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Humanized drug-resistant GH-type pituitary tumor cell strain and establishment method and application

A technology for pituitary tumors and cell lines, applied in the fields of biotechnology and microbial animal cell lines, can solve the problems of pituitary adenoma cell line establishment and culture difficulties that have not yet been seen, and achieve significant development potential and clinical application value

Inactive Publication Date: 2014-12-17
AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In view of the fact that most human pituitary tumors are benign, their cultivation is more difficult than other tumor cells. So far, there has been no report on the establishment of human pituitary adenoma cell lines.

Method used

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  • Humanized drug-resistant GH-type pituitary tumor cell strain and establishment method and application
  • Humanized drug-resistant GH-type pituitary tumor cell strain and establishment method and application
  • Humanized drug-resistant GH-type pituitary tumor cell strain and establishment method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 Obtaining and Primary Culture of Pituitary Adenoma Cells

[0026] (1) Source of specimen

[0027] Cao ××, male, 17 years old, a patient from the Neurosurgery Hospital of Shanghai Huashan Hospital, obtained the consent of the patient before the operation and signed the informed consent form. The specimen of pituitary adenoma was excised during the operation, and the postoperative pathological diagnosis was pituitary GH adenoma (such as Figure 4 shown);

[0028] (2) method

[0029] ①The surgically resected pituitary adenoma tissue was put into serum-free DMEM culture medium under aseptic operation, and sent to the laboratory immediately;

[0030] ② Rinse 2-3 times with sterile PBS in the ultra-clean bench to remove connective tissue, necrotic tissue and blood clots, cut the specimen into 1mm3 tissue pieces with ophthalmic scissors, add 5-6 times the amount of 2.5g / L trypsin ( Preheat to 37°C);

[0031] ③ Digest at 37°C for 30-50 minutes, and disperse on...

Embodiment 2

[0034] Example 2 Subculture of pituitary adenoma cells

[0035] (1) Source of specimen

[0036] Primary tumor cells in the above-mentioned embodiment 1

[0037] (2) method

[0038] ①Cultivate tumor cells to the fifth passage; blow the cell suspension with a pipette to blow it down from the bottom of the dish, make a single cell suspension and transfer it to a 15 ml centrifuge tube; centrifuge at 100×g for 5 min; Discard the supernatant, add 5 ml of complete culture medium to resuspend to make a single cell suspension; divide the cells into new culture dishes at a ratio of 1:2, shake well and put them in the incubator for culture;

[0039]②From the sixth generation, subculture once every 3 days, each division ratio is 1:2, and the culture medium uses D / F culture medium; the cells grow stably and continue to secrete GH. After the 20th generation, change to DMEM culture medium, the cells go through a 7-day slow growth period, and then continue to pass at a rate of one gene...

Embodiment 3

[0040] Example 3 Biological properties of huashanc-GH

[0041] The biological characteristics of the huashanc-GH cell line are tested and verified by conventional methods, including steps:

[0042] (1) Experimental method

[0043] ① Morphology

[0044] a. General shape

[0045] One week after cell inoculation, observe the growth morphology of the population cells;

[0046] b. Light mirror

[0047] Observe the morphological structure and growth characteristics of the cultured living cells under an inverted phase-contrast microscope;

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Abstract

The invention belongs to the field of biotechnology and microbe and animal cell lines and specifically relates to a humanized drug-resistant GH-type pituitary tumor cell strain huashanc-CH, especially to a first international humanized drug-resistant GH-type pituitary tumor cell strain huashanc-CH and its establishment method and application. Through isolated culture of pituitary tumor cells from a patient with clinical drug-resistant GH-type pituitary tumor, the humanized drug-resistant GH-type pituitary tumor cell strain huashanc-CH is obtained. The cell strain is passaged for 50 times for the first time and is mostly round or elliptical, cytoplasm is abundant, and nucleus is large and round. It shows through experimental results that the cell strain can be passaged stably, can secrete growth hormones continuously, is drug-resistant to somatostatin analogue, can be used to provide a necessary cell test tool for researches on pathogenic and drug-resistance mechanisms of pituitary tumor, research and development and screening of new drugs and new biotherapy and detection of related bio-engineering products and the like, and has exploring potential and application value.

Description

technical field [0001] The invention belongs to the fields of biotechnology and microbial animal cell lines, and relates to human tumor cells, in particular to a human-derived drug-resistant GH-type pituitary tumor cell line (huashanc-GH), especially to the world's first human-derived drug-resistant GH-type Pituitary tumor cell line (huashanc-GH) and its establishment method and use. Background technique [0002] Studies have shown that pituitary tumors are a group of tumors that arise from residual cells of the anterior and posterior pituitary glands and craniopharynx, accounting for about 10-15% of intracranial tumors; among them, growth hormone-type pituitary tumors are caused by growth hormone (Growth Hormone , GH) excessive secretion causes acromegaly (Acromegaly). According to reports, terminal hypertrophy is a serious neuroendocrine disease, which is mediated by the abnormal increase of insulin-like growth factor-I (Insulin-like Growth Factor-1, IGF-1) caused by exce...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/09A61K35/30A61P35/00C12Q1/04
Inventor 赵曜
Owner AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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