A method of preparing estradiol

An estradiol and cell technology, which is applied in the field of using induced mammalian cells to culture in vitro to prepare estradiol, can solve the problems of unpredictable preparation of estrogen, complex regulation network, difficulty in preparation of estrogen, etc., and achieves simple method and by-products. Fewer, milder effects

Active Publication Date: 2015-01-21
AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The production of estrogen involves the synergy of various cells and hormones. Due to the complexity of the regulatory network between various cells and hormones, it has brought difficulties for the preparation of estrogen by cell engineering.
, so far, there has been no report on the preparation of estrogen by cell engineering

Method used

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  • A method of preparing estradiol
  • A method of preparing estradiol
  • A method of preparing estradiol

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Experimental program
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Effect test

Embodiment 1

[0044] Human bone marrow-derived cells are used to induce and screen the cells described in the present invention using the compound trimethylcholanthracene (MCA), and use the cells to synthesize sexual estradiol in vitro; the cells have been obtained from the China Center for Type Culture Collection Deposit, deposit number CCTCC C2012173;

[0045] The preparation steps are:

[0046] (1) Human bone marrow-derived cells (HMBDCs, mainly composed of human bone marrow mesenchymal stem cells) were prepared by primary culture using existing technology (this example uses bone marrow flushing and adherence method), and the prepared cells were cultured in a medium containing 15% fetal DMEM medium with bovine serum, cultured at 37°C, 5% CO 2 , up to 10 5 cells / flask;

[0047] (2) Add MCA to the medium to make the final concentration 1ug / ml, and replace the medium containing MCA twice a week; MCA treatment ends after 1 week; cells are kept at 37°C, 5% CO 2 environment, continuous rou...

Embodiment 2

[0055] After induction by long-term culture in vitro using mouse bone marrow-derived cells (mBMDCs), the cells described in the present invention are produced and screened out, and the cells are used to synthesize sexual estradiol in vitro; the specific steps are:

[0056] (1) Isolate and purify mouse bone marrow-derived cells (mBMDCs), and continuously subculture them in vitro. After about 20 passages or about 6 consecutive cultures, the induced mBMDCs have the ability to produce estradiol;

[0057] (2) The concentration of estradiol produced by cells after the induction can reach up to 5000 pmol / L.

Embodiment 3

[0059] Mouse embryonic fibroblasts are used to induce the pluripotency genes Oct4, Sox2, Nanog, and c-KIT in vitro to produce and screen out the cells described in the present invention, and use the cells to synthesize estradiol in vitro; The steps are:

[0060] (1) Isolate and culture mouse embryonic fibroblasts;

[0061] (2) Using the method of gene introduction, transfer the pluripotency genes Oct4, Sox2, Nanog, c-KIT, screen the obtained cells, select the cells that can produce a higher concentration of estradiol, and proliferate and culture;

[0062] (3) The concentration of estradiol produced by the cells after the above induction can reach about 3000pmol / L.

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Abstract

The invention belongs to the field of biological cell engineering, relates to a method of preparing estradiol, and particularly relates to a method of preparing the estradiol by adoption of in-vitro culture of induced mammalian cells. In the method, a cell population generated by in-vitro special inducing of mammalian cells is adopted, is cultured in specific in-vitro conditions and can secret estrogen continuously. Secretion of the estrogen in the system fully imitates a synthesis process in the body. A process of producing the estradiol has characteristics of less side products, simple method and mild reactions and is nearly free of harm to the environment. Compared with the prior art, the method has irreplaceable advantages. In the method, mammalian somatic cells after in-vitro inducing under the special conditions have capability for producing the estradiol. The method provides a novel route for obtaining the estradiol.

Description

technical field [0001] The invention belongs to the field of biological cell engineering and relates to a method for preparing estradiol, in particular to a method for preparing estradiol by in vitro culture of induced mammalian cells. Background technique [0002] The prior art discloses that there are mainly three types of estrogens physiologically present in the human body: estradiol, estrone and estriol, which are mainly produced by ovaries, follicles, corpus luteum and placenta of pregnancy, and are of great significance for maintaining female secondary sexual characteristics , the lack of estrogen can cause multiple organ dysfunction in women's bodies. According to data, when women enter menopause, estrogen drops sharply. In the United States alone, the output value of estrogen drug replacement therapy for menopause has reached nearly 2 billion US dollars. [0003] At present, there are two main types of drugs used to supplement estrogen, one is estrone derived from h...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P33/00
Inventor 刘春芳吕元马展
Owner AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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