very interesting for us to study in the spatial organization of cells,” said Wang.

Experiments performed on purified proteins outside the cell suggest that NLRP3 forms a highly ordered disc-like structure; however, its structural organization in cells is poorly understood.

A new system for imaging structures in live cells

The researchers had to overcome significant technical challenges with traditional structural biology techniques to visualize NLRP3 assembling inside human cells. “The challenge was being able to target the structure precisely, especially as the structure is smaller than one micron,” explained Wang.

To preserve native structure while targeting specific regions of interest in the cell, the researchers used fluorescence-guided cryo–focused ion beam (cryo-FIB) milling and cryo–electron tomography (cryo-ET). This allowed the researchers to generate 3D reconstructions of cellular structures preserved in frozen ultra-thin sections of the cell.

“The first step was to acquire an initial fluorescent image to localize the target of interest, in this case, the NLRP3 inflammasome. We then used the focused ion beam as a precise milling tool to remove material from the top and bottom of the cell, keeping the region of interest centered within the remaining volume. By carefully adjusting the milling current, we were able to produce a lamella, around 100 to 200 nm thick, which was flat and suitable for high-quality imaging by cryo-ET,” Wang said.

Developing this method took years of work and required retooling the cryo-FIB instrument to include a light microscope to better direct the milling process.

“Using this new way of guiding the milling of cells, you're able to capture the biology you’re interested in almost 100% of the time,” Dr. Peter Dahlberg, an assistant professor at SLAC National Accelerator Laboratory and Stanford University, told Technology Networks.

The role of NLRP3 in cell division

Applying the new imaging technique to the NLRP3 inflammasome revealed a structure that looked surprisingly different from the structure often depicted in models. NLRP3 appeared to form a dense, gel-like cluster of proteins and signal molecules that accumulated around the centrosome.

“We had this picture from in vitro studies purifying these complexes and looking at them in isolation. While there's still insight to gain from that, looking at these things in their native environment can yield surprises like this, and it's worth remembering that what you see in situ isn't necessarily what you see out of the cell,” Dahlberg said.

By observing the formation of NLRP3, the researchers discovered that as the structure expands, it disrupts the positioning of centrioles—microtubule-based organelles found in pairs that together form the centrosome. Centriole separation is one of the earliest steps in cell division, but NLRP3 appears to trap the centrioles, preventing cell division. This observation helps explain why inflammasome activation and normal cell division are typically mutually exclusive.

Updating the toolkit for fighting inflammatory disease

Traditional pharmaceutical strategies targeting NLRP3 have focused on blocking movement within the inflammasome based on the belief that NLRP3 is a rigid protein structure. However, these new observations suggest that NLRP3 is in fact a flexible protein condensate, perhaps composed of fragments of the structures observed in in vitro isolated studies.

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“You can apply a different strategy for therapeutic design when targeting a protein condensate versus a protein complex structure. Condensates are driven by multiple interactions, and this opens up new opportunities to modulate the process,” said Wang.

While these new findings shed light on NLRP3 formation, the researchers caution against assuming it is a general principle applicable to all types of systems. Further research is required to determine whether the same mechanism applies to other inflammasomes and cell types outside of the human macrophages used in this study.

“One of the most exciting directions is to apply this customized fluorescence-guided cryo-FIB system to other protein assemblies, especially those that are challenging to study using the traditional structural approaches. And that's one of the future directions we are pushing,” concluded Dahlberg.

Reference: Wang J, Wu M, Xiao L, et al. Human NLRP3 inflammasome activation leads to formation of condensate at the microtubule organizing center. Sci Adv. 2026;12(13):eaee2473. doi: 10.1126/sciadv.aee2473

About the interviewees

Dr. Peter Dahlberg received his undergraduate degree in physics at McGill University in 2011 and his PhD in biophysics from the University of Chicago in 2016 for his work on developing ultrafast spectroscopy methods for the study of photosynthetic energy transfer. He then joined Stanford as a postdoc to work with W. E. Moerner and Wah Chiu to develop correlative light and electron microscopy methods. In 2021, he was awarded SLAC’s Panofsky Fellowship to continue his work on advanced correlative microscopy. He joined the departments of photon science and structural biology at Stanford as an assistant faculty member in 2025.

Dr. Phyllis Wang is a structural biologist and immunologist whose work focuses on revealing the in situ molecular architecture of innate immune protein machineries. Her ongoing research uses fluorescence-guided cryo-focused ion beam milling and cryo-electron tomography to study NLRP3 inflammasomes, while also contributing to the development of advanced integrated cryogenic imaging technologies. Beyond research, Wang is passionate about supporting women in STEM and fostering connections between science and the arts through community outreach and chamber music.