CN117007806A - Targeting LXR in liver macrophages for controlling slow hepatitis B progression - Google Patents

Abstract

The invention relates to the technical fields of biology and medicine, and discloses the use of targeting LXRα in liver macrophages to control the progression of chronic hepatitis B. Through a series of experiments such as constructing a mouse chronic hepatitis B model, macrophage function analysis, macrophage oil red staining, and liver macrophage LXRα expression detection, the present invention not only determines the progression of chronic hepatitis B (in order, acute inflammation, chronic inflammation, The effector cells of liver fibrosis and liver cancer are liver macrophages, and it is also clear that LXRα in liver macrophages is an inflammatory target that controls the progression of chronic hepatitis B, which lays the foundation for the realization of the "targeted inflammation" strategy in the treatment of chronic hepatitis B. Base.

Description

Targeting LXR in liver macrophages to control the progression of chronic hepatitis B

Technical field

The invention relates to the field of biological and medical technology, specifically targeting LXR in liver macrophages for controlling the progression of chronic hepatitis B.

Background technique

The danger of HBV infection is that persistent infection can initiate the dynamic progression of liver fibrosis, cirrhosis, and liver cancer. Currently, there are approximately 350-400 million chronic hepatitis B virus carriers worldwide, and 500,000-1 million patients die from chronic hepatitis B-related cirrhosis, hepatocellular carcinoma (HCC) and liver failure every year. Currently, there are 93 million patients with chronic hepatitis B infection in my country. It is predicted that by 2050, even under an ideal model, the number of chronic hepatitis B patients in my country will still exceed 27 million. Currently, there are no specific drugs targeting hepatitis B virus, and host targets have become an important way to treat chronic hepatitis B.

Clinically, more than half of HCC cases result from chronic HBV infection. The study of the intrahepatic microenvironment of HBV-related liver cancer combined with single-cell transcriptome studies confirmed that intrahepatic inflammation, especially inflammatory cells, plays a key role in promoting disease progression. Therefore, identifying key effector cells and inflammatory targets is the basis for the implementation of the "targeted inflammation" strategy in the treatment of chronic hepatitis B.

Contents of the invention

In response to the above problems, the present invention provides targeting LXRα in liver macrophages for controlling the progression of chronic hepatitis B.

In order to achieve the above objects, the present invention adopts the following technical solutions:

In a first aspect, the present invention provides a diagnostic and therapeutic marker for the progression of chronic hepatitis B, abnormal lipid metabolism in liver macrophages, or liver fibrosis and tumor formation. The marker is LXRα in liver macrophages.

Furthermore, the progression of chronic hepatitis B is acute inflammation, chronic inflammation, liver fibrosis and liver cancer.

In a second aspect, the present invention provides the application of LXRα in liver macrophages as a drug target for the progression of chronic hepatitis B, abnormal lipid metabolism of liver macrophages, or liver fibrosis and tumor formation.

In a third aspect, the present invention provides the application of a reagent for detecting LXRα in liver macrophages in preparing diagnostic tools for the progression of chronic hepatitis B, abnormal lipid metabolism in liver macrophages, or liver fibrosis and tumor formation.

Further, the tools include kits, chips, test strips or high-throughput sequencing platforms.

In a fourth aspect, the present invention provides a product for regulating LXRα content in liver macrophages.

Further, the regulation is enhancement, and the product that enhances the LXRα content in liver macrophages is an LXRα agonist or any reagent/kit that can cause/enhance LXR expression, mRNA vaccine, small nucleic acid, expression vector (viral expression vector , Crispr expression vector), viruses, etc.

In the fifth aspect, the present invention provides the use of reagents for regulating LXRα content in liver macrophages in preparing products to control the progression of chronic hepatitis B, reverse abnormal lipid metabolism in liver macrophages, or reverse liver fibrosis and tumor formation.

Further, the product is a drug, reagent or kit.

In the sixth aspect, the present invention provides a reagent or drug for controlling the progression of chronic hepatitis B, reversing abnormal lipid metabolism of hepatic macrophages, or reversing liver fibrosis and tumor formation. The reagent or drug includes a reagent or drug for regulating hepatic macrophages. Reagent for intracellular LXRα content.

In the seventh aspect, the present invention provides a kit for controlling the progression of chronic hepatitis B, reversing abnormal lipid metabolism in liver macrophages, or reversing liver fibrosis and tumor formation. The kit includes: for controlling the progression of chronic hepatitis B, Reagents or drugs that reverse abnormal lipid metabolism of liver macrophages or reverse liver fibrosis and tumor formation.

Compared with the prior art, the present invention has the following advantages:

The present invention not only determines that the effector cells responsible for the progression of chronic hepatitis B (acute inflammation, chronic inflammation, liver fibrosis and liver cancer in sequence) are liver macrophages, but also clarifies that LXRα in liver macrophages is the inflammatory target that controls the progression of chronic hepatitis B. , which lays the foundation for the implementation of the "targeted inflammation" strategy in the treatment of chronic hepatitis B.

Description of the drawings

Figure 1 shows the results of the natural progression of human chronic hepatitis B cancer transformation simulated by the mouse chronic hepatitis B model. A shows the results of the acute inflammation phase, chronic inflammation phase and liver fibrosis phase; B shows the results of the liver cancer phase.

Figure 2 shows the observation results of macrophages; A is the GO analysis of liver macrophages; B is the oil red staining results of macrophages in the liver of model mice; C is the co-culture of Hep AD38 and macrophages. Oil red staining results of macrophages.

Figure 3A shows the expression of LXRα in macrophages in the co-culture system after removing tetracycline; Figure 3B shows the changes in lipid droplets in macrophages after adding two LXR agonists respectively.

Figures 4A and B are respectively the detection results of liver fibrosis markers (TGF-β and MMP-2) in hepatocytes; Figures 4C and D are respectively the changes in the number and diameter of microspheres.

Detailed ways

The technical solution of the present invention will be described in detail below with reference to the embodiments of the present invention and the accompanying drawings. It should be noted that the following examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art.

1. A mouse chronic hepatitis B model was constructed by injecting lentivirus-packaged AAV-HBV type D vector into the tail vein of C57B/L mice, simulating the natural progression of human chronic hepatitis B cancer transformation, and found that it experienced an acute inflammatory phase, chronic Inflammation stage, liver fibrosis stage, and liver cancer stage (Figure 1A, 1B).

2. Functional enrichment (GO analysis) of macrophages (Kupffer cells) colonizing the liver in the single cell data of intrahepatic immune cells in mice at different stages after chronic hepatitis B infection was performed. It was found that with the progression of chronic hepatitis B, Kupffer cell lipids Metabolic function was significantly abnormal (Figure 2A). Further isolation of macrophages in the liver of model mice and oil red staining showed obvious accumulation of lipid droplets in the cells (Figure 2B).

Macrophages (M0) were first obtained from THP-1 cells by inducing phorbol ester (PMA) in vitro. The Hep AD38 cell line is an inducible human cell line that expresses HBV. It expresses HBV when tetracycline (Tet-) is withdrawn. After co-culture of M0 and Hep AD38(Tet-), lipid droplet aggregation was found in macrophages (Figure 2C).

The above results confirm that HBV infection with chronic hepatitis B can cause abnormal lipid metabolism in intrahepatic macrophages.

3. Further detection of the expression of LXRα found that the expression of LXRα, which is closely related to lipid metabolism, was significantly reduced in macrophages in the co-culture system (Figure 3A); after adding two LXR agonists respectively, the lipid droplets in macrophages were significantly reduced. decreased (Figure 3B). It shows that enhancing the expression of LXRα can reverse the abnormal lipid metabolism of Kupffer cells, thereby controlling the progression of chronic hepatitis B.

4. Co-culture of macrophages with abnormal lipid metabolism and Hep AD38 can cause a significant increase in liver fibrosis (TGF-β, MMP-2) markers in hepatocytes (Figure 4A, B), and increase the number of tumors in Hep AD38 cells. The stem cell formation ability is manifested by the increase in the number and diameter of microspheres (Figure 4C, D); adding LXRα agonist can significantly reverse the levels of related marker molecules and reduce the number and diameter of tumor stem cells forming spheres. It is confirmed that stimulating the expression of LXRα in macrophages can reverse liver fibrosis and tumor formation. It shows that liver fibrosis and tumor formation can be reversed by reversing abnormal lipid metabolism in Kupffer cells.

In addition, any reagents/kits, mRNA vaccines, small nucleic acids, expression vectors (viral expression vectors, Crispr expression vectors), viruses, etc. that can induce/enhance the expression of LXRα can also achieve the same effect as LXR agonists.

Although the present invention has been described in detail with general descriptions and specific embodiments above, it is obvious to those skilled in the art that some modifications or improvements can be made based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

Claims (11)

1. A diagnostic and therapeutic marker for the progression of chronic hepatitis B, abnormal lipid metabolism of liver macrophages, or liver fibrosis and tumor formation, characterized in that: the marker is LXRα in liver macrophages. 2. The marker according to claim 1, wherein the progression of chronic hepatitis B is acute inflammation, chronic inflammation, liver fibrosis and liver cancer in order. 3. The application of LXRα in liver macrophages as a drug target for the progression of chronic hepatitis B, abnormal lipid metabolism of liver macrophages, or liver fibrosis and tumor formation. 4. Application of reagents for detecting LXRα in liver macrophages in the preparation of diagnostic tools for the progression of chronic hepatitis B, abnormal lipid metabolism in liver macrophages, or liver fibrosis and tumor formation. 5. The application according to claim 4, characterized in that the tool includes a test kit, a chip, a test paper or a high-throughput sequencing platform. 6. A product for regulating LXRα content in liver macrophages. 7. The product for regulating LXRα content in liver macrophages according to claim 6, characterized in that: the regulation is enhancement, and the product for enhancing the LXRα content in liver macrophages is an LXRα agonist or any Reagents/kits, mRNA vaccines, small nucleic acids, expression vectors or viruses that can induce/enhance LXR expression. 8. Application of the reagent according to claim 6 or 7 in the preparation of products for controlling the progression of chronic hepatitis B, reversing abnormal lipid metabolism of liver macrophages, or reversing liver fibrosis and tumor formation. 9. The application according to claim 8, characterized in that the product is a medicine, a reagent or a kit. 10. A reagent or drug for controlling the progression of chronic hepatitis B, reversing abnormal lipid metabolism in liver macrophages, or reversing liver fibrosis and tumor formation, characterized in that: the reagent or drug includes a reagent or drug for regulating the intracellular content of liver macrophages. Reagent for LXRα content. 11. A kit for controlling the progression of chronic hepatitis B, reversing abnormal lipid metabolism in liver macrophages, or reversing liver fibrosis and tumor formation, characterized in that: the kit includes: a kit for controlling the progression of chronic hepatitis B, reversing liver Reagents or drugs that cause abnormal macrophage lipid metabolism or reverse liver fibrosis and tumor formation.