A new dimension in transplantation

Researchers at ETH Zurich have announced a technological breakthrough: they have developed a method with which mitochondria - the tiny power plants within a cell - can be transferred from one living cell to another with unsurpassed efficiency.

Organ transplantation at cellular level: researchers use a nanosyringe to suck up mitochondria (blue) from a living cell in order to transfer the organelles to another cell

Just as the human body can be divided into different organs - such as the heart, lungs, kidneys, intestines or liver - our cells also consist of several complementary and interdependent systems, which are referred to as organelles (i.e. small organs) in technical jargon. And just as the life of a person with kidney disease can sometimes be extended by several decades with the transplantation of a healthy kidney, individual cells could perhaps one day be refreshed with the transplantation of cell components.

The new findings of Julia Vorholt's research group from the Institute of Microbiology at ETH Zurich show that this thought experiment is not just a pipe dream, but has moved into the realm of technical feasibility. As the scientists have just reported in the journal PLos Biology, they have transplanted mitochondria from one living cell into another using a nanosyringe that they had previously developed.

The biochemical processes of cellular respiration, which evolved in bacteria more than two billion years ago, take place in these tiny cellular power plants. Later, some bacteria formed a close community with other cells, a so-called endosymbiosis. This plays a central role in the phylogenetic history of life on earth: it was this that enabled the development of fungi, plants and animals (including us humans), which are all made up of complex cells.

Mitochondrial transplantation (A) Scheme of mitochondrial transplantation using the cell-to-cell transfer approach: Mitochondria are extracted by FluidFM aspiration. Subsequently, the cantilever containing the extract is moved to a recipient cell and the extract is injected. (B) Image of a FluidFM cantilever prefilled with perfluorooctane after mitochondrial extraction, mitochondria are labeled with su9-mCherry. The extracted volume is about 0.8 pL. Scale bar: 10 μm. (C) Schematic of mitochondrial transplantation using mitochondria purified according to a standard mitochondrial purification protocol. The purified mitochondria are resuspended in HEPES-2 buffer and filled directly into the fluidic probe. The cells are injected one after the other. (D) Image of a FluidFM cantilever filled with mitochondria isolated from the mass and labeled with su9-mCherry. Scale bar: 10 μm. (E) Images of a recipient cell after transplantation of mitochondria via the cell-to-cell approach. The mitochondrial network of the host cells is labeled with su9-BFP, the graft with su9-mCherry. Scale bar: 10 μm. (F) Images of a recipient cell after mitochondrial transplantation by injection of isolated mitochondria, labeling as in c. Scale bar: 10 μm. (G) Evaluation of mitochondrial transplantation via the cell-to-cell approach after visual inspection and the isolated mitochondria injection approach. A total of 40 cells per approach were evaluated. (H) Absolute number of transplanted mitochondria from 22 individual cells scored for the cell-to-cell approach and injection of isolated mitochondria. (I) Fusion states of transplanted mitochondria 30 years after cell-to-cell transplantation. Mitochondria are visualized with different fluorescent labels for the graft (su9-mCherry) and for the host mitochondrial network (su9-BFP). Scale bars: 5 μm. (J) Fusion states of transplanted mitochondria 30 after injection of purified mitochondria, similar labeling as in g. Scale bar: 5 μm. (K) Degradation of transplanted mitochondria, the graft is divided into several smaller fluorescent vesicles (su9-mCherry) with no overlap of fluorescence with the labeled host cell mitochondrial network (su9-BFP). Scale bar: 5 μm. (L) Time-lapse image series of a single transplanted mitochondrion (su9-mCherry). The organelle donor was a HeLa cell, the recipient cell is a U2OS cell with a fluorescently labeled mitochondrial network (su9-BFP). Scale bar: 10 μm (source: [2])

This is how mitochondria have evolved from ancient bacteria over time: Organelles that are responsible for energy production in today's complex cells. In human cells, mitochondria form a thread-like dynamic network. "The threads react to negative pressure - and transform into a kind of string of pearls from which individual mitochondria split off," says Christoph Gäbelein, the first author of the article.

Schematic representation of organelle extraction and injection with FluidFM.
(A) The extraction volume is set by applying a negative pressure (-Δp). Pre-filling the probe with octadecafluorooctane enables optical and physical separation of the extract within the cantilever. (B) Selective extraction of organelle components by adjusting the aperture size and thus the range of applicable flow forces. Top row: Schematic representation of the extracted cell components within the cantilever. Middle row: Scanning electron micrographs of cantilever tips with different apertures. Bottom row: Range of applicable fluidic forces with matched FluidFM cantilevers. Scale bar: 2 μm. (C) Schematic representation of mitochondria injection into single cells by applying positive pressure (+Δp) once the cantilever is inserted into the recipient cell. ER, endoplasmic reticulum (source: [2])

Using angled-ended, cylindrical nanopipettes specially developed for this study, the researchers pierced through the cell membrane - and sucked in the spherical mitochondria. They then pierced the membrane of another cell and pumped the mitochondria back out of the nanopipette into the recipient cell. (see figure)

The position of the nanopipette is controlled by the laser light of a converted atomic force microscope. A pressure regulator adjusts the flow of liquid. This allows unimaginably small volumes in the femtoliter range to be moved during organelle transplantation. Both the donor and the acceptor cells survive this minimally invasive procedure (see illustration).

Over 80 % of the transplanted mitochondria also survive the operation. In most cells, the injected mitochondria begin to fuse with the filamentous network of the new cell after twenty minutes. "They are accepted by the host cell," says Julia Vorholt. Only in a few cells do they fall victim to the quality control of the new host cells - and are degraded.

"In the future, the technology presented here will enable applications in various areas of research," the researchers write. It is conceivable, for example, that it could be used to rejuvenate stem cells whose metabolic activity declines with increasing age. However, Vorholt's team has other plans. "We want to understand the processes that control the cooperation of the various cell compartments - and understand how endosymbioses evolve over time," says Vorholt.

https://ethz.ch

Literature

[1] Source: ETH Zurich
[2] Gäbelein CG, Feng Q, Sarajlic E, Zambelli T, Guillaume-Gentil O, Kornmann B, Vorholt JA. Mitochondria transplantation between living cells. PLoS Biol. 20: Published: March 23, 2022, doi: 10.1371/journal.pbio.3001576call_made