CN110680858A

CN110680858A - Application of Senden-4 in anti-tumor

Info

Publication number: CN110680858A
Application number: CN201911203720.7A
Authority: CN
Inventors: 都日娜; 赵鹏伟; 高耀星; 乌日娜; 杨丽敏
Original assignee: International Mongolian Medical Hospital Of Inner Mongolia Autonomous Region (mongolian Medical Research Institute Of Inner Mongolia Autonomous Region)
Current assignee: International Mongolian Medical Hospital Of Inner Mongolia Autonomous Region (mongolian Medical Research Institute Of Inner Mongolia Autonomous Region)
Priority date: 2019-11-29
Filing date: 2019-11-29
Publication date: 2020-01-14

Abstract

The invention discloses application of Senden-4 in tumor resistance, and in vitro studies show that: DEN-4 was effective in inhibiting the proliferation of melanoma cells A375, and high concentrations of DEN-4 were effective in inducing apoptosis of A375 cells. In addition, Senden-4 was able to efficiently induce the level of autophagy at A375, which is shown by the change of autophagy marker LC3 from type I to type II with increasing drug concentration. In addition, Senden-4 can inhibit the phosphorylation activation of AKT and ERK signal pathways, and has obvious regulation effect on jnk and p 38.

Description

Application of Senden-4 in anti-tumor

Technical Field

The invention relates to the technical field of biomedicine, in particular to application of Senden-4 in tumor resistance.

Background

Mongolian medicine compound Senden-4 decoction is a common anti-rheumatism medicine in Mongolian medicine clinic and is recorded in the Negmgolian patent medicine Standard. The medicine is composed of 4 medicines, namely 15g of shinyleaf yellowhorn, 9g of szechwan chinaberry fruit, 3g of cape jasmine fruit and 3g of myrobalan according to the sequence of monarch, minister, assistant and guide, has the functions of clearing heat and drying yellow water, is used for treating gout, arthritis, edema and other diseases, and has fresh reports on the application of Senden-4 in resisting tumors.

Disclosure of Invention

The invention aims to provide the application of the Senden-4 in resisting tumors so as to expand the application of the Senden-4.

In vitro studies, Senden-4 was effective in inhibiting the proliferation of melanoma cells A375, and could inhibit the proliferation of A375 by apoptosis. We found that high concentrations of Senden-4 were effective in inducing apoptosis in A375 cells. Annexin/PI flow cytometry significantly increased early apoptotic cells and late apoptotic cells with increasing drug concentration. In addition, Senden-4 was able to efficiently induce the level of autophagy at A375, which is shown by the change of autophagy marker LC3 from type I to type II with increasing drug concentration. In addition, Senden-4 can inhibit the phosphorylation activation of AKT and ERK signal pathways, and has obvious regulation effect on jnk and p 38.

Drawings

FIG. 1 is a schematic diagram showing the detection of cell proliferation by CCK-8 method.

FIG. 2 is a schematic diagram showing the effect of the Senden-4 extract on melanoma tumor cell apoptosis.

FIG. 3 is a graph showing the effect of various concentrations of Senden-4 extract on MAPK, AKT, and caspase-3 pathway proteins.

FIG. 4 is a graph showing the induction of apoptosis and autophagy by different concentrations of Senden-4 extract.

Detailed Description

The present invention will be described in detail with reference to specific examples. The following examples will assist those skilled in the art in further understanding the invention, but are not intended to limit the invention in any way. It should be noted that variations and modifications can be made by persons skilled in the art without departing from the spirit of the invention. All falling within the scope of the present invention.

Examples of the experiments

Preparing a Senden-4 extracting solution:

250g of shinyleaf yellowhorn, 150g of szechwan chinaberry fruit, 50g of myrobalan and 50g of gardenia are taken, crushed into fine powder, sieved, weighed according to the proportion and mixed evenly to obtain the Senden-4 powder.

Taking 100g of Senden 4 powder, adding 1000ml of 95% ethanol, soaking for 24h, filtering, removing ethanol by rotary evaporation of the filtrate to obtain a concentrated solution, and adding pure water to the volume before concentration to obtain an ethanol extract solution. Taking 100g of Senden 4 powder, adding 1000ml of pure water, soaking for 24h, and filtering to obtain filtrate, namely water extraction solution. Mixing the ethanol extract solution and the water extract solution in equal volume to obtain an extractive solution with gallic acid (effective component) concentration of about 300 μ g/ml.

The extract was adjusted to the corresponding concentration by adding distilled water.

CCK-8 method for detecting cell proliferation

A375 cells were treated with drug at concentrations of 0mg/ml, 5mg/ml, 1mg/ml, 200ug/ml, 50ug/ml, 10ug/ml, 2ug/ml, 500ng/ml, 100ng/ml, 20ng/ml, 5ng/ml, 1ng/ml for 72h, added to CCK8, incubated for 1h at 37 ℃ incubator and absorbance was measured at 450 nm. As a result, as shown in FIG. 1, the absorbance of the cells reached the lowest at 1mg/ml, and the absorbance at 200ug/ml was about half that of the normal cells.

Detecting apoptosis of A375 cells treated by Senden-4 extract with different concentrations by flow cytometry

A375 cells were treated with 1mg/ml, 200ug/ml, 50ug/ml and 0ug/ml of Senden-4 extract for 24h, respectively. The results showed that the proportion of apoptosis gradually increased with increasing drug concentration, as shown in figure 2.

Effect of different concentrations of drug on MAPK, AKT, and caspase-3 pathway proteins

A375 cells were treated with different concentrations of senden-4 extract: 2mg/ml, 1mg/ml, 0.5mg/ml, 0.25mg/ml, 0.125mg/ml, 62.5ug/ml and 0ug/ml (Senden-4 extract concentration) for 72 hours. Cellular proteins were extracted and detected using a western blot. The results show that: as shown in fig. 3, as the concentration of the senden-4 extract increased, the phosphorylated erk and phosphorylated akt signals gradually decreased, indicating that cell proliferation was inhibited.

As shown in FIG. 4, the low concentration of the Senden-4 extract significantly induced apoptosis and autophagy. Expression of type II LC3B was seen by treating A375 cells with 62.5ug/ml of Senden-4 extract.

In summary, the senden-4 extract can significantly inhibit the proliferation of a375 cells, and can inhibit the proliferation of a375 cells by apoptosis, and the high concentration of the senden-4 extract can effectively induce apoptosis of a375 cells, and Annexin/PI flow cytometry significantly increases early apoptotic cells and late apoptotic cells as the concentration of the drug increases. Annexin/PI flow cytometry significantly increased early apoptotic cells and late apoptotic cells with increasing drug concentration. In addition, the Senden-4 extract was effective in inducing autophagy at A375, which is shown by the change in autophagy marker LC3 from type I to type II with increasing drug concentration. In addition, the Senden-4 extract can inhibit the phosphorylation activation of AKT and ERK signal pathways, and has obvious regulation effect on jnk and p 38.

The foregoing description of specific embodiments of the present invention has been presented. It is to be understood that the present invention is not limited to the specific embodiments described above, and that various changes or modifications may be made by one skilled in the art within the scope of the appended claims without departing from the spirit of the invention. The embodiments and features of the embodiments of the present application may be combined with each other arbitrarily without conflict.

Claims

1. The application of Senden-4 in resisting tumors is characterized in that: senden-4 inhibits the phosphorylation activation of the AKT and ERK signaling pathways.

2. The use of claim 1, wherein: the Senden-4 can effectively inhibit the proliferation of melanoma cells A375.

3. Use according to claim 2, characterized in that: senden-4 can inhibit the proliferation of A375 by programmed death.

4. Use according to claim 2, characterized in that: senden-4, 1mg/ml, was effective in inducing apoptosis in A375 cells.

5. Use according to claim 2, characterized in that: senden-4 was effective in inducing autophagy levels of A375.