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Establishment method and application of novel rat hypertension model

A method for establishing a high blood pressure technology, which can be used in pharmaceutical formulations, preparations for in vivo testing, compound screening/testing, etc., and can solve problems such as elevated

Inactive Publication Date: 2019-04-23
GUANGXI INT ZHUANG MEDICINE HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] So far, there is no report that in vitro injection of intron-derived miRNA can lead to continuous increase in blood pressure in animals, and eventually maintain a stable high pressure state

Method used

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  • Establishment method and application of novel rat hypertension model
  • Establishment method and application of novel rat hypertension model
  • Establishment method and application of novel rat hypertension model

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1 Construction and detection of human intron-derived 27nt-miRNA high expression plasmid

[0040] The human intron-derived 27nt-miRNA nucleotide sequence was obtained through the Blast system in NCBI. According to this special nucleotide sequence, a pair of primers with opposite directions of the 5' ends were designed.

[0041] Use PCR to amplify the target gene, according to the reaction system: 1 μL template 1 (10ng / μL), 32.5 μL ddH2O, 10 μL 5×PS Buffer, 4 μL dNTP Mix (2.5 mM each), 1 μL upstream amplification primer (10 μM), 1 μL downstream amplification primer Booster primer (10 μM), 0.5 μL PrimeSTAR HS DNA polymerase, total volume 50 μL reaction.

[0042] Reaction program: 95°C for 5min (1Cycle), (98°C for 10s, 55°C for 10s, 72°C for 90s) 30Cycle, 72°C for 8min, 4°C for 24h.

[0043] After the above reaction system and reaction process, a large number of target gene fragments are obtained.

[0044] Digest the pSilencer 4.1-CMV puro / miR expression vector wi...

Embodiment 2

[0048] Example 2 Rat in vivo transfection and transfection determination

[0049] After purchasing the male rats, they were balancedly fed for one week, and the rats were randomly divided into two groups. Among them, the mixture of pSil-27-nt-miRNA plasmid mixed with DNA transfection reagent and diluted with NS was injected at high speed within 3-5s by tail vein injection method, which is called the intervention group. Another group used the same method to inject the mixture of blank plasmid and DNA transfection reagent diluted with physiological saline, which was the control group, and injected once every two days for 12 consecutive weeks. Normal diet, water supply and suitable environment.

[0050] Take the material to determine whether it is transfected into the body. After 12 weeks, the arterial tissue was taken, frozen at -80°C, and frozen sections were made, and the green fluorescence of the inner wall of the arterial tissue lumen was observed under a fluorescence micr...

Embodiment 3

[0051] The mensuration of embodiment 3 rat blood pressure

[0052] The rats in Example 2 were placed in a quiet environment, and the rat tail artery non-invasive blood pressure measuring instrument was used to measure the systolic blood pressure (SBP), diastolic blood pressure (DBP), mean blood pressure (MBP) and Heart rate (HR), the blood pressure of each rat was measured continuously at least 5 times each time, and the average value was taken. Measured once a week, continuous observation for 12 weeks. During the experiment, the rats were given normal diet and water, and the environment was suitable, and the condition was acceptable. The results showed that the SBP, DBP and MBP of the animals in the intervention group all increased first and then decreased. After the 6th week, the blood pressure of the rats continued to rise to the level of hypertension and remained unchanged. SBP, DBP and MBP of control group animal appear to increase first, then decline gradually, return ...

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Abstract

The invention discloses an establishment method and application of a novel rat hypertension model. The disclosed establishment method of the novel rat hypertension model includes the following steps that 1, a human-body intron-derived 27nt-microRNA sequence is provided to construct its high expression plasmids; 2, the high expression plasmids in the step 1 is mixed with an X-treme GENEHP DNA transfection reagent in a certain proportion, after dilution with normal saline, the reagent is injected instantaneously in a SD rat at high speed from the caudal vein of the rat, the blood pressure of therat is increased to a high level, and the novel rat hypertension model is formed. The rat hypertension model is an effective animal disease model, can be used for verifying the influence of human-body intron-derived 27nt-MicroRNA on the blood pressure and vasodilator factors of the normal rat and screening antihypertensive drugs, and has wide application prospects.

Description

technical field [0001] The invention relates to the technical field of gene recombination and animal instantaneous injection, in particular to a method for establishing a novel rat hypertension model and its application. Background technique [0002] Cardiovascular disease is one of the diseases with the highest morbidity and mortality in the world today, and hypertension is the most common cardiovascular disease, and it is also known as the "first killer" that seriously endangers human health. The World Health Organization (WHO) defines hypertension as systolic blood pressure ≥ 140 mmHg and / or diastolic blood pressure ≥ 90 mmHg in the absence of antihypertensive drugs. Hypertension is divided into 1, 2, and 3 grades: 1) Grade 1 hypertension, also known as mild hypertension, systolic blood pressure 140-159mmHg or diastolic blood pressure 90-99mmHg; 2) Grade 2 hypertension, also known as moderate hypertension Hypertension, systolic blood pressure 160-179 mmHg or diastolic bl...

Claims

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Application Information

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IPC IPC(8): A61K31/713A61K49/00
CPCA61K31/713A61K49/0008
Inventor 欧和生罗雪兰
Owner GUANGXI INT ZHUANG MEDICINE HOSPITAL
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