Unlock instant, AI-driven research and patent intelligence for your innovation.

Preparation method for stem cell thermosensitive gel with high EGF expression and application of stem cell thermosensitive gel

A temperature-sensitive gel and high-expression technology, applied in the field of cell biology, can solve the problems of high treatment cost and long skin wound healing time, and achieve the effects of increasing proliferation, promoting wound healing, and accelerating wound healing.

Pending Publication Date: 2021-04-27
东莞清实生物科技有限公司
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] In order to solve the problems of long skin wound healing time and high treatment cost in the prior art, the present invention provides a technology for modifying human umbilical cord mesenchymal stem cells with EGF, and the use of said human umbilical cord mesenchymal stem cells for treating wounds. use

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Preparation method for stem cell thermosensitive gel with high EGF expression and application of stem cell thermosensitive gel
  • Preparation method for stem cell thermosensitive gel with high EGF expression and application of stem cell thermosensitive gel
  • Preparation method for stem cell thermosensitive gel with high EGF expression and application of stem cell thermosensitive gel

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1: Inserting the gene EGF into the lentiviral vector pCDH-CMV-MCS-Puro

[0037] Entrust Beijing Huada Gene Technology Co., Ltd. to synthesize the EGF gene fragment, connect it to the PUC57 cloning vector, insert it into the pCDH-CMV-MCS-EF1-Puro vector after PCR amplification and double enzyme digestion (see figure 1 ), transformed into E.coli (stbl4), and the plasmid was extracted and purified using the plasmid purification kit of OMEGA Company.

Embodiment 2

[0038] Example 2: Preparation of UC-MSCs with high expression of EGF

[0039] (1) Preparation of UC-MSCs

[0040] The cooperating institution, Dongguan Binhaiwan Central Hospital, provided healthy newborn umbilical cords, kept them sterile at low temperature and sent them to the cell preparation laboratory within 3 hours. Separation of Huahe jelly tissue, chopped to 0.5mm 2 Adhesive culture of small pieces of different sizes, culture and subculture with UltraCULTURE medium, and observe the growth status and shape of the cells in a timely manner. When the growth status and shape are good, prepare a certain number of third-generation (P3) UC with a viability > 90% - After MSCs, carry out quality control, including: detection of bacteria, fungi, endotoxin, viability, surface markers, etc.

[0041] (2) Lentiviral packaging

[0042] 1. Take out the frozen 293T cells from the liquid nitrogen tank, quickly put them into a 37°C water bath and shake them quickly, try to dissolve the...

Embodiment 3

[0054] Example 3: Functional verification of UC-MSCs cells with high expression of EGF

[0055] (1) Proliferation experiment

[0056] (1) Co-culture with polyester membrane Transwell with 0.4 μm pore size, observe the effect of cytokines secreted by UC-MSCs with high EGF expression on the proliferation of model cells, inoculate model cells in the lower chamber, and inoculate UC-MSCs with high EGF expression in the upper chamber , the area ratio of the culture bottom of the upper chamber and the lower culture plate is 1:2.1.

[0057] (2) Inoculate 5×10 in a 6-well plate 5 HaCaT (human normal skin immortalized keratinocyte cell line) or HFF1 (human fibroblast cell line).

[0058] (3) The upper chamber was inoculated with 2×10 5 UC-MSCs with high expression of EGF were placed in a 6-well plate and co-cultured with HaCaT or HFF1. The control group was inoculated with UC-MSCs in the upper chamber, and 5×10 cells were inoculated in a 6-well plate. 5 HaCaT or HFF1 cells.

[0059...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a preparation method for stem cell thermosensitive gel with high EGF expression and application of the stem cell thermosensitive gel. Mesenchymal stem cells have self-renewal and multi-directional differentiation capability and an excellent skin tissue wound treating effect. EGF can be used for stimulating propagation of epidermal cells and endothelial cells and has a good treatment effect on repairing and healing of a treated wound surface. Liquid thermosensitive gel liniment is prepared from EGF-gene-modified umbilical mesenchymal stem cells, gelatinous solid is formed in the presence of a skin temperature after a wound is locally coated with the gel liniment, and thus, stem cells in gel can be fixed to the wound surface for a long time; a gel support is beneficial to maintaining of stem property and multi-directional differentiation potential of the stem cells, and the probability that the stem cells are directionally migrated to the wound surface, cover a local skin defected region and are differentiated into skin tissue cells is improved greatly; and Successful-colonized stem cells express the EGF and are continuously secreted to a local region of the wound surface so as to propagate local epidermal and endothelial cells, and the complete healing of the wound surface is accelerated finally under the joint action of three factors.

Description

technical field [0001] The invention relates to the field of cell biology, in particular to a preparation method and application of a stem cell temperature-sensitive gel with high EGF expression. Background technique [0002] The skin is the largest organ of the human body, located on the surface of the human body, and is the first line of defense of the human body. Skin is composed of epidermis, dermis and subcutaneous tissue, in addition to rich blood vessels, lymphatic vessels, nerves, muscles and skin appendages (including hair, hair follicles, sebaceous glands, sweat glands, nails). The epidermis is differentiated from the ectoderm, which belongs to stratified squamous epithelium, and is mainly composed of two types of cells: keratinocytes and dendritic cells; according to the differentiation stage and characteristics of keratinocytes, the epidermis is viewed from the inside to the outside under the light microscope Divided into basal layer, spinous layer, granular lay...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/18A61K9/06A61K47/34A61P17/02C12N15/867C12N5/10A61K35/28
CPCA61K35/28A61K38/1808A61K9/06A61K47/34A61K9/0014A61P17/02C12N15/86C12N5/0668C07K14/485C12N2740/15043C12N2510/00A61K2300/00
Inventor 王冰李治寰李艳群卢佳豪张伟涛
Owner 东莞清实生物科技有限公司