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Skin-derived fibroblast having tendon regeneration effect and use thereof

A technology for fibroblasts, tendons, applied in the field of pharmaceutical compositions for tendon regeneration and therapy, to achieve high expression levels

Pending Publication Date: 2022-08-02
TEGO SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although there is an autologous cell therapy using tenocytes as a similar cell therapy, its disadvantage is that it needs to be accompanied by the collection of the patient's normal tendon tissue to obtain the raw cells of the cell therapy

Method used

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  • Skin-derived fibroblast having tendon regeneration effect and use thereof
  • Skin-derived fibroblast having tendon regeneration effect and use thereof
  • Skin-derived fibroblast having tendon regeneration effect and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Culture of skin-derived fibroblasts

[0044] The normal skin tissue isolated from the donor was minced, and only fibroblasts were isolated by disintegration. The isolated fibroblasts were primary cultured in Dulbecco's Modified Eagle's medium (DMEM) / F12 (Invitrogen, USA) containing 10% fetal bovine serum (FBS). primary culture), the culture conditions were maintained at 37°C, 10% CO 2 humid environment. After subculturing at intervals of about 5 to 6 days, a Master Cell Bank (MCB) was established at passage 2. Then, it was further subcultured at intervals of about 5 to 6 days, and a large Working Cell Bank (WCB) was obtained in passage 6 ( figure 1 ). When the cells in the petri dish were filled to about 70 to 80%, cells were separated using Trypsin-EDTA to carry out cell subculture. In most in vitro cell cultures, 5% CO 2 Used as a gas composition, but in the present invention, 10% CO is used 2 , to keep the pH of the medium exactly at 7.4.

[0045]...

Embodiment 2

[0047] Example 2: Confirmation of properties of skin-derived fibroblasts

[0048] 2-1. Proliferation and morphological characteristics of skin-derived fibroblasts

[0049] The skin-derived fibroblasts obtained in Example 1 were cultured from passage 2 to passage 10 under the same culture conditions, and the doubling time of the cells was calculated by counting the number of cells at the time of initial plating and the total number of cells after the cells were obtained (doubling time). As a result, it took an average of 30 hours from passage 2 to passage 10 and showed similar values ​​for the two different media.

[0050] In addition, from the results of confirming the cell proliferation level (PDL) according to the following mathematical formula 1, it was calculated that the skin-derived fibroblasts obtained in Example 1 were 2.7 to 4.5 per passage ( figure 2 , lower graph; N=3, mean standard deviation). This result is in contrast to the short cell lifespan of tenocytes in...

Embodiment 3

[0064] Example 3: Confirmation of the quality of skin-derived fibroblasts

[0065] The skin-derived fibroblasts of the cryopreserved cell bank were thawed and then re-cultured to confirm the quality. As a result, the recultured skin-derived fibroblasts exhibited excellent thawing rate and cell survival rate, and it was confirmed that the cell quality was maintained regardless of the freezing period (Table 1).

[0066] Table 1

[0067]

[0068]

[0069] In addition, the results of the foreign virus negative test and the microorganism residue test that meet the cell therapy approval standards of the Korean Ministry of Food and Drug Safety have confirmed that there are no foreign viruses and microorganisms remaining in the cell bank.

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PUM

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Abstract

The present invention relates to a human skin-derived fibroblast having a tendon regeneration effect, and a pharmaceutical composition for tendon regeneration and treatment comprising the same as an active ingredient, the fibroblast derived from the same kind of skin being similar to a tendon cell in cell morphology, according to the present invention, the expression level of the waveform protein as the fibroblast marker is high, a large amount of extracellular matrix protein is expressed, and the synthesis of collagen can be increased by acting on TGF-beta and ERK signal transmission mechanisms, such that the damaged tendon can be effectively regenerated.

Description

technical field [0001] The present invention relates to a skin-derived fibroblast having a tendon regeneration effect and a pharmaceutical composition for tendon regeneration and treatment comprising the same as an active ingredient. Background technique [0002] Tendon is a connective tissue that connects muscle and bone and transmits the force generated from muscle to bone to cause joint movement, which is composed of extracellular matrix such as tenocyte and collagen. matrix, ECM). Tenocytes are tendon-specific fibroblasts that play a role in the production and maintenance of tendon matrix. Tendon injuries caused by trauma often occur in sports, work, and daily life. Even minor trauma can cause tendon inflammation or partial tear due to degenerative changes due to aging. [0003] A rotator cuff tear refers to the supraspinatus, infraspinatus, teres minor, and subscapularis muscles that surround the scapula and help the arm move freely. The tendons of the four muscles a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077C07K14/78C07K14/495A61K35/33A61P21/00
CPCC07K14/495A61K35/33C07K14/78A61P21/00A61P19/04A61K35/12A61K9/0024A61K47/36A61K47/42C12N5/0656C12N2533/54C12N2533/52
Inventor 全世华韩职铉金伦希李智惠
Owner TEGO SCI
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