Establishment method and application of paraquat-resistant cell line of human lung cancer

A technology of paraquat and cell lines, applied in the field of cell engineering, to achieve the effect of improving the success rate of induction and obviously resisting the toxicity of PQ

Active Publication Date: 2019-08-13
THE FIRST AFFILIATED HOSPITAL OF WENZHOU MEDICAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Through searching, it is found that there is no literature report on the establishment of human lung cancer drug-resistant paraquat cell line by using the human lung cancer cell line A549 as the induction object at present.

Method used

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  • Establishment method and application of paraquat-resistant cell line of human lung cancer
  • Establishment method and application of paraquat-resistant cell line of human lung cancer
  • Establishment method and application of paraquat-resistant cell line of human lung cancer

Examples

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Effect test

Embodiment 1

[0028] Example 1 Induction and establishment of paraquat-resistant cell lines of human lung cancer

[0029] The human lung cancer paraquat-resistant cell line A549 / PQ was established according to the following steps: (1) A549 cells were routinely cultured in 1640 medium containing 10% fetal bovine serum at 37°C and 5% CO 2 . Cells in the logarithmic growth phase were used for experiments each time. (2) Establish cell drug-resistant strains by using PQ induction method with increasing concentration and periodic exposure: the concentration of PQ in the medium first starts from 100 μmol / L, and after adding the drug for 24 hours, change the medium to remove the drug, and wait for the cells to re-grow to the logarithmic phase Afterwards, the same concentration of PQ was added again for stimulation, and this was repeated 3 times. Then extend the exposure time to 48h (100μmol / L exposure for 48h), change the medium to remove the drug, and after the cells re-grow to the logarithmic p...

Embodiment 2

[0031] Example 2 Morphological observation of human lung cancer paraquat-resistant cell line A549 / PQ and human lung cancer A549 cells

[0032] The human lung cancer cell lines A549 and A549 / PQ cells in the logarithmic growth phase were routinely digested and passaged, and the cell morphology was observed with a Leica optical microscope 24 hours after the cells were completely adhered to the wall, and the photos were taken with a Nikon camera. The results are as follows: figure 1 shown.

Embodiment 3

[0033] Example 3 Cell Counting Method Drawing Human Lung Cancer Paraquat-resistant Cell Line A549 / PQ and Human Lung Cancer Cell Line A549 Cell Growth Curve and Determination of Doubling Time

[0034] Take 50,000 parental A549 cells in mid-logarithmic growth and good growth conditions, and human lung cancer paraquat-resistant cell line A549 / PQ cells to make cell suspensions and inoculate them into culture flasks, 21 flasks of each cell. After 24 hours, the cell counting was started, and the counting was continued for seven days, with three flasks of cells per day, and the cell growth curve was drawn according to the counting results. Then calculate the doubling time (DT) according to the formula, DT=t×lg2 / (lgNt-lgN0), t is the cell culture time, N0 is the average number of cells measured after inoculation 24h, and Nt is the average number of cells after culturing th value, the result is as figure 2 and 3 shown.

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Abstract

A human lung cancer cell strain A549 is adopted as an inducing object, an in-vitro inducing method for performing periodic exposure and gradually increasing the concentration of paraquat is adopted, and a paraquat-resistant cell strain A549 / PQ is established. The cell strain is preserved in China General Microbiological Culture Collection Center at June 16th 2015, and the preservation number is CGMCC NO.10892. The cell strain has the typical paraquat-resistant characteristic, and the biological characteristic provides a research model for further performing research on reversing of toxicity of paraquat, extracting an evaluation marker of paraquat poisoning and exploring a new anti-oxidant way, and has high scientific research value and production and application value.

Description

technical field [0001] The invention belongs to the technical field of cell engineering and relates to a human cell line, in particular to a paraquat-resistant human lung cancer cell line and its establishment method and application. Background technique [0002] Paraquat (PQ) is a high-efficiency herbicide widely used in the world. It is highly toxic to the human body. It can cause poisoning through absorption in the digestive tract, skin, respiratory tract, and veins. There is no effective treatment for it in clinical practice. Measures, the mortality rate of patients as high as 40 to 80%. The production and consumption of PQ in my country rank first in the world. In recent years, the incidence of PQ poisoning in China has increased exponentially, and the prevention and control situation is very severe. Although my country began to gradually ban the production and sales of PQ water in June 2015, other dosage forms will continue to be produced and used. Therefore, paraqu...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/09C12N1/36
Inventor 卢中秋洪广亮谭佳平刘佳明丁颖威赵光举
Owner THE FIRST AFFILIATED HOSPITAL OF WENZHOU MEDICAL UNIV
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