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Application of macrophage in blood vessel barrier protection, and prevention, inhibition and treatment of ovarian cancer ascites

A technology for macrophages and blood vessel protection, applied in the field of diagnosis and treatment of gynecological diseases, to achieve the effect of protecting blood vessel barriers

Active Publication Date: 2020-06-05
THE FIFTH AFFILIATED HOSPITAL SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Tumor treatment regimens targeting TAMs can inhibit tumor growth to a certain extent, but patients commonly suffer side effects of facial edema, suggesting that macrophages have other functions to regulate blood vessels that need to be discovered

Method used

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  • Application of macrophage in blood vessel barrier protection, and prevention, inhibition and treatment of ovarian cancer ascites
  • Application of macrophage in blood vessel barrier protection, and prevention, inhibition and treatment of ovarian cancer ascites
  • Application of macrophage in blood vessel barrier protection, and prevention, inhibition and treatment of ovarian cancer ascites

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 M2 macrophages reduce the permeability of endothelial cells by inhibiting VCAM-1

[0036] 1. Experimental process of penetration analysis of tetramethylrhodamine dextran (TRITC-dextran, mw=70kDa)

[0037] The transport of TRITC-dextran (70kDa) in co-culture of endothelial cells and macrophages was investigated to determine the permeability of endothelial cells. will be 10 5 Human umbilical vein endothelial cells were plated in the upper chamber of the transwell, and after 24 hours, different subtypes of macrophages were added to the upper chamber and co-cultured for another 24 hours. After incubating 2 mg / mL TRITC-dextran in the upper chamber for 3 hours, the fluorescence intensity of TRITC-dextran was measured at an excitation wavelength of 530 nm and an emission wavelength of 540 nm, respectively.

[0038] 2. Immunofluorescence staining experiment process

[0039] For immunofluorescence analysis of cultured cells, cells grown on coverslips were fixed, mem...

Embodiment 2

[0045] Example 2 VLA-4 is down-regulated in the co-culture system with M2 macrophages and endothelial cells

[0046] Different subtypes of macrophages and endothelial cells were co-cultured to investigate the expression of VCAM-1 ligand VLA-4, the results are as follows image 3 . from image 3 As can be seen in A, the expression of VLA-4 in M2 macrophages was down-regulated in the co-culture system, which is consistent with the down-regulation of VCAM-1 expression in the co-culture system. from image 3 Immunofluorescence of B shows attenuated expression of VLA-4 in M2 macrophages in the co-culture system.

[0047] In this example, the overexpression and knockdown models of VLA-4 in macrophages were also established by plasmid transfection. VLA-4 knockdown macrophages and macrophages pretreated with VLA-4 inhibitor (CDP323) ( image 3 C) After addition to endothelial cells, decreased phosphorylation of VE-cadherin was observed, and increased phosphorylation of VE-cadheri...

Embodiment 3

[0048] Example 3 Reduced ROS generation when endothelial cells were co-cultured with M2 macrophages

[0049] In order to verify whether ROS, as a downstream molecule of VCAM-1, participates in the role of macrophages in regulating the permeability of endothelial cells, this example detected the expression of ROS in different co-culture systems, proving that ROS is a permeability-inducing factor.

[0050] Reactive oxygen species levels were determined using the dichlorodihydrofluorescein diacetate (DCFH-DA) assay. Cells were resuspended in PBS and stained with 10 μM DCFH-DA for 20 min at 37°C in the dark. After washing 3 times with PBS, PBS buffer was added to the cells. The relative fluorescence intensity at the indicated time was recorded using a flow cytometer with an excitation wavelength of 488 nm and an emission wavelength of 525 nm. The result is as Figure 5 and Figure 6 shown, from Figure 5 In the left panel of A, it can be seen that the reactive oxygen species ...

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Abstract

The invention belongs to the technical field of gynecological disease diagnosis and treatment, and discloses application of an M2 macrophage in blood vessel barrier protection, and prevention, inhibition and treatment of ovarian cancer ascites. The M2 macrophage is researched to reconstruct a molecular mechanism of a damaged blood vessel barrier, a discovery result shows that the M2 macrophage inhibits the expression of a VCAM-1 (vascular cell adhesion molecule-1) in endothelial cells, and therefore, an ROS (reactive oxygen species) level and the phosphorylation of VE-cadherin can be lowered to protect the blood vessel barrier. In addition, an ovarian cancer animal model is constructed and is combined with a clinical sample to discover that the macrophage in the ovarian cancer regulates the blood vessel barrier through a VLA-4 (very late antigen-4) / VCAM-1 approach to affect the development of malignant ascites, so that a new approach is confirmed, and a new strategy is provided for treating the ovarian cancer ascites.

Description

technical field [0001] The invention relates to the technical field of diagnosis and treatment of gynecological diseases, more specifically, the application of macrophages in protecting blood vessel barriers, and the prevention, inhibition and treatment of ovarian cancer ascites. Background technique [0002] Ovarian malignant tumor is one of the common malignant tumors of female reproductive organs, the incidence rate is second only to cervical cancer and uterine body cancer. Among ovarian malignant tumors, epithelial carcinoma (EOC) is the most common, which poses a serious threat to women's lives. [0003] Ovarian epithelial carcinoma accounts for 50-70% of ovarian tumors. Due to the lack of early diagnosis methods and early symptoms, 70% of patients are at an advanced stage when they are diagnosed. Malignant ascites is an ascites caused by cancer, and epithelial ovarian cancer is more likely to occur than any other type of tumor, mainly because EOC can metastasize to the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/15A61K45/06A61P35/00A61P7/10G01N33/574
CPCA61K35/15A61K45/06A61P7/10A61P35/00G01N33/57449G01N33/57484A61K2300/00
Inventor 何欢欢张诗博王莉杰张浩培
Owner THE FIFTH AFFILIATED HOSPITAL SUN YAT SEN UNIV
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