3D Imaging Shows How Cytotoxic T Lymphocytes Kill Target Cells

Cytotoxic T lymphocytes are the body’s specialized “killer” cells, precisely eliminating infected or cancerous cells. Their action relies on a specialized exchange zone called the “immune synapse,” where they release active molecules to destroy the target cell without damaging neighboring ones. Until now, the fine organization of these structures remained difficult to observe. A study conducted by the University of Geneva (UNIGE) and the Lausanne University Hospital (CHUV) has made it possible to visualize these mechanisms in three dimensions in a near-native state. Published in Cell Reports, the study reveals how the molecular organization of cytotoxic T cells underpins their function, opening new perspectives in immuno-oncology.

During infection or cancer, cytotoxic T lymphocytes attach to their target and establish an exchange zone known as the immune synapse, then release toxic molecules that trigger the death of the targeted cell. This mechanism enables precise and controlled destruction, essential for protecting the body while avoiding damage to nearby healthy cells.

By providing a three-dimensional and near-native view of these processes, this work establishes a reference framework for analyzing how immune cells function.

Although this process has been widely studied, its organization at the nanometer scale in intact human cells remained difficult to access. One of the main obstacles lies in sample preparation methods, which can alter fragile cellular structures. Existing imaging approaches often involve trade-offs between resolution, observable volume, and preservation of structures.

A technique to see the invisible

To overcome these limitations, a study by UNIGE and CHUV-UNIL, supported by the ISREC Foundation TANDEM program, relied on cryo-expansion microscopy (cryo-ExM). “This technique involves instantaneously freezing cells at very high speed, placing them in a so-called vitreous state, where water solidifies without forming crystals and thus faithfully preserves biological structures. The samples are then physically expanded using an absorbent hydrogel, making it possible to observe their internal organization with great precision while maintaining their near-native architecture,” explains Virginie Hamel, senior lecturer in the Department of Molecular and Cellular Biology at the Faculty of Science of UNIGE.

“Our work reveals that at the point of contact between the immune cell and its target, the membrane forms a kind of dome, whose structure appears to be linked to adhesion interactions  and to the internal organization of the cell,” notes Florent Lemaître, postdoctoral researcher in the Department of Molecular and Cellular Biology at the Faculty of Science of UNIGE and first author of the study. The research team also visualized cytotoxic granules -responsible for destroying target cells – with an unprecedented level of detail. The study shows that these structures can vary in their organization, with one or more “cores” concentrating the active molecules that enable the destruction of the target cell.

From cells to patients

“We extended this approach to human tumor tissues, making it possible to directly observe T lymphocytes infiltrating tumors and their cytotoxic machinery at the nanometer scale. This allows us to study immune responses directly in their clinical context and to better understand the mechanisms that determine their effectiveness,” explains Benita Wolf, Chief Resident and associate researcher in the Department of Clinical Oncology at CHUV, who co-led the study.

By providing a three-dimensional and near-native view of these processes, this work establishes a reference framework for analyzing how immune cells function. It could help improve therapeutic strategies, particularly in immuno-oncology, by enabling a better understanding of the mechanisms that determine the effectiveness - or limitations – of the immune response.

“Our work reveals that at the point of contact between the immune cell and its target, the membrane forms a kind of dome, whose structure appears to be linked to adhesion interactions  and to the internal organization of the cell,” notes Florent Lemaître, postdoctoral researcher in the Department of Molecular and Cellular Biology at the Faculty of Science of UNIGE and first author of the study. The research team also visualized cytotoxic granules -responsible for destroying target cells – with an unprecedented level of detail. The study shows that these structures can vary in their organization, with one or more “cores” concentrating the active molecules that enable the destruction of the target cell.

From cells to patients

“We extended this approach to human tumor tissues, making it possible to directly observe T lymphocytes infiltrating tumors and their cytotoxic machinery at the nanometer scale. This allows us to study immune responses directly in their clinical context and to better understand the mechanisms that determine their effectiveness,” explains Benita Wolf, Chief Resident and associate researcher in the Department of Clinical Oncology at CHUV, who co-led the study.

By providing a three-dimensional and near-native view of these processes, this work establishes a reference framework for analyzing how immune cells function. It could help improve therapeutic strategies, particularly in immuno-oncology, by enabling a better understanding of the mechanisms that determine the effectiveness - or limitations - of the immune response.