Application of attenuated salmonella in preparation of medicine for treating lung cancer pleural effusion
A Salmonella and pleural effusion technology, applied in the application field of attenuated Salmonella in the preparation of pleural effusion drugs for the treatment of lung cancer, to achieve the effect of increasing survival time, improving pleural effusion, and improving quality of life
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Embodiment 1
[0046] Construction of mouse lung cancer pleural effusion model
[0047] Experimental cells:
[0048] LLC cells were donated by the School of Life Sciences, Nanjing University.
[0049] Experimental animals:
[0050] Wild-type C57 mice (C57BL / 6J wild-type mice): SPF grade, 6-8 weeks old, mass 16-20 g, purchased from Changzhou Cavens Laboratory Animal Co., Ltd.
[0051] Main experimental equipment:
[0052] Bacterial culture shaker was purchased from Shanghai Zhicheng Analytical Instrument Manufacturing Co., Ltd.; pipette was purchased from Glison Company; desktop common centrifuge, desktop low-temperature refrigerated centrifuge and biological spectrophotometer were purchased from Eppendorf Company; PH instrument was purchased from Sartorius Company; Nucleic acid electrophoresis and protein vertical electrophoresis system were purchased from Bio-Rad Company of the United States; PCR instrument was purchased from Takara Company of Japan; autoclave was purchased from Zealway ...
Embodiment 2
[0060] In vitro growth curve determination of attenuated Salmonella VNP20009
[0061] The VNP20009 stored at -80°C was inoculated into LB liquid medium, recovered and cultured overnight in a bacterial incubator at 37°C, and then transferred to fresh LB at 1:100 the next day and cultured to an OD of 0.6-0.8. Take the same amount of VNP20009 strain. Transfer to an equal amount of fresh LB and place it in a 37°C bacterial incubator for 7 hours, sample once every 0.5 hours from the time the bacteria were transferred to LB (ie 0h), and use a biospectrophotometer to measure the OD600 nm of the bacteria value, plot the bacterial growth curve ( figure 2 ).
[0062] Attenuated Salmonella VNP20009 in the treatment of pleural effusion in mice with lung cancer and pleural effusion
[0063] Take lung cancer pleural effusion model mice, use isoflurane anesthesia machine to lightly anesthetize, use forceps to pick up the skin of the 5-6 intercostal space on the left side of the sternum of...
Embodiment 3
[0069] Construction of mouse lung cancer pleural effusion model
[0070] (1) In vitro cell culture
[0071] The LLC cells were taken out from a liquid nitrogen tank or -80°C refrigerator, thawed at 37°C for 1 min to recover the cells, centrifuged at 1000 rpm / min for 3 min, and cultured in 90% DMEN high-glucose medium and 10% fetal bovine serum in a 37°C 5% carbon dioxide incubator. After the cells entered the exponential growth phase and covered the bottom of the culture flask, the iron-walled cells were digested with trypsin and washed out with PBS. The total number of cells using the hemocytometer technique was used, and the proportion of viable cells was calculated using the trypan blue detection kit. When the proportion of viable cells reached more than 90%, the LLC cell suspension was diluted to 1x10 with normal saline. 6 spare, figure 1 .
[0072] (2) Construction of mouse lung cancer pleural effusion model
[0073] Take 30 6-8 week old C57BL / 6J wild-type mice, half ...
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