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Application of histone methyltransferase inhibitor in improving animal round sperm injection embryo development efficiency

A technology of methyltransferase and embryo development, which is applied in the field of medicine, can solve the problems of low implantation rate, low live birth rate, and high miscarriage rate, and achieve the effect of increasing the blastocyst development rate and improving the feasibility

Active Publication Date: 2021-08-10
SOUTHERN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although ROSI technology has been applied in a variety of animals including mice, rabbits, monkeys and humans, there are still problems such as low implantation rate, high miscarriage rate, and low live birth rate

Method used

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  • Application of histone methyltransferase inhibitor in improving animal round sperm injection embryo development efficiency
  • Application of histone methyltransferase inhibitor in improving animal round sperm injection embryo development efficiency
  • Application of histone methyltransferase inhibitor in improving animal round sperm injection embryo development efficiency

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Example 1 Effect of Histone Methyltransferase Inhibitor A366 on Improving the Developmental Efficiency of Round Sperm Injection Embryos

[0082] 1. Oocyte Collection

[0083] (1) Prepare egg storage dishes: use 3.5cm imported Eppendorf embryo culture dishes, make eight 30μL culture drops with M16 medium for each dish, add 2.5mL paraffin oil, submerge the culture medium, and put them in the incubator (37°C , 5%CO 2 ) Pre-balanced for 3h.

[0084] (2) Select 10-week-old C57BL6 / N female mice (C57) in the estrous period, weighing about 18-25 g, purchased from Weitong Lihua. Calculate the unit of hormone injection according to body weight, inject 8U per 20g of body weight; inject pregnant horse serum gonadotropin (PMSG) intraperitoneally at around 7:30 in the evening of the same day, and inject human chorionic gonadotropin 48 hours after PMSG starts Hormone (Human chorionic gonadotropin, HCG) was used to induce superovulation; eggs were retrieved 13.5 hours after HCG inje...

Embodiment 2

[0116] Example 2 Effect of concentration and action time of histone methyltransferase inhibitor A366 on blastocyst development rate of embryos injected with round sperm

[0117] 1. The effect of the concentration of histone methyltransferase inhibitor A366 on the blastocyst development rate of round sperm injection embryos

[0118] This test process is identical with step 1, 2, and 3 in embodiment 1, and the difference is only in:

[0119] (1) The B6D2F1 strain of mice was used in this experiment, which is the F1 offspring obtained from the mating of DBA / 2N male mice and C57BL / 6N female mice. C57BL / 6N and DBA / 2N were purchased from Weitong Lihua.

[0120] (2) The concentrations of histone methyltransferase inhibitory A366 in the activation solution and KSOM culture solution in the third step of the test process were 150 nM, 300 nM and 600 nM respectively, and each treatment was repeated 3 times.

[0121] The result is as Figure 4 It is shown that the histone methyltransfera...

Embodiment 3

[0127] Example 3 Effect of Histone Methyltransferase Inhibitors on Improving the Blastocyst Development Rate of Round Sperm Injected Embryos

[0128] This test process is identical with step 1, 2, and 3 in embodiment 1, and the difference is only in:

[0129] (1) The B6D2F1 strain of mice was used in this experiment, which is the F1 offspring obtained from the mating of DBA / 2N male mice and C57BL / 6N female mice. C57BL / 6N and DBA / 2N were purchased from Weitong Lihua.

[0130] (2) The type and concentration of histone methyltransferase in the chemical activation solution containing histone methyltransferase and KSOM culture medium were different during the test: Figure 6"A366-300 nM" means A366 with a concentration of 300nM; "A366-150 nM" means A366 with a concentration of 150nM; ); "BIX01294-100 nM" indicates BIX-01294 at a concentration of 100 nM; "BRD4770-2.5 μM" indicates BRD4770 at a concentration of 2.5 μM; "BRD4770-1.25 μM" indicates BRD4770 at a concentration of 1.25 μ...

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Abstract

The invention belongs to the field of medicines, and discloses application of a histone methyltransferase inhibitor to improvement of animal round sperm injection embryo development efficiency. The invention discloses the application of the histone methyltransferase inhibitor in improving the animal round sperm injection embryo development efficiency for the first time. Compared with an existing round sperm injection technology, the blastocyst development rate, implantation rate and living yield of the round sperm injection embryo of the animal can be remarkably improved through the histone methyltransferase inhibitor under the condition that transgenic risks are not introduced; in addition, the round sperms treated by the histone methyltransferase inhibitor can be injected to generate the live F0 generation to normally breed offspring, a good round sperm injection embryo in-vitro culture system is established, and the feasibility of the round sperm injection technology is improved.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to the application of a histone methyltransferase inhibitor in improving the developmental efficiency of animal circular sperm injection embryos. Background technique [0002] There are no mature sperm or elongated sperm cells in the testes and epididymis of patients with severe azoospermia, so they cannot obtain genetic offspring by intracytoplasmic sperm injection (ICSI). The round spermatid injection (ROSI) technology is to inject round sperm into oocytes and fertilize oocytes through auxiliary activation, thereby helping these azoospermic patients obtain genetic offspring. Although the ROSI technique has been applied in a variety of animals including mice, rabbits, monkeys, and humans, there are still problems such as low implantation rate, high abortion rate, and low live birth rate. Therefore, improving the developmental potential of ROSI embryos can strongly promote the c...

Claims

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Application Information

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IPC IPC(8): C12N5/073C12N5/075C12N5/076
CPCC12N5/0604C12N5/0609C12N5/061C12N2501/72C12N2500/12C12N2500/16C12N2500/20C12N2501/999
Inventor 赵小阳王京任少芳周偲马琳子李朝晖罗芳常港
Owner SOUTHERN MEDICAL UNIVERSITY
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