2022 Fiscal Year Research-status Report
Identification of early stage malignancy changes in the structural and biophysical proporties of exosomes by atomic force microscopy (AFM)-based nano-mechanical measurements
| Project/Area Number |
20K05321
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| Research Institution | Kanazawa University |
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Principal Investigator |
YURTSEVER AYHAN 金沢大学, ナノ生命科学研究所, 特任助教 (00761529)
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| Project Period (FY) |
2020-04-01 – 2024-03-31
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| Keywords | exosomes / Atomic force Microscopy / Nanomechanics / Cells / 3D-AFM |
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| Outline of Annual Research Achievements |
We applied 3D-AFM to assess the structural and nanomechanical properties of exosomes released from different types of cells in a physiologically
relevant environment. We established A method for studying the interaction of molecules exposed on the exosomes by using AFM. Exploring the molecule exposed on the native exosome surface will deepen our understanding of the physiological function of exosomes in cancer progression. In addition, we performed AFM measurements to identify the exosomes in heterogeneous extracellular vesicles (EVs) and analysis their biophysical properties. There is no method that can isolate exosomes (EXOs) or microvesicles (MVs) without including other EVs. As a substitute for EXOs, intraluminal vesicles (ILVs), which are the origin of EXOs were isolated from cells. After separating EVs by the anion exchange column, we performed AFM nanomechanical characterization to determine the biophysical properties of exosomes and other EV subtypes.
AFM nanomechanical measurements determined the rapture characteristic of EVs. We have found differences in their rapture mechanics, implying difference of membrane structure related to elastic fibers. Mass spectroscopic analyses were performed to determine their protein and lipid compositions.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
Research progress is sufficient. We achieved most of our previously planned research objectives. We determined the structural and mechanical characteristic of normal and cancer-derived exosomes for different cell types.
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| Strategy for Future Research Activity |
The biochemical composition of exosomes will be determined using functionalized AFM probes with biomolecules specific to exosome surface receptors and proteins Furthermore, we will perform nanomechnaical measurements of exosomes from different cell types (PC-9, PC-9 BrM4 and WM266.4, WM266.4 BrM3 cells).
The internal composition content of exosomes, such as miRNA and DNAs, will be determined by high resolution AFM imaging and force mapping by rupturing the lipid membrane of exosomes.
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| Causes of Carryover |
I had planned to attend domestic and international conferences, but due to corona many conferences were canceled or went online. Because of that, I couldn't go on a business trip, so the plan has changed; some funds will be used in the next fiscal year.
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