Nanotechnology Now - Press Release: Preserving the goods: A new technique for isolating intact lysosomes from cell cultures: Scientists advance the study of fragile digestive organelles by developing strategy to rapidly extract them from cells using magnetic nanoparticles

Home > Press > Preserving the goods: A new technique for isolating intact lysosomes from cell cultures: Scientists advance the study of fragile digestive organelles by developing strategy to rapidly extract them from cells using magnetic nanoparticles

Once magnetic nanoparticles have naturally accumulated in a cells lysosomes through the endocytic pathway, the cell membrane is ruptured. and its contents sifted for 30 minutes using magnets. At the end of this process, intact lysosomes can be retrieved from the magnets and used to study their structure, metabolites, and protein composition.

CREDIT
Shinya Maenosono from JAIST.

Abstract:
The correct functioning of our cells rests upon the precise orchestration of many complex processes and organelles. Lysosomesvital cell organellesare enzyme-filled subunits found within many animal cells that help break down and reuse macromolecules, such as proteins, lipids, and nucleotides. Besides their function in cellular digestion and waste management, lysosomes also participate in amino acid signaling, which stimulates protein synthesis alongside other effects.

Preserving the goods: A new technique for isolating intact lysosomes from cell cultures: Scientists advance the study of fragile digestive organelles by developing strategy to rapidly extract them from cells using magnetic nanoparticles

Ishikawa, Japan | Posted on January 7th, 2022

Given that a lot of diseases are caused by defects in lysosome function, it is no surprise that researchers have been actively trying to understand these organelles for decades. But there are only a few techniques that allow the extraction of lysosomes from within a cell. The most common method is called density gradient ultracentrifugation. It involves gently breaking the cell membrane and applying a centrifugal force to the cells contents. This separates the cell components by density. Unfortunately, some other organelles have the same density as lysosomes, resulting in samples with impurities. Moreover, the process takes so long that by the time it finishes, many lysosomal proteins have already been lost and/or degraded.

A more advanced technique, called immunoprecipitation, relies on modifying the surface proteins of lysosomes so that they can be captured by magnetic beads covered in specially tailored antibodies. While this approach produces purer results, the protein composition of the extracted lysosomes is modified by the procedure and, thus, subsequent protein analyses can be compromised. It is clear, then, that we need to find a better way to extract lysosomes from cells.

Fortunately, a team of scientists led by Prof. Shinya Maenosono from the Japan Advanced Institute of Science and Technology (JAIST) has stepped up to the plate and developed a novel strategy to quickly separate intact lysosomes with high purity. This study was published in ACS Nano and also included Prof. Kazuaki Matsumura and Associate Prof. Yuichi Hiratsuka from JAIST, and Prof. Tomohiko Taguchi of Tohoku University, Japan.

Their strategy is centered around the use of magnetic-plasmonic hybrid nanoparticles (MPNPs) made of silver and an ironcobalt alloy and covered in a compound called amino dextran (aDxt). The basis for this approach is that the aDxt-covered MPNPs are naturally ingested by the cells through endocytosis, which culminates inside lysosomes. Once enough MPNPs have accumulated inside the lysosomes, the cells can be gently crushed, and the lysosomes retrieved using magnets.

For this method to work, it is essential that MPNPs are located only within lysosomes and not in other organelles. This is where plasmon imaging comes in handy, as the distinct way plasmonic nanoparticles interact with light makes them easy to visualize with an optical microscope. By coloring each type of organelle in the endocytic pathway differently using immunostaining and checking how the location of MPNPs overlaps with them, the researchers determined the precise time it takes most MPNPs to reach the lysosomes. In turn, this ensures that the separation process yields lysosome samples with high purity.

Afterwards, the team analyzed the effects of temperature and magnetic separation time on the protein composition of the extracted lysosomes. Their results showed that protein loss was remarkably quick, even at temperatures as low as 4°C. Fortunately, their approach was fast enough to extract intact lysosomes, as Prof. Maenosono highlights: We found that the maximum time required to isolate lysosomes after cell rupture was 30 minutes, which is substantially shorter than the time required using centrifugation-based techniques, which typical require a minimum separation time of several hours.

Overall, this new technique will help researchers explore the fragile metabolites of lysosomes and how they change in response to stimuli. In turn, this shall pave the way to new insights into disorders related to lysosomal dysfunction. In this regard, Prof. Maenosono remarks: Given the profound relation of lysosomes with many cellular metabolites, a deeper understanding of lysosomal function is necessary to determine its regulation in different cell states. Therefore, our technique can contribute to better understanding and treatment of lysosomal diseases in the future. Moreover, this new approach could be modified to extract other organelles besides lysosomes. Hopefully, this study will put us closer to understanding the contents of cells to a much higher degree.

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About Japan Advanced Institute of Science and Technology (JAIST)
Founded in 1990 in Ishikawa prefecture, the Japan Advanced Institute of Science and Technology (JAIST) was the first independent national graduate school in Japan. Now, after 30 years of steady progress, JAIST has become one of Japans top-ranking universities. JAIST counts with multiple satellite campuses and strives to foster capable leaders with a state-of-the-art education system where diversity is key; about 40% of its alumni are international students. The university has a unique style of graduate education based on a carefully designed coursework-oriented curriculum to ensure that its students have a solid foundation on which to carry out cutting-edge research. JAIST also works closely both with local and overseas communities by promoting industryacademia collaborative research.

About Professor Shinya Maenosono from Japan Advanced Institute of Science and Technology, Japan

Shinya Maenosono received a PhD in Chemical Systems Engineering from the University of Tokyo, Japan, in 2002. He then joined the Japan Advanced Institute of Science and Technology (JAIST) as an Associate Professor in 2006. In 2012, he became a full Professor at JAIST, where he leads a research group. His research in JAIST has focused on the wet chemical synthesis of semiconductor nanoparticles with controlled size, shape, and composition for applications in energy conversion devices, as well as the synthesis and bioapplication of monometallic and alloyed multimetallic nanoparticles. He has published over 120 papers and 12 book chapters.

Funding information

This study was supported by the Grant-in-Aid for Young Scientists from the Japan Society for the Promotion of Science (grant no. 21K14506)

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Contacts:
Shinya Maenosono
Japan Advanced Institute of Science and Technology

Office: +81-761-51-1611

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