Establishment and applications of human pituitary adenoma cell line

A pituitary adenoma and cell line technology, which is applied in the field of preparing human pituitary adenoma cell lines, can solve the problems of excessive growth of fibroblasts and the influence of successfully cultured cell lines, and achieves fast growth rate, stable passage and high tumor formation rate. Effect

Inactive Publication Date: 2017-12-12
任远 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

It is worth noting that fibroblast overgrowth is inevitable in pituitary ad

Method used

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  • Establishment and applications of human pituitary adenoma cell line
  • Establishment and applications of human pituitary adenoma cell line
  • Establishment and applications of human pituitary adenoma cell line

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Embodiment 1: the establishment of human pituitary adenoma cell line and animal model

[0048] Immunodeficient mouse SCID Beige CB17.Cg-PrkdcscidLystbg / Crl (4 weeks, male, 20 g) was purchased from Weitong Lihua Biotechnology Co., Ltd.

[0049] 1.1 Method: Through transsphenoidal single nostril pituitary adenoma resection, non-functional pituitary adenoma tumor tissue was obtained, and then placed in a 15ml sterile test tube containing 1% double-antibody (penicillin and streptomycin) DMEM culture solution , try to transport to the cell culture room at 4°C. In the cell sterile culture platform, use PBS solution containing 1% double antibody (penicillin and streptomycin) to wash the tissue pieces three times, then use sterile surgical scissors to carefully cut the tissue into small pieces in a disposable sterile culture dish. Small pieces with a diameter of about 2 mm were washed with PBS solution, and then transferred to another sterile petri dish, and the PBS solution w...

Embodiment 2

[0061] Example 2: Confirmation of Human Pituitary Adenoma Cell Lines

[0062] 5×10 of the human nonfunctional pituitary adenoma cell line HNFPA obtained according to Example 1 6 Cells were seeded in 10ml DMEM medium containing 10% FBS, 1U / 100ml insulin, 0.1mg / 100ml heparin and 0.1mg / 100ml non-essential amino acids in 5% CO 2 , Culture at 37°C. When the cells form a single cell layer attached to the wall, discard the culture medium, wash twice with warm PBS, then add 0.25% trypsin digestion solution (including EDTA) to submerge the monolayer, suck the digestion solution quickly after 3 minutes, add 3 times the volume of fresh DMEM medium containing 10% FBS was neutralized and pipetted repeatedly, and inoculated with fresh DMEM medium containing 10% FBS, 1U / 100ml insulin, 0.1mg / 100ml heparin and 0.1 mg / 100ml of non-essential amino acids in DMEM medium in 5% CO 2 , 37 ℃ continue to culture passage.

[0063] The inoculated cells can grow adherently within 1 hour. Depending on ...

Embodiment 3

[0075] Example 3: Subcutaneous transplantation of non-functional pituitary adenoma model intervention experiment

[0076] Intervention experiment was carried out by using the patient's own non-functional pituitary adenoma severe immunodeficiency mouse SCID BEIGE tissue block subcutaneous xenograft model prepared in Example 1. Severe immunodeficiency mouse SCID BEIGE was purchased from Weitong Lihua Biotechnology Co., Ltd. The patient has signed the informed consent.

[0077] The experimental steps are as follows:

[0078] 1. SCID BEIGE was anesthetized by intraperitoneal injection of 2% pentobarbital sodium (2mg / kg), and the fresh tissue with nonfunctional adenoma cells was cut into 3×3×3mm 3 The tissue block was transplanted subcutaneously in the left axilla of SCID BEIGE, and the SCID BEIGE xenograft tumor model of severe immunodeficiency mouse SCID BEIGE was established corresponding to the surgical tissue block of non-functioning pituitary adenoma. History and image ana...

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Abstract

The invention relates to a method for preparing a human pituitary adenoma cell line, the human pituitary adenoma cell line obtained through the method, and applications of the human pituitary adenoma cell line in medicine research and development. The method comprises the following steps: (a) transplanting the human pituitary adenoma to a non-human animal suffering from immune deficiency; (b) when the weight of the transplanted pituitary adenoma achieves 15% or above of the body weight of the non-human animal or when the size of the transplanted tumor does not increase, taking out the pituitary adenoma, and transplanting the pituitary adenoma into the new non-human animal suffering from immune deficiency; (c) repeating the step (b), when the Ki67 positive expression ratio of the pituitary adenoma cells is 20% or above, taking out the pituitary adenoma; and (d) culturing the pituitary adenoma cells which are taken out, thus obtaining the pituitary adenoma cell line.

Description

technical field [0001] The invention relates to a method for preparing human pituitary adenoma cell line, the human pituitary adenoma cell line obtained by the method and its application in medicine research and development. Background technique [0002] Pituitary adenoma is one of the common clinical benign intracranial tumors, accounting for 10%-15% of intracranial tumors. In recent years, the incidence of pituitary tumors has been on the rise, and pituitary tumors usually occur in young adults. Non-function pituitary adenoma (NFPA) with abnormal hormone secretion often has severe local space-occupying symptoms, often leading to blindness and other symptoms. Hypersecretion of hormones by pituitary tumors can lead to increased levels of prolactin (PRL), adrenocorticotropic hormone (ACTH) and other hormones in patients, resulting in moon face, central obesity, acne, increased hair growth, and high blood pressure in patients , Secondary diabetes, osteoporosis and other symp...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12N15/87A01K67/027
CPCA01K67/0271A01K2207/12A01K2227/105A01K2267/0331C12N5/0693
Inventor 任远王任直
Owner 任远
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