Application of young blood plasma exosome in preparation of medicine for treating senescence-induced degenerative diseases
A technology for degenerative diseases, young people, applied in the biological field, can solve problems such as mitochondrial function decline
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Embodiment 1
[0018] Example 1 Extraction of plasma exosomes
[0019] The method of the above-mentioned plasma exosome extraction kit comprises the following steps:
[0020] (1) Ultrasonic cleaning of the ultracentrifuge special centrifuge tube one day in advance, and drying.
[0021] (2) Set the water bath to 37°C half an hour before the operation.
[0022] (3) Take out the subpackaged frozen plasma, thaw in a water bath at 25-37°C, and place it on ice after thawing.
[0023] (4) Centrifuge at 500g for 5 minutes at 4°C, and take the supernatant.
[0024] (5) Centrifuge at 3000g for 25 minutes at 4°C, and take the supernatant.
[0025] (6) Centrifuge at 10,000g for 60 minutes at 4°C, and take the supernatant.
[0026] (7) First add an appropriate amount of PBS to the 15ml centrifuge tube to dilute, try not to have air bubbles.
[0027] (8) Pour the plasma into the centrifuge tube without air bubbles. Rinse the tube wall and tube cover without air bubbles. The mark faces outward and sli...
Embodiment 2
[0037] After extracting the plasma exosomes from young mice, the exosomes were injected into the old C57BL / 6J mice according to the plasma exosome injection procedure shown in Example 1, and then the mouse hippocampus and muscle tissue proteins were extracted and detected by western bolt Changes of differentially expressed genes in each tissue, see the results figure 1 .
Embodiment 3
[0039] After the plasma exosomes were extracted, the exosomes were co-cultured with nerve cells (NE4C) and muscle cells (C2C12), respectively, and the following indicators were detected after 24 hours:
[0040] (1) The changes of differentially expressed genes in cells were detected by western bolt;
[0041] (2) Detection of cell proliferation ability by EdU;
[0042] (3) The mRNA expression of maker P21, a marker of aging, was detected by qPCR;
[0043] (4) Detect the change of ATP generation rate by ATP generation rate detection kit;
[0044] (5) Detect the changes of mitochondrial complex V activity by the mitochondrial complex V activity detection kit;
[0045] (6) Detection of mitochondrial DNA changes by qPCR;
[0046] (7) The changes of mitochondrial respiratory chain were detected by seahorse instrument mitochondrial stress kit.
[0047] see the results figure 2 .
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