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Method for controlling apoptosis and its application in inhibiting tumor growth

A control method and cell generation technology, applied in the fields of biotechnology and medicine, can solve the problems of inability to obtain further application, penetrating cell damage, etc., and achieve the effect of inhibiting growth and high safety

Inactive Publication Date: 2019-09-03
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the use of light as a means of regulation has some limitations that prevent it from being further applied, such as the limited penetration of light in the living body and the damage to cells caused by thermal effects.

Method used

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  • Method for controlling apoptosis and its application in inhibiting tumor growth

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Hela cells of appropriate density were evenly spread into 12-well plates and cultured for 24 hours, transfected with MscL plasmid, and after 48 hours, the expression of the target protein was verified by a fluorescence microscope, and then fresh culture medium was replaced, and ultrasonic radiation with a power of 15W and a frequency of 6MHz was carried out. The time is 30 minutes. After continuing to culture for 24 hours, the cells were washed with phosphate buffered saline, and then stained with Calcein-AM / PI mixed fluorescent dye for dead and alive cells for 10 minutes, and finally flow cytometry was performed to detect that the apoptosis rate was about 60%.

Embodiment 2

[0037] 4T1 cells with appropriate density were evenly spread into 12-well plates and cultured for 24 hours, transfected with MscL plasmid, and after 48 hours, the expression of the target protein was verified by a fluorescent microscope, and then fresh culture medium was replaced, and ultrasonic radiation with a power of 15W and a frequency of 6MHz was carried out. The time is 60 minutes. After continuing to culture for 24 hours, the cells were washed with phosphate buffered saline, and then stained with Calcein-AM / PI mixed fluorescent dye for dead and alive cells for 10 minutes. Finally, flow cytometry was performed to detect that the apoptosis rate was about 80%.

Embodiment 3

[0039] A subcutaneous model of breast cancer in mice was constructed, and when the tumor diameter reached about 4-6 mm, group treatment began. They were divided into 4 groups with 5 mice in each group. The specific treatment plan is as follows:

[0040] Group 1: Local injection of phosphate buffered saline

[0041] Group 2: local injection of phosphate buffer, and ultrasonic treatment for 7 days.

[0042] Group 3: local injection of in vivo transfection reagent carrying MscL plasmid.

[0043] Group 4: local injection of in vivo transfection reagent carrying MscL plasmid, and ultrasonic treatment for 7 days. figure 1 Cell apoptosis after ultrasonic radiation for different time.

[0044] After the treatment, the body weight and tumor volume of the mice were measured every day. The tumor volume of the mice in the fourth group was significantly smaller than that of the other three groups, which can effectively inhibit the growth of the tumor.

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Abstract

The invention provides a method for controlling apoptosis and its application in inhibiting tumor growth. The method uses ultrasound to open a mechanically sensitive ion channel protein expressed by acell to cause apoptosis. Ultrasonic radiation to the cells allows the mechanically sensitive ion channel protein expressed by the cell to respond to the ultrasonic wave, and the ion channel opens togenerate ion flux, which causes the dynamic balance of the ion concentration inside and outside the cell to break, and causes the apoptosis of cells. By controlling the power, frequency and radiationtime of the ultrasonic wave, cells can be controlled to produce varying degrees of apoptosis. The mechanism of sound-activated apoptosis can effectively inhibit the growth of tumor cells.

Description

technical field [0001] The invention relates to the fields of biotechnology and medicine technology, in particular to a method for controlling cell apoptosis and its application in inhibiting tumor growth. Background technique [0002] Recent studies have shown that light-sensitive ion channel proteins can be opened under the conditions of corresponding wavelengths of light, allowing ions to flow inside and outside cells, and then regulating cells to produce a series of ion-mediated effects. However, there are some limitations in the use of light as a means of regulation, which prevent further applications, such as the limited penetration of light in the living body and the damage to cells caused by thermal effects. Ultrasound is a widely used, safe, and penetrative means of regulation. It is expected to achieve better results by using ultrasound instead of laser to regulate cells. And at present, the research on mechanosensitive ion channel proteins that can respond to ult...

Claims

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Application Information

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IPC IPC(8): C12N13/00
CPCC12N13/00
Inventor 常津王天歌王汉杰何天地张英英
Owner TIANJIN UNIV
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