Method for screening and culturing extracellular hair follicle stem cell matrix for clinic treatment level cell therapy

A hair follicle stem cell and cell therapy technology, applied in the field of biological stem cell technology, can solve the problems of short laying cycle, low fibroblast adhesion force, loss, etc., and achieve the effects of promoting wound healing, promoting skin diseases, and prolonging transplantation time.

Inactive Publication Date: 2016-01-20
AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Therefore, if the trophoblast is not used, it is possible that the differentiation of skin cells during long-term culture may cause uncontrollable development of subsequent research;
[0036] Method 6 (homologous fibroblast attachment screening method): Scholar Jin Yan et al. used references to improve scholars HagarB and Dunnwald mouse fibroblasts, changed the donor fibroblasts pre-expanded in vitro as the attachment substrate as a trophoblast, and cultured homologous fibroblasts. Epidermal cells, but the patent does not specify the detailed steps of how to prepare the trophoblast plate, the plating cycle is short, the fibroblast adhesion force is low, and the repeated PBS cleaning and purification process of screening will lose part of the cultured fibro

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  • Method for screening and culturing extracellular hair follicle stem cell matrix for clinic treatment level cell therapy
  • Method for screening and culturing extracellular hair follicle stem cell matrix for clinic treatment level cell therapy
  • Method for screening and culturing extracellular hair follicle stem cell matrix for clinic treatment level cell therapy

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Embodiment 1

[0104] Embodiment 1 The specific implementation steps and methods of the preparation of the hair follicle stem cell highly mimic in vivo cell matrix of the present invention:

[0105] Step 1: Skin disinfection process: take the donor skin including the following (embryonic skin, adult prepuce outer plate, scalp, armpit skin, and trunk skin can all be used) faulty skin with epidermis and dermis structure, cut into tissue pieces and soak in Dilute it in a solution containing 0.05-0.1% benzalkonium bromide with normal saline for 10-15 minutes, disinfect it, and wash the residual benzalkonium bromide repeatedly with sterile normal saline.

[0106] Step 2: Freeze-dry at low temperature -60-80°C, vacuum up to 20-110pa, remove more than 90-95% of the water in the tissue block, take it out, seal it and pack it in vacuum, and sterilize it by irradiation at a dose of 25-60KGY. The finished product can be stored at room temperature for up to 5 years. To start, you only need to soak at 3...

Embodiment 2

[0131] Embodiment 2 The method for culturing hair follicle stem cells for clinical therapeutic grade cell therapy in the highly mimicked in vivo extracellular matrix of the present invention includes:

[0132] Screening of skin samples:

[0133] The screening sites for hair follicle stem cells include embryonic skin, outer plate of foreskin, scalp, beard, and armpit skin (preferably skin tissue with a lot of hair).

[0134] Under the microscope, fix the skin (head, beard, armpit hair, and pubic hair) with a dissecting needle, use ophthalmic forceps to clamp the root of the vibrissa hair follicle close to the epidermis, pull out the complete hair follicle along the direction of hair growth, and carefully scratch it with ophthalmic scissors The dermis sheath, and the hair follicles are peeled off, the connection between the dermal papilla and the dermis sheath is cut off, and part of the hair shaft and hair root fat is cut off. Shred hair follicle tissue to obtain uniform parti...

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Abstract

The invention discloses culturing, screening and amplifying techniques for hair follicle stem cells for clinic treatment level cell therapy. The bottlenecks are always research hotspots and difficulties in the biological stem cell field; currently, by carrying out separation and purification by virtue of remarkable adhering properties of a basement membrane, the hair follicle stem cells are obtained by virtue of three-dimensional culture and amplification measures. By depending on amplification anchorage-dependent cells, a three-dimensional high-simulation in-vivo extracellular matrix system is provided for screening and separating the levels of target cells. The method is a novel culture technique for culture in vitro of biological cells of stem cells of adults (embryo skin) in a high-simulation in-vivo adhesion environment. Skin seed hair follicle cells of targeted tissue engineering cultured through adhesion screening in the environment of a high-simulation substrate have high abdication capacity and multiplication capacity and can form a differentiated cortex structure in vitro. By screening immunogenic cells, immune escape is safely achieved, the transplant time is prolonged, the clinic application of tissue engineering is increased, the healing of wounds is promoted, and a solution with absolute advantages for the clinic treatment level cell therapy is provided for diseases of skin.

Description

1. Technical field: [0001] The invention belongs to the field of biological stem cell technology and tissue engineering. It relates to a technique for constructing a highly simulated in vivo attachment environment in vitro, separating and screening, and cultivating and expanding hair follicle stem cells. Obtain active seed stem cells from hair follicle stem cells used in tissue engineering skin construction technology. 2. Background technology: [0002] The skin is the largest organ of the human body. It plays a very important physiological role in resisting biological invasion, ultraviolet radiation, preventing water loss, regulating body temperature and maintaining individual appearance. It is an important part of the human immune system, and skin cells are the composition The basic unit of this organ, the basic research field of skin cells is an important scientific research way to reveal skin health, development, prevention and treatment. [0003] The extracellular mat...

Claims

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Application Information

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IPC IPC(8): C12N5/074
Inventor 朱宁文王凌仪贾正亓发芝李伟刘天一任捷苏丽娜刘建兰
Owner AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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