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Method for promoting in-vitro maturing of human immature oocyte by utilizing 3D printing technology

A 3D printing and oocyte technology, applied in the field of biological 3D printing, can solve the problems affecting the maturation process of oocyte culture in vitro, and achieve the effect of strong repeatability and good stability

Inactive Publication Date: 2015-11-11
ZHEJIANG UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Existing studies have shown that many factors may affect the in vitro maturation process of oocytes

Method used

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  • Method for promoting in-vitro maturing of human immature oocyte by utilizing 3D printing technology
  • Method for promoting in-vitro maturing of human immature oocyte by utilizing 3D printing technology

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035]Example 1 Using sodium alginate hydrogel to construct artificial granulosa cell hemispheres to promote the maturation of immature oocytes

[0036] Put the collected granulosa cells into a cell culture flask for culture, the medium is TCM199 medium, add 10% fetal bovine serum and 0.1% penicillin-streptomycin mixture, place at 37°C with 5% CO 2 Cultivate in the incubator for 3-7 days, take the cells in the logarithmic growth phase and digest them with 0.25% trypsin, resuspend the cells and mix them evenly with sodium alginate, add 0.02M Ca 2+ Stand to form a hydrogel, and the final concentration of granular cells in each mL of hydrogel is about 10 5 pieces.

[0037] The granulosa cell-sodium alginate hybrid gel is used as ink for three-dimensional printing, and the printed graphics are precisely designed using computer-aided design technology. According to the shape of the follicle, a bowl-shaped structure is designed, such as figure 1 As shown in the figure, 1 is the XY...

Embodiment 2

[0042] Example 2 Using collagen hydrogel to construct artificial granulosa cell hemispheres to promote the maturation of immature oocytes

[0043] Put the collected granulosa cells into a cell culture flask for culture, the medium is TCM199 medium, add 10% fetal bovine serum and 0.1% penicillin-streptomycin mixture, place at 37°C with 5% CO 2 Cultivate in the incubator for 2-7 days, take the cells in the logarithmic growth phase and digest them with 0.25% trypsin. The concentration of granule cells is 10 7 pieces.

[0044] The granulosa cell-collagen hybrid gel is used as ink for three-dimensional printing, and computer-aided design technology is used to accurately design the printed graphics. According to the shape of the follicle, it is designed into a bowl-shaped structure. The wall of this hemispherical structure is made of granulosa cells mixed with collagen hydrogel, with a diameter of 10 mm and a wall thickness of 100 μm, so as to simulate the wall granulosa cells in ...

Embodiment 3

[0049] Example 3 Using gelatin hydrogel to construct artificial granulosa cell hemispheres to promote maturation of immature oocytes

[0050] Put the collected granulosa cells into a cell culture flask for culture, the medium is TCM199 medium, add 10% fetal bovine serum and 0.1% penicillin-streptomycin mixture, and place in a 37°C 5% CO2 incubator for 3- On the 7th, the cells in the logarithmic growth phase were digested with 0.25% trypsin, and the cells were resuspended and mixed with gelatin solution (dissolved in hot water at 65-70°C and then cooled down to about 37°C) and mixed evenly at 35°C. Set into a hydrogel, the final particle cell concentration in each mL of hydrogel is 10 6 pieces.

[0051] The granulosa cell-gelatin hybrid gel is used as ink for three-dimensional printing, and the printed graphics are precisely designed using computer-aided design technology. According to the shape of the follicle, it is designed into a bowl-shaped structure. The wall of this he...

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Abstract

The invention discloses a method for promoting in-vitro maturing of human immature oocyte by utilizing a 3D printing technology. The method comprises the following steps: separating and collecting granular cells in clinic, propagating the granular cells in an in-vitro culture mode for 2 to 7 days; evenly mixing biological hydrogel with the pre-processed granular cells, printing artificial follicles by a biological 3D printing technology; adding a culture liquid into the artificial follicles, wherein the volume percentage content of human mature follicular fluid of the culture liquid is not less than 10%; culturing for 24 to 48 hours, then adding an immature oocyte-cumulus granular cell composite body, and culturing the 3D cells for 24 to 48 hours to promote the maturing. In the provided method, a 3D cell printing technology is adopted, human granular cells are used to construct an immature oocyte 3D culturing system, and multiple growth factors and simulated follicle culture liquid are added at the same time to create a simulated human follicle so as to promote the in-vitro maturing of immature oocyte.

Description

technical field [0001] The invention belongs to the technical field of biological 3D printing, relates to mechanical processing of biological materials and its application, in particular to a method for promoting the in vitro maturation of immature oocytes by using 3D printing technology to prepare artificial follicles. Background technique [0002] According to statistics, the infertility rate among the population of childbearing age in my country has reached 12.5%, and there are nearly 50 million infertility patients in the country. It has become a major social problem and deserves great attention from the whole society. The introduction of modern assisted reproduction (IVF) has brought hope to solve fertility problems for infertile patients. More available mature oocytes are obtained through controlled superovulation, and then artificial in vitro fertilization is carried out through sperm microinjection technology. Finally, the fertilized eggs cultured in vitro are implan...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/075
Inventor 金帆张帆马梁全胜蔡立义王丽雅王大辉姜中石张诚程张无忧王祺婧
Owner ZHEJIANG UNIV
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