Protective effect of cyanidin-3-glucoside on hacat cell injury induced by UVB
A technology of glucoside and cyanidin, which is applied in the direction of medical preparations containing active ingredients, cosmetic preparations, skin care preparations, etc., to improve the survival rate, prevent and treat skin photodamage, and reduce apoptosis. Effect
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Embodiment 1
[0051] Example 1 Study of cyanidin-3-glucoside interfering with UVB-damaged HaCaT cells ROS / COX-2 pathway
[0052] 1. Experimental method
[0053] 1. HaCaT cell culture
[0054] (1) Cell recovery
[0055] 1) Take out the cryopreservation tube containing HaCaT cells from the -80°C refrigerator, put it into warm water at 37°C immediately, and shake it from time to time to make it evenly heated and melt as soon as possible;
[0056] 2) When the cryopreservation solution is completely dissolved, take out the cryopreservation tube, suck out the cell suspension in a sterile table, inject it into 9ml cell culture medium, blow it evenly, and centrifuge at 1000rpm×5min;
[0057] 3) Discard the supernatant, add 6ml of culture medium, pipette the discrete cells, transfer to a 10mm culture dish, and place in 5% CO 2 , Cultured in a 37°C incubator.
[0058] (2) Cell culture
[0059] 1) Cells were grown in DMEM medium containing 10% FBS, placed in 5% CO 2 , Cultivated in a 37°C incuba...
Embodiment 2
[0170] Example 2 Effect of C3G on apoptosis of HaCaT cells after UVB irradiation
[0171] 1. Apoptosis detection——Annexin V-FITC / PI double staining flow cytometry detection
[0172] The principle of Annexin V-FITC / PI double staining method for detecting cell apoptosis: in the early stage of cell apoptosis, phosphatidylserine (PS) located inside the cell membrane can migrate to the outside of the cell membrane, and Annexin V, as a phospholipid-binding protein, It has a high binding force to PS, so the fluorescence intensity of Annexin V-labeled fluorescein (FITC) can reflect early apoptotic cells; at the same time, because PS of necrotic cells is also exposed outside the cell membrane and is highly stained to PI, the binding Apoptotic (early apoptosis) and necrotic (late apoptosis) cells can be distinguished using PI exclusion. On the scatter diagram of flow cytometry analysis, Annexin V and PI double negatives represent normal active cells (lower left quadrant); Annexin V pos...
Embodiment 3
[0178] Example 3 Effect of C3G on Mitochondrial Membrane Potential of HaCaT Cells After UVB Irradiation
[0179] 1. Mitochondrial membrane potential (Δψm) detection—JC-1 staining flow cytometry detection
[0180]The principle of JC-1 detecting the change of mitochondrial membrane potential: JC-1 is a cationic lipid fluorescent dye, which has two states of monomer and polymer. exist in multimeric form. In normal cells, due to the polarity of the mitochondrial membrane potential (Δψm), JC-1 is rapidly absorbed into the mitochondria depending on the polarity of Δψm, and forms multimers in the mitochondria due to its increased concentration, and the multimers emit The light is red fluorescence, which can be detected by the red (FL-2) channel of the flow cytometer; when cells undergo apoptosis, the mitochondrial membrane potential is depolarized, resulting in the release of JC-1 from the mitochondria, reducing its concentration , so it exists in the form of a monomer and emits gr...
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