Preparation and application of cell-eliminating pig corium substrate without cytotoxicity

Abstract

In the present invention, the porcine decellularized dermal matrix is ​​prepared by placing the dermal matrix in a 0.5Mol / L sodium hydroxide solution prepared with triple-distilled water and continuously shaking it, then rinsing it with triple-distilled water and phosphoric acid balanced saline, and freeze-drying , packed and sterilized; pig acellular dermal matrix is ​​used as a scaffold for in vitro culture of epidermal cells to prepare tissue-engineered composite skin. After the dermal matrix is ​​coated with type I bovine collagen dissolved in acetic acid, the pH is adjusted to 7.0-7.4, and then placed successively Soak in phosphate balanced salt solution and serum-free medium, take out the inoculated and passaged epidermal cells, and then place them in CO 2 Serum-free culture in the incubator; application of porcine acellular dermal matrix, involving the construction of tissue-engineered composite skin with porcine acellular dermal matrix as a scaffold and the preparation of tissue-engineered composite skin as a substitute for human skin.

Description

technical field [0001] The invention belongs to the technical field of medical skin tissue engineering, and in particular relates to a preparation method of non-cytotoxic porcine acellular dermal matrix; a method for constructing tissue engineering composite skin with non-cytotoxic porcine acellular dermal matrix as a scaffold; and a non-cytotoxic porcine acellular dermal matrix. Application of cellular dermal matrix in constructing tissue engineered composite skin and preparing tissue engineered composite skin as a skin substitute. Background technique [0002] At present, the preparation methods of porcine acellular dermal matrix as human skin substitute mainly include enzyme digestion method, hypertonic salt-detergent treatment method and balanced salt solution method. Among them, the enzyme digestion method has been reported most at home and abroad. It uses exogenous protease to remove the epidermal structure at 4°C or 37°C, and then supplemented with other methods, suc...

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Problems solved by technology

In the above two methods, some chemical reagents used in the preparation, such as Tritonx-100, SDS, etc., have a strong cell lysis effect, which is difficult to completely remove, resulting in a low success rate of cell culture using the dermal matrix as a scaffold, and poor reproducibility; balance The saline solution method is to place sterile skin pieces in 37°C phosphate-buffered saline solution to activate the endogenous proteases of the skin, thereby separating the epidermis and dermis, and then remove all the memory of the dermis by repeated freeze-thawing and gamma-ray irradiation. Living cells

But this method, first, the production process is relatively complicated, and the processing time is long, about 2 weeks; second, it is easy to be polluted during the preparation process; third, the antigenicity is not completely removed