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SiRNA-loaded nano-liposome hybrid micelle and preparation method and application thereof

A kind of micellar and nanotechnology, applied in the direction of pharmaceutical formula, medical preparations with non-active ingredients, medical preparations containing active ingredients, etc., can solve the problem of no treatment method, etc., and achieve the effect of simple steps and lower concentration

Active Publication Date: 2021-07-16
ZHENGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is currently no effective treatment for pregnant women with preeclampsia other than termination of pregnancy

Method used

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  • SiRNA-loaded nano-liposome hybrid micelle and preparation method and application thereof
  • SiRNA-loaded nano-liposome hybrid micelle and preparation method and application thereof
  • SiRNA-loaded nano-liposome hybrid micelle and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Preparation of siRNA-loaded nano-lipid hybrid micelles

[0053] 1) Preparation of siRNA@DLin-MC3-DMA complex:

[0054] 1.1) Preparation of DLin-MC3-DMA solution: Weigh 0.5mg DLin-MC3-DMA, add 500μL of absolute ethanol to it, fully dissolve to obtain 1mg / mL DLin-MC3-DMA solution, store at -20°C , and set aside; take 200μLDLin-MC3-DMA solution and add absolute ethanol to dilute to 2mL, mix thoroughly with ultrasound, place in a round-bottomed flask, place in a constant temperature water bath at 37°C, remove the solvent by rotary evaporation for 15min, add 4mL of phosphate buffer, Hydrate at 37°C for 20 minutes and sonicate for 5 minutes to obtain a 0.1 mg / mL DLin-MC3-DMA micellar solution, store at 4°C for later use.

[0055] 1.2) Preparation of siRNA solution: Take 1OD siRNA and centrifuge at 4000rpm / min for 3min, then slowly open the cap of the tube, add 125μL of DEPC water, shake and dissolve to obtain a 20μM siRNA solution, and then divide the siRNA solution in a ste...

experiment example 1

[0069] Characterization of physicochemical properties of siRNA-loaded nanolipid hybrid micelles

[0070] 1.1 Agarose gel electrophoresis retardation of siRNAs@DLin-MC3-DMA complex

[0071] The preparation process of agarose gel is as follows: accurately weigh 1.2g of agarose in a conical flask, add 40mL of electrophoresis buffer (1×TBE), boil and dissolve in a microwave oven, observe that there is no insoluble matter, and pour it into the gel after cooling slightly. Insert a comb into the gel plate, let it stand at room temperature for 30 minutes, and conduct the experiment after the gel solidifies. Mix the loading buffer (5×RNA LoadingBuffer) and the sample at a ratio of 1:5 on a clean parafilm, and the sample is the prepared DLin-MC 3 - siRNA@DLin-MC with volume ratios of DMA micelles to siRNA of 30:1, 25:1, 20:1, 15:1, 10:1 3 - DMA complex and free siRNA solution to be tested.

[0072] The result is as figure 2 Shown when DLin-MC 3 -When the volume ratio of DMA to siR...

experiment example 2

[0086] 2.1 Determination of particle size and zeta potential of siRNA-loaded nanolipid hybrid micelles

[0087] 2.1.1 The effect of the addition ratio of phospholipids and cholesterol

[0088] Weigh cholesterol and phospholipids with a mass ratio of 1:4, 1:5, and 1:6, respectively, and the subsequent related steps are the same as in Example 1. After the ultrasound is completed, a blank liposome (liposome) is obtained, and 1 mL is taken to detect the particle size and potential , and the results are shown in Table 2.

[0089] Table 2

[0090] No. Cholesterol: Phospholipids Particle size / nm PDI Potential / mV 1 1:4 155.2±4.90 0.290±0.026 -4.03±1.701 2 1:5 137.1±5.90 0.259±0.034 -7.33±2.921 3 1:6 150.0±1.73 0.303±0.023 -3.19±0.587

[0091] As can be seen from Table 2, when the mass ratio of cholesterol and phospholipids measured by the Malvern particle size analyzer through dynamic light scattering is 1:5, the average particle diamete...

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Abstract

The invention discloses a siRNA-loaded nano-liposome hybrid micelle. The siRNA-loaded nano-liposome hybrid micelle comprises siRNA, cationic liposome and a lipid outer layer, wherein the the siRNA is coated by the cationic liposome from inside to outside. The siRNA provided by the invention can selectively silence three sFLT1 mRNAs subtypes of sFLT1 mainly causingover-express of a placenta, without decreasing the full-length FLT1 mRNA. The positively charged cationic liposome and the negatively charged siRNA are combined to form a compound, so that the stability of the siRNA is improved. After the compound is wrapped by the lipid outer layer, the placenta can be targeted, so that siRNAs are accumulated at the placenta, the uptake of cells to a delivery system is improved, and the escape of endosome is promoted. The invention further discloses a preparation method of the micelle and application of the micelle in preparation of drugs for treating preeclampsia, providing a new thought for gene therapy of preeclampsia.

Description

technical field [0001] The invention belongs to the technical field of medicines, and relates to a siRNA-loaded hybrid micelle and a preparation method and application thereof. Background technique [0002] Preeclampsia is a placental induced hypertensive disorder of pregnancy. Globally, preeclampsia causes nearly 76,000 maternal and 500,000 infant deaths each year. Women with preeclampsia usually rapidly develop hypertension after 20 weeks of gestation, often accompanied by proteinuria, kidney damage, HELLP (hemolysis, elevated liver enzymes and low platelets) syndrome, seizures, stroke, severe Can be life-threatening and cause death. The developmental effects on the fetus are growth restriction in the uterus, nervous system damage (such as cerebral palsy) caused by hypoxia, and severe death. Currently, there is no effective treatment for pregnant women with preeclampsia, other than termination of pregnancy. [0003] Maternal symptoms of preeclampsia are caused by abnor...

Claims

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Application Information

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IPC IPC(8): A61K9/107A61K31/713A61K47/24A61K47/28A61K47/18A61P9/12A61P15/00
CPCA61K31/713A61K9/1075A61K47/24A61K47/28A61K47/18A61P15/00A61P9/12
Inventor 刘洋张启蒙王童高兴丽
Owner ZHENGZHOU UNIV
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