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Application of Nestin gene and protein in treatment of organ fibrosis

An organ fibrosis and gene technology, applied in the preparation of organ fibrosis treatment drugs, the application of Nestin gene and protein in the treatment of organ fibrosis, can solve problems such as poor prognosis and affect the quality of life of patients, and achieve slowing down. The effect of progress, good application value and prospect

Active Publication Date: 2021-03-30
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006]In addition, although some anti-fibrotic drugs have been discovered recently, they have poor prognosis, can only slow down the progress of the disease and have serious side effects, thereby affecting the quality of life of patients

Method used

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  • Application of Nestin gene and protein in treatment of organ fibrosis
  • Application of Nestin gene and protein in treatment of organ fibrosis
  • Application of Nestin gene and protein in treatment of organ fibrosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Example 1 Relationship between liver fibrosis and Nestin expression in mice

[0055] 1. Construction of a mouse model of liver fibrosis

[0056] 8-14-week-old male C57BL / 6 mice (Nanjing University Institute of Model Animals) were treated with the following regimens: Oil, CCl4 2w, CCl4 4w, CCl 4 ; Normal diet, DDC 1w, DDC 4w. Mice were injected intraperitoneally with 5 μl / g of 20% CCl4 (Sigma-Aldrich, 289116) (diluted in corn oil (Sigma-Aldrich, 23-0230)) or the same volume of corn oil, twice a week; or regular feeding containing 0.1% DDC (Sigma-Aldrich, 137030) conventional mouse chow, Normaldiet group did not do any treatment. On the 28th day, the mouse liver tissue was taken for the experiment.

[0057] 2. Histopathological evaluation of mouse liver fibrosis model

[0058] After mice were euthanized, mouse liver tissue was perfused with 4% paraformaldehyde. After tissue sections were made, PSR, IHC (rabbit anti-Nestin: Millipore, ABD69; mouse anti-α-SMA: Abcam, a...

Embodiment 2

[0067] Example 2 The effect of interfering with Nestin on the fibrosis of mouse primary HSCs cell line and LX2 cell line

[0068] 1. Isolation and culture of mouse primary HSCs and culture of LX2 cell line

[0069] Mouse livers were digested by reverse stepwise perfusion using a solution containing Pronase (Sigma-Aldrich, 11459643001) and Collagenase (Sigma-Aldrich, 11213865001). The cell suspension was centrifuged at 50 g and the supernatant containing non-parenchymal hepatocytes was collected. HSCs were purified from non-parenchymal hepatocytes by density gradient centrifugation and incubated at 37°C, 5% CO. 2 cultured in DMEM containing 10% fetal bovine serum (FBS).

[0070] LX2 cell line (ATCC) at 37°C, 5% CO 2 cultured in DMEM containing 10% fetal bovine serum (FBS).

[0071] 2. Construction of cell lines that stably interfere with Nestin

[0072] Construct Nestin lentivirus interference vector, and construct a stable Nestin interference cell line by viral packaging ...

Embodiment 3

[0086] Example 3 Effects of AAV6-ShNestin on liver fibrosis in mice

[0087] 1. Adeno-associated virus 6 (AAV6) and CCl4 / DDC treatment

[0088]Adeno-associated virus serotype 6 (AAV6-ShNES) expressing ShNestin under the CMV promoter was purchased from Hanheng Biotechnology Co., Ltd. (Shanghai, China). Twenty-four 8-14-week-old male C57BL / 6 mice (Nanjing University Institute of Model Animals) were divided into 4 groups and were treated with the following regimens: Oil+AAV6-shControl (n=6), CCl4+AAV6-shControl ( n=6), CCl4+AAV6-ShNes#1 (n=6), CCl4+AAV6-ShNes#2 (n=6); Normal diet+AAV6-shControl (n=6), DDC+AAV6-shControl (n=6) =6), DDC+AAV6-ShNes#1 (n=6), DDC+AAV6-ShNes#2 (n=6). Mice were injected intraperitoneally with 5 μl / g of 20% CCl4 (Sigma-Aldrich, 289116) (diluted in corn oil (Sigma-Aldrich, 23-0230)) or the same volume of corn oil, twice a week; or regular feeding containing 0.1% Conventional mouse chow for DDC (Sigma-Aldrich, 137030). After the mice were anesthetized ...

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Abstract

The invention discloses an application of a Nestin gene and protein in treating organ fibrosis. The researches show that the Nestin can be used as an important marker of organ fibrosis, meanwhile, theNestin has a remarkable treatment effect on the organ fibrosis by interfering / inhibiting the Nestin, and fibrosis related indexes such as collagen content are reduced by interfering endogenous expression of the Nestin, so that the organ fibrosis is alleviated or relieved. Therefore, the invention provides a brand-new thought and method for targeted therapy of organ fibrosis, and has good application value and prospect.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and relates to the application of Nestin gene and protein in the treatment of organ fibrosis. More specifically, it relates to the application of Nestin gene and protein as a therapeutic target for organ fibrosis and the application in preparing a drug for treating organ fibrosis. Background technique [0002] Fibrosis is a wound-healing response to acute or chronic cellular injury, characterized by abnormal activation of fibroblasts and excessive deposition of extracellular matrix (ECM), and in highly progressive cases may also lead to scar tissue and ultimately Organ dysfunction and even organ failure. Fibrosis can occur in almost every organ, especially in the liver and lung. It is considered to be the main cause of morbidity and mortality in the world. Statistics show that about 45% of patients who die from the disease can be attributed to fibrosis. disease. [0003] Pulmonary fibrosis...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883G01N33/68A61K45/00A61K31/7105A61P11/00A61P1/16A61P9/00A61P13/12A61P43/00
CPCC12Q1/6883G01N33/6893A61K45/00A61K31/7105A61P11/00A61P1/16A61P9/00A61P13/12A61P43/00G01N2800/7052
Inventor 项鹏汪建成王涛
Owner SUN YAT SEN UNIV