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Tendon tissue engineered seed cell-hypodermal fibroblast

A fibroblast, seed cell technology, applied in the field of medicine and biomedical engineering, can solve the problems of lack of seed cells, limited clinical application, slow proliferation, etc.

Inactive Publication Date: 2004-06-30
上海第二医科大学附属第九人民医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Studies have shown that tenocytes seeded in biodegradable materials must reach a certain amount of cells, but tenocytes are the basic functional unit of tendon, with a high degree of differentiation, relatively slow proliferation, and even lose the ability to enter the proliferative phase after multiple passages. The lack of cells has become one of the difficulties in tendon tissue engineering, which also limits its clinical application

Method used

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  • Tendon tissue engineered seed cell-hypodermal fibroblast
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Examples

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preparation example Construction

[0075] The preparation method of the tissue-engineered tendon of the present invention is simple and convenient. A certain number of seed cells are mixed with a pharmaceutically acceptable biodegradable material, and then coated with a biofilm, or the seed cells are inoculated on a biofilm-coated biological in degradable materials.

[0076] The shape of the tissue-engineered tendon graft of the present invention is not particularly limited, and can be shaped arbitrarily according to the shape of tissue defect. Usually, grafts are long strips. Taking a cylinder-like graft as an example, its sides are covered with a membranous covering, and its ends may or may not be covered with a membranous covering.

[0077] The biofilm typically covers at least 60%, preferably at least 70%, more preferably at least 80%, most preferably at least 90% of the external surface area of ​​the biodegradable material.

[0078]The seed cell concentration in the tissue engineered tendon of the presen...

Embodiment 1

[0087] Isolation and culture of fibroblasts

[0088] Five young domestic pigs, weighing about 10kg, were provided by Chuansha Breeding Factory.

[0089] Under general anesthesia and aseptic conditions, the skin tissue of the flank of the pig was taken and cut into 2×2×2mm 3 The large and small tissue pieces were washed twice with phosphate buffer (PBS, containing 100 U / ml each of penicillin and streptomycin), added twice the volume of 1 mg / ml type II collagenase (Worthington, Freehold, NJ, USA), and placed at 37°C Digested on a constant temperature shaker for 4 hours, filtered and centrifuged with a 150-mesh nylon mesh, washed twice with PBS, counted, stained with trypan blue to check the viability of fibroblasts, and used 1×10 6 / plate density (petri dish diameter 100mm) culture cells, culture medium is Ham'F-12 (Gibco, Gland, Island, NY, USA) containing 10% fetal bovine serum, L-glutamine 300ug / ml, vitamin C50ug / ml, penicillin, streptomycin each 100U / ml, the fibroblasts ...

Embodiment 2

[0091] Isolation and expansion of tenocytes in vitro

[0092] In this example, the enzymatic digestion technique was used to separate the tenocytes in the tendon tissue, and expanded and cultured them in vitro.

[0093] (1) Isolation of Tenocytes

[0094] Cut out 3 cm of superficial flexor tendon on the left side of the pig under aseptic conditions, and cut the tendon tissue into 1×2×2mm 3 Put the tissue block into a centrifuge tube and wash it three times with PBS, add 0.25% type II collagenase, place it on a shaking table at 37°C for digestion, and collect tenocytes at 4, 5, and 7 hours respectively. Isolated tenocytes were obtained by the following harvesting steps:

[0095] 1. Pass the digestive juice through a 150-mesh filter to remove tissue fragments, centrifuge the filtrate in a centrifuge at 1500 rpm for 5 minutes, and discard the supernatant.

[0096] 2. Add 5ml of PBS, vortex and centrifuge to discard the supernatant, repeat 3 times.

[0097] 3. Add 10ml of DMEM...

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Abstract

The present invention provides a tissue engineering tendon graft. It includes: (a). pharmaceutically-acceptable bio-degradable material.; (b) seed cell, said seed cell can be inoculated in the described bio-degradable material and is selected from (i) fibroblast; and (ii), mixture of fibroblast and tendon cell and / or bone marrow matrix stem cell according to the ratio of 1:10-10000:1; and (c) bio-membrane, the exterior of said bio-degradable material is covered with said bio-membrane. Said invented graft can beused for effectively curing musculotendinous defect. Said invention also provides its preparation and application.

Description

technical field [0001] The invention relates to the fields of medicine and biomedical engineering, and more specifically relates to a tissue-engineered tendon constructed with fibroblasts, a preparation method and application thereof. Background technique [0002] Tendon is a bundle of dense collagen fibrous connective tissue that connects the belly of skeletal muscle to bone, and is an integral part of skeletal muscle. The structure of the tendon can be regarded as the continuation and degeneration of the muscle. The tendons of fresh specimens are silvery-white, cord-like, and tough in texture. There are ripples on the surface in the relaxed state. If the tension exceeds 4% of the relaxed state, the ripples will disappear, and the ripples will reappear after the tension is removed. Tendons and muscles differ markedly in morphology, structure, and function. Muscle is a highly specialized tissue with contractility, rich blood supply, strong metabolism, and strong anti-infe...

Claims

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Application Information

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IPC IPC(8): A61L27/00C12N5/00
Inventor 曹谊林刘伟崔磊
Owner 上海第二医科大学附属第九人民医院
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