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Anti-inflammation effect and application of immuno-regulatory molecule Rhbdd3

A technology of use and inflammatory factors, which is applied in the fields of biotechnology and medicine, and can solve problems such as unclear regulatory effects

Inactive Publication Date: 2015-11-25
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

However, the role of Rhbdd3 molecules in the regulation of dendritic cell activation and infectious diseases (such as endotoxin-induced infections) is still unclear

Method used

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  • Anti-inflammation effect and application of immuno-regulatory molecule Rhbdd3
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  • Anti-inflammation effect and application of immuno-regulatory molecule Rhbdd3

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0122] Example 1: Interfering with the expression of Rhbdd3 promotes the activation of dendritic cells and the production of inflammatory factors

[0123] Get mouse humerus and femur bone marrow cells, culture 3 days with RPMI1640 medium containing recombinant GM-CSF (10ng / ml) and IL-4 (1ng / ml) (purchased from Peprotech), remove non-adherent cells, and the remaining Cells were further cultured with RPMI1640 medium at the same concentration of GM-CSF and IL-4 for two days. Collect the semi-suspended cells and sort them with anti-mouse CD11c magnetic beads (purchased from MiltenyiBiotech) to obtain CD11c + Dendritic Cells.

[0124] The mouse dendritic cells obtained above were transfected with the interfering RNA (Si-Rhbdd3) against Rhbdd3 (Si-Rhbdd3) and the mock control (Si-mimetic) (the transfection reagent INTERFERin was purchased from Polyplus Company), and cultured in fresh RPMI1640 after 36 hours base.

[0125] The interfering RNA (Si-Rhbdd3) against Rhbdd3 and the m...

Embodiment 2

[0136] Example 2: Inhibitory effect of overexpression of Rhbdd3 on activation of dendritic cells and production of inflammatory factors

[0137]First import Rhbdd3(NM_177370) cDNA into AdMax TM The adenovirus shuttle plasmid in the viral vector system (purchased from Microbix Biosystems) was used to co-transfect HEK293 cells (purchased from ATCC) with the high shuttle plasmid and the adenovirus genome plasmid for DNA recombination. The amplification and purification of recombinant adenovirus can be carried out with reference to "AdenovirusVectorConstructionManual" ("Adenovirus Vector Construction Guide", MicrobixBiosystems Company Document: (03 / 00A) 01-1) (purification kit was purchased from MicrobixBiosystems Company), and Ad- Rhbdd3.

[0138] Mouse dendritic cells were infected with Ad-Rhbdd3 at an MOI of 50:1 (the preparation method was the same as in Example 1), and fresh RPMI1640 medium was replaced after 36 hours.

[0139] Infected dendritic cells (1×10 6 cells / ml),...

Embodiment 3

[0143] Example 3: Inhibitory effect of overexpression of Rhbdd3 on the production of inflammatory factors in serum and its effect on endotoxic shock Effects on mouse survival

[0144] A replication-deficient adenoviral vector of Rhbdd3 was constructed (same as Example 2). Will 1×10 7 PFUAd-Rhbdd3 was injected into the tail vein of mice (8-week-old male SDF-grade C57BL6 mice, purchased from SipperBK), and LPS (15 mg / kg) was injected intraperitoneally 48 hours later. After 4 hours, the mouse serum was collected by taking blood from the eyeball, and the expression levels of inflammatory factors IL-6 and TNFα in the serum were detected by the CBA method (purchased from BDpharmingen Company), and the survival rate of the mice was dynamically observed.

[0145] The results of the mouse survival test were as follows: Figure 6 shown.

[0146] The results showed that: the overexpression of Ad-Rhbdd3 adenovirus in mice can improve the survival rate of mice to 60% (60h after LPS ...

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Abstract

The invention relates to anti-inflammation effect and application of an immuno-regulatory molecule Rhbdd3, in particular to protein 3 (namely Rhbdd3) or Rhbdd3 coding sequence having an rhomboid structure domain and application of promoter or inhibitor in preparation of drug used for regulating and controlling dendritic cell activation and / or inflammatory factor level or a reagent box, and further relates to application and an application method of the above substance in resisting infection and treating chronic inflammatory diseases caused by infection and corresponding drug, drug combination or reagent box. The immuno-regulator molecule can be used for inhibiting activation of infected dendritic cells and excessive generation of inflammatory factors, increasing survival rate and survival time of organisms in endotoxic shock and protecting visceral organs like lung after being infected from inflammatory damage or can be used for immunoregulation, and has wide application prospect.

Description

technical field [0001] The invention belongs to the field of biotechnology and medicine, in particular, the invention relates to protein 3 (rhomboid domain containing 3, Rhbdd3) containing rhomboid domain in the prevention or treatment of diseases or symptoms related to the activation state of dendritic cells, treatment or prevention of infection and Inflammatory diseases caused by infection, effects in the control of inflammatory injury, mechanism of action, method of implementation and use. Background technique [0002] Infection is a very harmful clinical common disease, which can lead to local and systemic infectious manifestations of the body, and can even eventually lead to serious consequences such as septic shock, endotoxic shock, and multiple organ dysfunction syndrome (multiple organdysfunction syndrome, MODS) , once severe infection leads to shock or multiple organ failure, it causes high mortality (~50-80%) performance. Common clinical bacterial infections, such...

Claims

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Application Information

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IPC IPC(8): A61K38/45A61K39/395A61K48/00A61K45/00A61P35/00A61P37/02A61P37/06A61P37/08A61P31/04A61P31/10A61P31/12A61P33/00A61P29/00
Inventor 刘娟曹雪涛韩超峰刘书逊
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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