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Human renal vascular smooth muscle lipoma primary cell, human renal vascular smooth muscle lipoma progeny cell and applications thereof

A technology of primary cells and renal blood vessels, applied in the field of cell biology, can solve the problems of easy loss of tumor cell heterogeneity and tumor cell characteristics, and achieve the effect of stable cell characteristics

Active Publication Date: 2021-09-21
武汉赛尔朗灵科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there are only a handful of tumor cell lines used in basic research and drug sensitivity testing in the world, and the tumor cell lines have serious defects, that is, the heterogeneity of tumor cells and the stability of tumor cells in vivo are easily lost during in vitro culture. The primary cultured tumor cells have become an ideal in vitro experimental model for researchers because they retain their original genetic characteristics, maintain good correlation with the clinic, and can more truly reflect the internal characteristics of the body.
No primary human renal angiomyolipoma cells are currently available to researchers

Method used

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  • Human renal vascular smooth muscle lipoma primary cell, human renal vascular smooth muscle lipoma progeny cell and applications thereof
  • Human renal vascular smooth muscle lipoma primary cell, human renal vascular smooth muscle lipoma progeny cell and applications thereof
  • Human renal vascular smooth muscle lipoma primary cell, human renal vascular smooth muscle lipoma progeny cell and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Primary isolation and culture of XHRAML-06

[0042] (1) Fresh human renal angiomyolipoma resection clinical specimens were obtained from Wuhan Union Medical College Hospital through the hospital ethics committee, with the consent of the patient or the patient's guardian and after signing the informed consent.

[0043] (2) Immediately put the resected specimen into the pre-cooled sterile tissue preservation solution (containing 1000U / ml penicillin, 1000ug / ml streptomycin sulfate, 2.5ug / ml amphotericin and 50ug / ml gentamycin) Into the collection tube, and immediately put into a 4 ℃ sample transport box, transported to the laboratory within 4 hours for cell separation.

[0044] (3) Primary isolation and culture: Obtain the tissue in a biological safety cabinet, wash it once with absolute ethanol, and wash it twice with 1xPBS (pH7.2-7.4), remove blood vessels, fat and For the necrotic tissue part, cut the tissue to a minced shape with dissecting scissors. Add 20...

Embodiment 2

[0046] Example 2 Subculture of primary cells of human renal angiomyolipoma

[0047] (1) When the abundance of cells cultured in T25 culture flask reaches 80%, rinse the cells twice with 1xPBS (pH7.2-7.4), add 1ml of 0.05% trypsin-EDTA to digest the monolayer cells for 2-3min.

[0048] (2) Add 2 ml of PBS containing 10% FBS to stop the digestion.

[0049] (3) Centrifuge at 1000rpm for 4min, remove the supernatant, collect the cell pellet, resuspend with 1ml CORT medium, replenish the medium, and put it into a T25 culture bottle for cultivation according to the ratio of 1 to 2.

[0050] The human renal angiomyolipoma primary cells subcultured according to the above-mentioned method, the cell growth curve obtained by culturing is as follows: figure 2 , continuous passage for 43 days, the cell multiplication multiple reached 34, and the primary cells of human renal angiomyolipoma of the present invention can still maintain a proliferative state and grow normally.

Embodiment 3

[0051] Example 3 Marker Expression Identification of Human Renal Angiomyolipoma Primary Cells

[0052] After the human renal angiomyolipoma primary cell XHRAML-06 obtained above was cultured, the specific marker expression of the cell was identified, and the specific steps were as follows:

[0053] (1) In the culture plate, soak the slide with the cells climbed in PBS for 3 times, each time for 3 minutes;

[0054] (2) Fix slides with 4% paraformaldehyde for 15 minutes, soak and wash slides with PBS 3 times, 3 minutes each time;

[0055] (3) Permeabilize with 0.5% Triton X-100 (prepared in PBS) at room temperature for 20 minutes (this step is omitted for antigens expressed on the cell membrane);

[0056] (4) PBS soaked slides 3 times, 3 minutes each time, blotted the PBS with absorbent paper, added normal goat serum dropwise on the slides, sealed at room temperature for 30 minutes;

[0057] (5) Absorb the blocking solution with absorbent paper, without washing, add a sufficie...

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Abstract

The invention discloses human renal vascular smooth muscle lipoma primary cell, human renal vascular smooth muscle lipoma progeny cell and an applications thereof The cell is named as a human renal vascular smooth muscle lipoma primary cell XHRAML-06, and the preservation number is C202082. The donor of the primary cell is a clinical renal vascular smooth muscle lipoma patient. The primary cell has in-vitro soft agar tumorigenesis ability and nude mouse subcutaneous tumorigenesis ability, and shows different drug toxic reactions to rapamycin, lobaplatin and carboplatin. The cell provided by the invention can be used for analyzing the pathogenesis of the human renal vascular smooth muscle lipoma in vitro and in vivo and detecting related characteristics such as drug sensitivity in vitro, and an experimental material closer to clinical tumor biological characteristics is provided for the research of the human renal vascular smooth muscle lipoma.

Description

technical field [0001] The invention belongs to the technical field of cell biology, and in particular relates to a primary cell of human renal angiomyolipoma, a progeny cell and an application thereof. Background technique [0002] According to the 2016 WHO classification of renal tumors, renal angiomyolipoma (RAML) is classified as a mesenchymal tumor that mainly occurs in adults. Histologically, there are two types of RAML, one is classical RAML and the other is epithelioid RAML. Classical RAML is a benign mesenchymal neoplasm composed of adipose tissue, spindle-shaped and epithelioid smooth muscle cells, and thick-walled vessels. Epithelioid RAML is characterized by epithelioid cell proliferation on the basis of classical RAML structure, showing infiltrating and destructive growth, and is a mesenchymal tumor with malignant potential. RAML accounts for 8.7%-29.4% of renal tumors. Because it can lead to common complications of spontaneous rupture and bleeding, it can ca...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12N5/071C12Q1/02C12R1/91
CPCC12N5/0693C12N5/0691G01N33/5011G01N33/5061C12N2503/02G01N2500/10
Inventor 刘红亚章小平肖文王前进刘霞李继新
Owner 武汉赛尔朗灵科技有限公司
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