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Method for screening medicine for restraining tumor proliferation and promoting cell apoptosis by disturbing nidogen expression

A tumor proliferation and tumor cell line technology, which is used in the field of screening drugs that inhibit tumor proliferation and promote cell apoptosis

Inactive Publication Date: 2010-06-09
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there is no research report on the effect of nestin on tumor cell proliferation, apoptosis and unknown functions

Method used

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  • Method for screening medicine for restraining tumor proliferation and promoting cell apoptosis by disturbing nidogen expression
  • Method for screening medicine for restraining tumor proliferation and promoting cell apoptosis by disturbing nidogen expression
  • Method for screening medicine for restraining tumor proliferation and promoting cell apoptosis by disturbing nidogen expression

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1: Immunohistochemical detection of nestin expression in tumors of different malignancies

[0028] The pathological tissue sections used in this experiment are mainly from the Department of Pathology of the First Affiliated Hospital of Sun Yat-sen University and the Department of Pathology of the Cancer Prevention and Control Center of Sun Yat-sen University, including: highly and poorly differentiated gastrointestinal stromal tumors; highly and poorly differentiated angiosarcoma, cavernous vessels Tumor; Well-differentiated, moderately differentiated and poorly differentiated pancreatic adenocarcinoma; Well-differentiated, moderately differentiated and poorly differentiated hepatocellular carcinoma; Well-differentiated and poorly differentiated intrahepatic cholangiocarcinoma; Embryonal rhabdomyosarcoma, alveolar rhabdomyosarcoma, polymorphic Sexual rhabdomyosarcoma, malignant melanocytoma, testicular stromal cell tumor, retinoblastoma, benign and malignant isle...

Embodiment 2

[0040] Example 2: Establishment of the culture of mouse melanocytoma B16 and the monitoring method for nestin expression

[0041] The animals used in this experiment were clean-grade C57BL / 6 mice, 6-8 weeks old, male or female, purchased from the Medical Experimental Animal Center of Sun Yat-sen University.

[0042] main reagent

[0043] Fluorescent immunochemical reagents:

[0044] Primary antibody: mouse anti-rat / mouse Nestin antibody (Nestin), Chemicon

[0045] Mouse anti-BrdU antibody, DSHB company

[0046] Mouse anti-mouse PCNA antibody, Cell Signaling Technology

[0047] Rabbit anti-mouse Ki67 antibody, Abcam

[0048] Secondary antibody: Cy3-conjugated goat anti-mouse IgG (H+L), Jackson ImmunoResearch

[0049] R-PE conjugated goat anti-rabbit IgG (H+L), SouthernBiotech

[0050] Serum: goat serum blocking solution (ready-to-use), Wuhan Boster Bioengineering Company

[0051] S-P Immunohistochemical Kit (Immunohistochemical Kit), Maixin Company

[0052] Cell Culture...

Embodiment 3

[0098] Example 3: Effect of short-term down-regulation of nestin expression on proliferation and apoptosis of mouse melanoma B16

[0099] Three siRNA sequences were designed, as follows:

[0100] siNM_016701_001

[0101] Target sequence: AAAGGAGAATTGCAATTCA

[0102] Sense strand (5′-3′): AAAGGAGAAUUGCAAUUCA dTdT

[0103] Antisense strand (3′-5′): dTdT UUUCCUCUUAACGUUAAGU

[0104] siNM_016701_002

[0105] Target sequence: CAAAGCCTCTTAGAAATGA

[0106] Sense strand (5′-3′): CAAAGCCUCUUAGAAAUGA dTdT

[0107] Antisense strand (3′-5′): dTdT GUUUCGGAGAAUCUUUACU

[0108] siNM_016701_003

[0109] Target sequence: GTGAGACTCTGGAATGCAA

[0110] Sense strand (5′-3′): GUGAGACUCUGGAAUGCAA dTdT

[0111] Antisense strand (3′-5′): dTdT CACUCUGAGACCUUACGUU

[0112] negative control

[0113] Target sequence: TTCTCCGAACGTGTCACGT

[0114] Sense strand (5′-3′): UUCUCCGAACGUGUCACGU dTdT

[0115] Antisense strand (3′-5′): dTdT AAGAGGCUUGCACAGUGCA

[0116] Experimental method: Digest p5 m...

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Abstract

The invention relates to a method for screening a medicine for restraining tumor proliferation and promoting cell apoptosis by disturbing nidogen expression. The method comprises the following steps of: A. providing a tumor specific cell line and condition of culture; B. identifying the expression of the expression nidogen of the tumor cell line; C. adding a medicine to be screened; and D. culturing for 48 hours or above; detecting the expression of the nidogen; and comparing the detection result with the result of the step B; if the situation of down-regulation nidogen expression is generated, the medicine to be screened becomes the precursor medicine for restraining tumor proliferation and / or promoting cell apoptosis. The invention obtains direct evidence of the relation between the nidogen and the growth, proliferation, apoptosis, migration and other vital movements of tumor cell and proves that the establishment of the method for screening the medicine for restraining tumor proliferation and promoting cell apoptosis by disturbing the nidogen expression is feasible. The invention provides the method for screening the medicine for restraining tumor proliferation and promoting cell apoptosis for the first time.

Description

technical field [0001] The invention belongs to the field of molecular biology, and relates to a method for screening drugs for inhibiting tumor growth and promoting cell apoptosis, in particular to a method for screening drugs for inhibiting tumor growth and promoting cell apoptosis by interfering with the expression of nestin . Background technique [0002] Cytoskeletal proteins play important roles in cell motility, division, information transfer, energy conversion, metabolic regulation, and fibroblast morphology. The cytoskeleton can be divided into microtubules, microfilaments, intermediate fibers or intermediate filaments, and microbeams according to the size of the fiber diameter. The distribution of intermediate fibers is strictly tissue specific. This point has been applied in the clinical differential diagnosis of tumors to identify the tissue source of tumor cells. Nestin (nestin) is a VI intermediate filament protein. In mammals, nestin was first found expres...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/48G01N33/53C12Q1/02
Inventor 项鹏杨旭辉夏添
Owner SUN YAT SEN UNIV