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Application of box Forkhead box (FOXA2) in preparation of medicine for treating chronic liver disease

A chronic liver disease, transcription factor technology, applied in gene therapy, drug combination, pharmaceutical formulation, etc., can solve problems such as unreported effects

Inactive Publication Date: 2016-01-20
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These studies suggest that FOXA2 is an important regulator of hepatocyte differentiation, function and epithelial phenotype, but the role of FOXA2 in the development of liver fibrosis has not been reported so far

Method used

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  • Application of box Forkhead box (FOXA2) in preparation of medicine for treating chronic liver disease
  • Application of box Forkhead box (FOXA2) in preparation of medicine for treating chronic liver disease
  • Application of box Forkhead box (FOXA2) in preparation of medicine for treating chronic liver disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Detection of changes in FOXA2mRNA in liver tissue by Real-timePCR

[0043] Trizol method (RNAiso, Takara) was used to extract rat DMN model and BDL model (see YueHY, YinC, XieWF, et al. Hepatocytenuclearfactor4alphaattenuateshepaticfibrosisinrats.Gut.2010; 59:236-46. for the preparation method) liver tissue RNA and measure it with a spectrophotometer its OD 260 The value was formulated into a working concentration (1 μg / μl), and the RNA integrity was detected by 1% agarose gel electrophoresis.

[0044] Add 1 μg RNA to 4 μl 5×PrimeScript RT mastermix (reverse transcription kit), add DEPC water to make up the volume to 20 μl, react at 37°C for 15 minutes; react at 85°C for 5s to inactivate reverse transcriptase, and the reverse transcription product can be obtained. After diluting the reverse transcription product, take 1 μl as a template for FOXA2 amplification, and use β-actin as an internal reference to perform PCR reaction under the same reaction condition...

Embodiment 2

[0049] Example 2: Inhibitory effect of up-regulating hepatic FOXA2 on the process of liver fibrosis in rats

[0050] Male SD rats were randomly divided into 4 groups, 10 in each group. Common food feeding, free drinking water, day and night lighting alternately. Group 1 was given intraperitoneal injection of normal saline as the negative control group; Groups 2 to 4 were liver fibrosis model groups, and DMN was injected intraperitoneally at a dose of 10 mg / ml, three times a week for a total of 4 weeks, to prepare DMN. Induced rat liver fibrosis model. The second group was set as the model control group; the third group was compared with the lentivirus Lv-GFP group (for the preparation method, see WangJ, ZhuCP, HuPF. FOXA2 suppresses the metastasis of hepatocellular carcinoma partially through matrix metalloproteinase-9 inhibition. Carcinogenesis. 2014; 35 (11): 2576-83.); The 4 groups were set as the lentivirus Lv-hFOXA2 treatment group (see WangJ, ZhuCP, HuPF for the prepar...

Embodiment 3

[0053] Embodiment 3: Immunohistochemical method detects mouse CCl 4 Changes in the expression of FOXA2 protein in model liver tissue.

[0054] Preparation of CCl 4 Injured liver fibrosis model: Male C57 mice were fed with normal feed, free to drink water, and illuminated alternately between day and night. To CCl 4 According to the intraperitoneal injection at a dose of 1ml / kg, two consecutive injections per week for a total of 5 weeks to prepare CCl 4 Induced mouse liver fibrosis model (eg Figure 4 shown).

[0055] Normal mice and CCl 4 4μm serial sections of model mouse liver tissue wax blocks, fixed in a 60°C oven for 30 minutes, dewaxed to water, 3% H 2 o 2 Place at room temperature for 10 minutes to remove endogenous peroxidase, boil citrate buffer for antigen retrieval, add 1:20 normal rabbit serum to block at room temperature for 30 minutes, add FOXA2 antibody (1:1000) dropwise at 4°C overnight; the next day PBS (pH7 .4) Wash 3 times, 5 min each time; add secondar...

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Abstract

The invention relates to the fields of molecular biology, cytobiology, gene engineering and clinical medicine such as disease treatment, and provides application of a Forkhead box (FOXA2) in preparation of medicine for treating a chronic liver disease. Particularly, the FOXA2 is used for treating liver fibrosis and liver cirrhosis. Researches prove that a role in protecting liver cells can be effectively produced by controlling liver cell FOXA2 gene expression, and therefore a new means for treating the chronic liver disease is provided.

Description

technical field [0001] The present invention relates to the fields of molecular biology, cell biology, genetic engineering and clinical medicine such as disease treatment. Specifically, the present invention relates to the treatment of human chronic liver disease and liver cirrhosis by using the forkhead-like transcription factor FOXA2 (HNF3β). Background technique [0002] Chronic liver disease caused by hepatitis virus infection, drug damage, immune damage, alcohol, metabolism and other factors can progress to liver cirrhosis, liver cancer, or even liver failure if it persists, which is life-threatening. In this course of disease, liver fibrosis is currently considered to be the only reversible pathological process. Therefore, the treatment of liver fibrosis has become a key link in preventing chronic liver disease from progressing to end-stage liver disease. [0003] Liver fibrosis is the liver's repair response to chronic damage such as viral infection, and is characte...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K38/17A61P1/16
Inventor 谢渭芬张新王维王剑
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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