JPH01230365A - Cell aggressive medical material and its manufacture - Google Patents
Cell aggressive medical material and its manufactureInfo
- Publication number
- JPH01230365A JPH01230365A JP63053836A JP5383688A JPH01230365A JP H01230365 A JPH01230365 A JP H01230365A JP 63053836 A JP63053836 A JP 63053836A JP 5383688 A JP5383688 A JP 5383688A JP H01230365 A JPH01230365 A JP H01230365A
- Authority
- JP
- Japan
- Prior art keywords
- collagen
- cross
- processed
- medical material
- heat
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 239000012567 medical material Substances 0.000 title claims description 25
- 238000004519 manufacturing process Methods 0.000 title claims description 7
- 108010035532 Collagen Proteins 0.000 claims abstract description 52
- 102000008186 Collagen Human genes 0.000 claims abstract description 52
- 229920001436 collagen Polymers 0.000 claims abstract description 52
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 239000003431 cross linking reagent Substances 0.000 claims abstract description 16
- 230000003176 fibrotic effect Effects 0.000 claims description 9
- 238000010438 heat treatment Methods 0.000 claims description 8
- 239000000463 material Substances 0.000 claims description 8
- 108010045569 atelocollagen Proteins 0.000 claims description 7
- 238000004132 cross linking Methods 0.000 abstract description 18
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 abstract description 2
- 150000001299 aldehydes Chemical class 0.000 abstract description 2
- 238000012545 processing Methods 0.000 abstract description 2
- 239000000427 antigen Substances 0.000 abstract 1
- 102000036639 antigens Human genes 0.000 abstract 1
- 108091007433 antigens Proteins 0.000 abstract 1
- 230000000644 propagated effect Effects 0.000 abstract 1
- 210000001519 tissue Anatomy 0.000 description 17
- 210000004027 cell Anatomy 0.000 description 10
- 239000000243 solution Substances 0.000 description 9
- 102000029816 Collagenase Human genes 0.000 description 6
- 108060005980 Collagenase Proteins 0.000 description 6
- 230000000052 comparative effect Effects 0.000 description 6
- 230000000704 physical effect Effects 0.000 description 6
- 229960002424 collagenase Drugs 0.000 description 5
- 238000004108 freeze drying Methods 0.000 description 5
- 238000012360 testing method Methods 0.000 description 5
- 238000010382 chemical cross-linking Methods 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 239000012153 distilled water Substances 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 239000012948 isocyanate Substances 0.000 description 3
- 150000002513 isocyanates Chemical class 0.000 description 3
- PMMYEEVYMWASQN-DMTCNVIQSA-N Hydroxyproline Chemical compound O[C@H]1CN[C@H](C(O)=O)C1 PMMYEEVYMWASQN-DMTCNVIQSA-N 0.000 description 2
- 241000700159 Rattus Species 0.000 description 2
- 239000003513 alkali Substances 0.000 description 2
- 239000002473 artificial blood Substances 0.000 description 2
- 239000012237 artificial material Substances 0.000 description 2
- 210000004204 blood vessel Anatomy 0.000 description 2
- 230000004709 cell invasion Effects 0.000 description 2
- 238000000354 decomposition reaction Methods 0.000 description 2
- 230000018044 dehydration Effects 0.000 description 2
- 238000006297 dehydration reaction Methods 0.000 description 2
- 238000004925 denaturation Methods 0.000 description 2
- 230000036425 denaturation Effects 0.000 description 2
- PMMYEEVYMWASQN-UHFFFAOYSA-N dl-hydroxyproline Natural products OC1C[NH2+]C(C([O-])=O)C1 PMMYEEVYMWASQN-UHFFFAOYSA-N 0.000 description 2
- 229960002591 hydroxyproline Drugs 0.000 description 2
- 238000002513 implantation Methods 0.000 description 2
- 210000000056 organ Anatomy 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 210000003491 skin Anatomy 0.000 description 2
- 238000007920 subcutaneous administration Methods 0.000 description 2
- FGMPLJWBKKVCDB-UHFFFAOYSA-N trans-L-hydroxy-proline Natural products ON1CCCC1C(O)=O FGMPLJWBKKVCDB-UHFFFAOYSA-N 0.000 description 2
- WCMYLBJNTQIIMH-UHFFFAOYSA-N 1,6-diisocyanatohexane;ethanol Chemical compound CCO.O=C=NCCCCCCN=C=O WCMYLBJNTQIIMH-UHFFFAOYSA-N 0.000 description 1
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide Substances CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 description 1
- FPQQSJJWHUJYPU-UHFFFAOYSA-N 3-(dimethylamino)propyliminomethylidene-ethylazanium;chloride Chemical compound Cl.CCN=C=NCCCN(C)C FPQQSJJWHUJYPU-UHFFFAOYSA-N 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 239000005057 Hexamethylene diisocyanate Substances 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 102000057297 Pepsin A Human genes 0.000 description 1
- 108090000284 Pepsin A Proteins 0.000 description 1
- 230000005856 abnormality Effects 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000002983 circular dichroism Methods 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000008094 contradictory effect Effects 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 210000004207 dermis Anatomy 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000002500 effect on skin Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 210000002950 fibroblast Anatomy 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- RRAMGCGOFNQTLD-UHFFFAOYSA-N hexamethylene diisocyanate Chemical compound O=C=NCCCCCCN=C=O RRAMGCGOFNQTLD-UHFFFAOYSA-N 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 230000008105 immune reaction Effects 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000005847 immunogenicity Effects 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 229940111202 pepsin Drugs 0.000 description 1
- 230000000737 periodic effect Effects 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 239000004814 polyurethane Substances 0.000 description 1
- 229920002635 polyurethane Polymers 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 239000001488 sodium phosphate Substances 0.000 description 1
- 229910000162 sodium phosphate Inorganic materials 0.000 description 1
- 238000000807 solvent casting Methods 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 229920001059 synthetic polymer Polymers 0.000 description 1
- 230000017423 tissue regeneration Effects 0.000 description 1
- 238000002054 transplantation Methods 0.000 description 1
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
Landscapes
- Materials For Medical Uses (AREA)
Abstract
Description
【発明の詳細な説明】
[産業上の利用分野]
本発明は細胞侵入性医用材料およびその製造法に関する
。DETAILED DESCRIPTION OF THE INVENTION [Industrial Field of Application] The present invention relates to a cell-invasive medical material and a method for producing the same.
さらに詳しくは本発明は架橋構造を有するコラーゲンを
水の存在下で加熱処理した変性コラ−ケンからなる細胞
侵入性医用材料およびその製造法に関する。More specifically, the present invention relates to a cell-invasive medical material made of modified collagen obtained by heat-treating collagen having a crosslinked structure in the presence of water, and a method for producing the same.
本発明の医用材料は生体内に埋入されて生体組織と同化
され、あるいは創傷面に被覆されて真皮組織に変換され
るので医学および生物学の分野において人工皮膚、人工
血管等として利用される。The medical material of the present invention is implanted in a living body and assimilated with living tissue, or is coated on a wound surface and converted into dermal tissue, so it can be used as artificial skin, artificial blood vessels, etc. in the fields of medicine and biology. .
[従来の技術]
生体組織に何らかの異常か生じた場合、自己の他部位の
組織あるいは、親族など免疫原性の少ない個体からの同
種移植が好ましいがそのような供給が困難な場合には人
工物をもってそれに代替するという発想は古くから存在
した。しかし、当然免疫拒絶反応の対象となるケースか
多く、そのため組織や免疫系細胞から不感作であるよう
な、いわゆる組織反応が低い物質を求める努力が続けら
れている。ポリウレタンを代表とする合成高分子を、よ
り疎水化させる方向の研究などはその一例一 つ
−
である。また、これとは全く正反対に、免疫反応を引き
起こす前に速かに物質か組織と同化]−2でL2まうこ
とにより器官としての機能を付与するという考え方があ
る。人工物としては生体由来材料であるコラーゲン等を
選択し、線維芽細胞等組織修復機能を持った細胞を早期
に侵入させて、結合組織様の組織を構築さけて1」的の
組織をおおわせ免疫反応を免れる考え方で、後者の方か
より理想に近い形である。[Prior art] When some abnormality occurs in living tissue, it is preferable to transplant allografts from tissues from other parts of the body or from individuals with less immunogenicity such as relatives, but if such supply is difficult, artificial The idea of replacing it with something has existed for a long time. However, in many cases, they are naturally subject to immune rejection, and therefore efforts are being made to find substances with low tissue reactions, which are insensitizing to tissues and immune system cells. One example is research into making synthetic polymers, such as polyurethane, more hydrophobic.
− is. In addition, in the exact opposite direction, there is the idea that organs can be given functions as organs by quickly assimilating them into substances or tissues]-2 before triggering an immune response. As the artificial material, collagen, etc., which is a bio-derived material, is selected, and cells with tissue repair functions such as fibroblasts are allowed to invade at an early stage, and the tissue is covered with 1" tissue, avoiding the construction of a connective tissue-like tissue. The idea is to avoid immune reactions, and the latter is closer to the ideal.
[発明か解決しようとする問題点〕
コラーゲンを用いた人工材料は生体由来であるため、確
かに細胞組織に対する親和性は大きいと考えられるもの
の、生体内でコラゲナーゼにより容易に分解・吸収され
るものである。そこで使用するにあっては、何らかの手
段で架橋を導入し、物性面の強化をはかる必要かある。[Problem to be solved by the invention] Since artificial materials using collagen are derived from living organisms, they are thought to have a high affinity for cell tissues, but they are easily degraded and absorbed by collagenase in living organisms. It is. When used in this situation, it is necessary to introduce crosslinking by some means to strengthen the physical properties.
架橋法としては加熱による脱水架橋、薬品を用いる化学
的架橋等を採用し得る。このうち、熱脱水架橋は薬品処
理に比べ安全性が高いか、物性的にコラゲナーゼ酵素に
対する耐性か化学的架橋に対して低い。そこで、化学的
架橋を熱架橋と併用させたり、化学的架橋単独で用いる
手段が選択される。これを実施すると、物性的での性質
向上が著しい。例えば、110℃の温度で真空下に24
時間装いて熱的な架橋を導入した場合には、コラゲナー
ゼ3 unit/ ml riに37℃下で静置すると
1日以内に溶解するのに対し、イソシアネート系架橋の
みを施した場合にはコラゲナーゼ1oounit/ m
l中に37℃で70経過しても形態に変化か見られない
。ところか、かかる強固な架橋を導入すると、導入前に
コラーゲンが有していた細胞、組織に対する親和性が大
幅に低下し、細胞侵入が阻止される傾向が出現する。つ
まり物性面の強化と、細胞、組織に対する親和性とい・
う生物学的性能の向1−とは、両立か困難な相反する事
象であり、満足する材料は従来求め得なかった。As the crosslinking method, dehydration crosslinking by heating, chemical crosslinking using chemicals, etc. can be employed. Among these, thermal dehydration crosslinking is safer than chemical treatment, and its physical properties are less resistant to collagenase enzymes or chemical crosslinking. Therefore, a method is selected in which chemical crosslinking is used in combination with thermal crosslinking, or chemical crosslinking is used alone. When this is carried out, the physical properties are significantly improved. For example, 24 hours under vacuum at a temperature of 110°C.
When thermal cross-linking is introduced over time, it dissolves within one day when left standing at 37°C in 3 units/ml of collagenase, whereas when only isocyanate-based cross-linking is applied, 1 unit of collagenase / m
No change in morphology was observed even after 70 hours at 37°C. However, when such strong crosslinks are introduced, the affinity of collagen for cells and tissues that it had before introduction is significantly reduced, and cell invasion tends to be inhibited. In other words, strengthening physical properties and affinity for cells and tissues.
The objective of biological performance (1) is a contradictory phenomenon that is difficult to reconcile, and it has not been possible to find a material that satisfies it.
[問題点を解決するための手段]
本発明の目的は、生体に埋入または創傷面に被覆した際
に生体内の分解酵素に対して抵抗性を有し、一定期間必
要な機械的強度を保持し、かつ細胞、組織に対する親和
性が良好で増殖した細胞が容易にその内部に入り込みや
すい医用材料およびその製造法を提供することにある。[Means for Solving the Problems] The object of the present invention is to have a material that is resistant to degrading enzymes in the living body when implanted in a living body or coated on a wound surface, and that maintains the necessary mechanical strength for a certain period of time. It is an object of the present invention to provide a medical material that can be retained, has good affinity for cells and tissues, and into which proliferated cells can easily enter, and a method for producing the same.
かかる本発明の1」的は以下の構成によって達成される
。The first objective of the present invention is achieved by the following configuration.
1)架橋構造を有するコラーゲンを水の存在下で加熱処
理した変性コラーゲンからなる細胞侵入性医用材料。1) A cell-invasive medical material made of denatured collagen obtained by heat-treating collagen with a crosslinked structure in the presence of water.
2)コラーゲンか再構成された線維化コラーゲンである
1項の医用材料。2) The medical material according to item 1, which is collagen or reconstituted fibrotic collagen.
3)線維化コラーゲンが線維化アテロコラーゲンである
2項の医用材料。3) The medical material according to Item 2, wherein the fibrotic collagen is fibrotic atelocollagen.
4)コラーゲンを加熱処理または架橋剤で処理して架橋
構造を有するコラーゲンを調製17、次いでこれを水の
存在下で50〜125℃で加熱することを特徴とする1
項の細胞侵入性医用材料の製造法。4) Collagen is heat-treated or treated with a cross-linking agent to prepare collagen having a cross-linked structure 17, and then this is heated at 50 to 125°C in the presence of water 1
2. Method for producing cell-invasive medical materials.
本発明の変性コラーゲンは、牛真皮由来のコラ−ケンを
酸またはアルカリ処理し、得られた玉重鎖へリックスを
有するコラーゲンを加熱処理または架橋剤で処理し、次
いで水の存在下で50〜125℃で加熱することによっ
て得られる。原料コラーゲンは、酸またはアルカリ処理
したコラ−ケンをさらにプロクターゼまたはペプシンに
よりその分子末端のテロペプチドを消化除去し、抗原性
を無くしたものか好ましい。The denatured collagen of the present invention is obtained by treating collagen derived from bovine dermis with an acid or alkali, heat-treating the obtained collagen having a helix of a tama-helix chain, or treating it with a crosslinking agent, and then heating it in the presence of water for 50 to Obtained by heating at 125°C. Preferably, the raw material collagen is one that has been made acid- or alkali-treated, and is further digested with protase or pepsin to remove the telopeptide at the molecular end, thereby eliminating antigenicity.
コラーゲンの架橋は、常法に従ってコラーゲンを加熱処
理するか架橋剤で処理することによって実施される。Cross-linking of collagen is carried out by heat-treating collagen or treating it with a cross-linking agent according to a conventional method.
加熱処理による場合は、コラーゲンを真空ドで110℃
に24時間以上保持して脱水するのか望ましい。In the case of heat treatment, the collagen is heated to 110°C in a vacuum.
It is preferable to dehydrate the water by holding it for 24 hours or more.
架橋剤で処理する場合は、架橋剤には特に制限はなく、
グルタルアルデヒドのようなアルデヒド系架橋剤、ヘキ
ザメチレンジイソシアネートのようなイソシアネート系
架橋剤、1−エチル−3−(3−ジメチルアミノプロピ
ル)カルボジイミド塩酸塩のようなカルボシト系架橋剤
等が使用される。When processing with a crosslinking agent, there are no particular restrictions on the crosslinking agent;
An aldehyde crosslinking agent such as glutaraldehyde, an isocyanate crosslinking agent such as hexamethylene diisocyanate, a carboside crosslinking agent such as 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride, etc. are used. .
架橋度が低すぎると医用材料としての十分な物理的強度
かjすられす、逆に高ずぎるとコラーゲンの構造・性質
か損イ)れるので避けるべきである。If the degree of crosslinking is too low, it will not have sufficient physical strength as a medical material, and if it is too high, the structure and properties of collagen will be impaired, so it should be avoided.
0.01−5%(w/v) 、好ましくは1〜3%(w
/v)架橋剤濃度で架橋させると適当な架橋度のコラー
ゲンが得られる。0.01-5% (w/v), preferably 1-3% (w/v)
/v) Collagen with an appropriate degree of crosslinking can be obtained by crosslinking at a crosslinking agent concentration.
架橋か導入されるべきコラーク゛ンは、二重鎖へリック
スを有する分散状の水溶性のものでは架橋しても物性か
あまり向上しないので、分散状コラ−ケンを37℃でり
ん酸系の緩衝液を用いて中和処理し、生体内にあるよう
な周期性線維構造をもつ再構成された線維化コラーゲン
の形にすることか好ましい。これにより架橋処理との相
乗効果で物性が飛躍的に向上する。If the collagen to be cross-linked is water-soluble and has double-stranded helices, its physical properties will not improve much even if it is cross-linked. It is preferable to perform a neutralization treatment using a fibrous collagen to form reconstituted fibrotic collagen having a periodic fibrous structure similar to that found in vivo. This dramatically improves the physical properties due to the synergistic effect with the crosslinking treatment.
架橋を導入されたコラーゲンはさらに水の存在下で50
〜125℃好ましくは90〜121℃で20分〜1時間
加熱される。The cross-linked collagen is further diluted with 50 ml of water in the presence of water.
It is heated at ~125°C, preferably 90-121°C for 20 minutes to 1 hour.
本発明の変性コラーゲンはコラーゲンの架橋の前または
後に適当な形態に成型される。例えばコラーゲン水溶I
fkから、ソルヘントキャスト法によりフィルムをある
いは凍結乾燥法により多孔性スポンジを作製し、次いで
これを加熱処理または架橋剤処理する。あるいはコラー
ゲン水溶液に架橋剤を加えて架橋し、ゲルを形成させ次
いで成型される。The denatured collagen of the present invention is molded into a suitable form before or after collagen crosslinking. For example, collagen water soluble I
From fk, a film is produced by a solvent casting method or a porous sponge is produced by a freeze-drying method, and then this is heat-treated or treated with a cross-linking agent. Alternatively, a crosslinking agent is added to the collagen aqueous solution to cause crosslinking to form a gel, which is then molded.
本発明の変性コラーゲンは部分的にコラーゲン分子鎖が
変性しているため物性の安定性と細胞侵入性の両方の性
質を具備していると思イつれる。コラーゲンの変性度は
へソックス構造の含量によって示される。ヘリックス含
量とは、コラ−ケン特有の二重鎖ヘリックスの含量を意
味し、変性コラーゲンではこのヘリックスがランダムコ
イル化しているためへソックス含量か変性度に対応する
。Since the denatured collagen of the present invention has partially denatured collagen molecular chains, it is thought to have both physical stability and cell invasion properties. The degree of collagen denaturation is indicated by the content of hesox structures. The helix content refers to the content of double-stranded helices unique to Kolaken, and since the helices in denatured collagen are randomly coiled, it corresponds to the hesox content or the degree of denaturation.
このヘリックス含量は円偏光2色性分光計(CD)や赤
外分光光度計(IR)で測定することができる(P、[
7,Gordon et ah; Macromole
cules、 l (6)954 (1974))。本
発明の変性コラーゲンのへソックス含量は0〜80%で
あり、より好ましくは40〜70%である。This helical content can be measured using a circular dichroism spectrometer (CD) or an infrared spectrophotometer (IR) (P, [
7, Gordon et ah; Macromole
cules, l (6) 954 (1974)). The hesox content of the denatured collagen of the present invention is 0 to 80%, more preferably 40 to 70%.
以下に実施例および試験例を示して本発明をさらに具体
的に説明する。EXAMPLES The present invention will be explained in more detail by showing Examples and Test Examples below.
実施例 ]
アテロコラーゲン]、0グラムをpH3,0の希塩酸に
溶解して、0.3%(W/V)の溶液を調整する。この
溶液を4℃の恒温槽に入れ撹拌しながら、りん酸緩衝液
を加え、終濃度が、0.1%(w/v)アテロコラーゲ
ン、30mMりん酸−2−ナトリウム、100mM
NaCρであるコラ−ケン溶液を調製した。次いて、3
7℃の恒温槽に1日浸漬し、線維化アテロコラーゲン液
を得た。この液を遠心分離(5000r、p、m、 、
1.0分)して、濃縮し、 0.3%(w/v)線維
化アテロコラーゲン溶液を調製した。この溶液を一30
℃で急速凍結した後、凍結乾燥をおこないスポンジを作
製した。このスポンジを1%(W/V)へキザメチレン
・ジイソシアネートのエタノール溶液に室温で24時間
浸漬し、架橋を導入した。未架橋のイソシアネートを水
洗除去した後に、このスポンジを蒸留水に浸漬した状態
で滅菌瓶を用いて、121℃、30分の加熱処理をおこ
なった。Example] Atelocollagen], 0 g is dissolved in dilute hydrochloric acid of pH 3.0 to prepare a 0.3% (W/V) solution. This solution was placed in a constant temperature bath at 4°C, and while stirring, phosphate buffer was added to adjust the final concentration to 0.1% (w/v) atelocollagen, 30mM sodium phosphate, 100mM
A Kolaken solution, which is NaCρ, was prepared. Next, 3
It was immersed in a constant temperature bath at 7°C for one day to obtain a fibrotic atelocollagen solution. This liquid was centrifuged (5000r, p, m,
1.0 min) and concentrated to prepare a 0.3% (w/v) fibrotic atelocollagen solution. Add this solution to 130
After rapid freezing at ℃, freeze-drying was performed to prepare a sponge. This sponge was immersed in a 1% (w/v) hexamethylene diisocyanate ethanol solution at room temperature for 24 hours to introduce crosslinking. After removing uncrosslinked isocyanate with water, the sponge was immersed in distilled water and heated in a sterilized bottle at 121° C. for 30 minutes.
さらに凍結乾燥後110℃12時間真空下で加熱滅菌し
て医用材料を得た。Furthermore, after freeze-drying, the material was heat sterilized at 110° C. under vacuum for 12 hours to obtain a medical material.
実施例 2
実施例1と同様に調製し、架橋を導入したスポンジを蒸
留水に浸漬し90℃、30分の加熱処理をおこなった。Example 2 A crosslinked sponge prepared in the same manner as in Example 1 was immersed in distilled water and heat-treated at 90° C. for 30 minutes.
さらに、凍結乾燥後110℃,2時間真空下で加熱滅菌
して医用材料を得た。Furthermore, after freeze-drying, the material was heat sterilized at 110° C. under vacuum for 2 hours to obtain a medical material.
比較例 1
実施例1と同様に調製し、架橋を導入したスポンジを蒸
留水に浸漬し、40℃、30分の加熱処理をおこなった
。さらに、凍結乾燥後110℃,2時間真空下で加熱滅
菌し医用材料を得た。Comparative Example 1 A cross-linked sponge prepared in the same manner as in Example 1 was immersed in distilled water and heat-treated at 40° C. for 30 minutes. Furthermore, after freeze-drying, the material was sterilized by heating under vacuum at 110° C. for 2 hours to obtain a medical material.
比較例 2
実施例]と同様に調製し、架橋を導入したスポンジを1
10℃、2時間真空下で加熱滅菌し医用材料を得た。Comparative Example 2 A sponge prepared in the same manner as in Example 2 and introduced crosslinking was
The material was heat sterilized at 10° C. under vacuum for 2 hours to obtain a medical material.
試験例 1
各種医用材料の皮下埋入実験
実施例1,2および比較例1,2で得られたスポンジに
ついて組織適合性をみる一つの指標としてラットの皮下
埋入試験をおこなった。試験は、ラットの背に約+−、
5cmの切り込みをつくり、1 cmX 1cmXL5
mmの寸法の実施例]、2および比較例1,2で得ら
れたスポンジをこの切り込みにより作られた空隙に挿入
し、傷口を縫合糸で閉じ、移植後7[E、1.4日]」
に動物を情死せしめ背筋」二の皮膚組織を切り出し、通
常の組織切片標本作製法に従いスポンジへの細胞侵入性
について、観察をおこなった。結果を表1に示す。表1
からも明らかなように、水分存在下での加熱処理により
、細胞侵入性の著しい向上が認められた。Test Example 1 Subcutaneous implantation experiment of various medical materials A subcutaneous implantation test was conducted on rats as an indicator of tissue compatibility for the sponges obtained in Examples 1 and 2 and Comparative Examples 1 and 2. The test is carried out on the rat's back at approximately +-,
Make a 5cm incision, 1cmX 1cmXL5
The sponge obtained in Example 2 and Comparative Examples 1 and 2 with dimensions in mm was inserted into the gap created by this incision, the wound was closed with suture, and 7 [E, 1.4 days] after transplantation. ”
Animals were sacrificed to death, skin tissue from the back muscle was cut out, and the ability of cells to invade the sponge was observed using standard tissue section preparation methods. The results are shown in Table 1. Table 1
As is clear from the above, heat treatment in the presence of water significantly improved cell invasiveness.
(以下余白)
試験例 2
医用材料の酵素による分解
実施例1,2および比較例1および2で得られた各スポ
ンジを100unit/mlの細菌由来コラゲナーセ溶
液中に37℃で24時間インキュベートした。(Leaving space below) Test Example 2 Enzymatic Decomposition of Medical Materials Each sponge obtained in Examples 1 and 2 and Comparative Examples 1 and 2 was incubated in a 100 unit/ml bacterial collagenase solution at 37°C for 24 hours.
その後経時的に溶液中に溶出したコラーゲンのハイドロ
キシプロリン量を測定し、初期のコラーゲンのハイドロ
キシプロリン量との比により分解性(%)を算出した。Thereafter, the amount of hydroxyproline in the collagen eluted into the solution was measured over time, and the degradability (%) was calculated from the ratio to the amount of hydroxyproline in the initial collagen.
結果を表2に示す。The results are shown in Table 2.
表 2 スポンジの酵素による分解
表2から本発明の医用旧材は、比較例とほぼ同等の酵素
抵抗性を有していることかわかる。Table 2 Decomposition of Sponge by Enzymes From Table 2, it can be seen that the old medical material of the present invention has almost the same enzyme resistance as the comparative example.
[発明の効果]
本発明の医用+」料は、架橋構造を有するコラー−1’
3−
一 12 −
ゲンを水の存在下で加熱処理した変性コラーゲンからな
るため、生体内に埋入あるいは創傷面に被覆された際に
、コラゲナーゼに対して抵抗性を有し、一定期間、必要
とされる機械的強度を保持することができるとともに、
生体適合性に優れその内部に増殖した細胞が容易に入り
込むことができる。アテロコラーゲンを原料として得ら
れる医用材料は抗原性を有しないので、特に望ましい。[Effect of the invention] The medical material of the present invention has a crosslinked structure of Collar-1'
Since it is made of denatured collagen obtained by heat-treating 3-1 12-gen in the presence of water, it has resistance to collagenase when implanted in a living body or coated on a wound surface, and does not need to be used for a certain period of time. In addition to being able to maintain the mechanical strength that is said to be
It has excellent biocompatibility and cells that have proliferated inside it can easily enter. Medical materials obtained using atelocollagen as a raw material are particularly desirable because they do not have antigenicity.
かかる本発明の医用材料は例えば生体内留置人工心臓、
人工血管等や深度熱傷時の人工被覆祠として好適に利用
される。Such medical materials of the present invention can be used, for example, in indwelling artificial hearts,
It is suitable for use as an artificial covering shrine for artificial blood vessels and deep burns.
本発明はさらに上記医用材料の好適な製造法を提供する
。即ちコラーゲンを加熱処理または架橋剤で処理して架
橋構造を有するコラーゲンを調製し、次いでこれを水の
存在下で50〜125℃で加熱することによって本発明
の細胞侵入性医用材料を簡便に製造することができる。The present invention further provides a suitable method for producing the above medical material. That is, the cell-invasive medical material of the present invention can be easily produced by heat-treating collagen or treating it with a cross-linking agent to prepare collagen having a cross-linked structure, and then heating it at 50 to 125°C in the presence of water. can do.
Claims (1)
理した変性コラーゲンからなる細胞侵入性医用材料。 2)コラーゲンが線維化コラーゲンである請求項1の医
用材料。 3)線維化コラーゲンが線維化アテロコラーゲンである
請求項2の医用材料。 4)コラーゲンを加熱処理または架橋剤で処理して架橋
構造を有するコラーゲンを調製し、次いでこれを水の存
在下で50〜125℃で加熱することを特徴とする請求
項1の細胞侵入性医用材料の製造法。[Scope of Claims] 1) A cell-invasive medical material comprising denatured collagen obtained by heat-treating collagen having a crosslinked structure in the presence of water. 2) The medical material according to claim 1, wherein the collagen is fibrotic collagen. 3) The medical material according to claim 2, wherein the fibrotic collagen is fibrotic atelocollagen. 4) The cell-invasive medical use according to claim 1, characterized in that collagen having a crosslinked structure is prepared by heat treatment or treatment with a crosslinking agent, and then this is heated at 50 to 125°C in the presence of water. Method of manufacturing materials.
Priority Applications (6)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP63053836A JP2610471B2 (en) | 1988-03-09 | 1988-03-09 | Cell-invasive medical material and method for producing the same |
DE1989609933 DE68909933T2 (en) | 1988-03-09 | 1989-03-09 | FOR CELL-PULLABLE MEDICAL MATERIAL AND ARTIFICIAL SKIN. |
PCT/JP1989/000258 WO1989008466A1 (en) | 1988-03-09 | 1989-03-09 | Medical material permitting cells to enter thereinto and artificial skin |
AU32125/89A AU632471B2 (en) | 1988-03-09 | 1989-03-09 | Medical material permitting cells to enter thereinto and artificial skin |
EP19890903231 EP0411124B1 (en) | 1988-03-09 | 1989-03-09 | Medical material permitting cells to enter thereinto and artificial skin |
US07/970,955 US5350583A (en) | 1988-03-09 | 1992-11-03 | Cell-penetrable medical material and artificial skin |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP63053836A JP2610471B2 (en) | 1988-03-09 | 1988-03-09 | Cell-invasive medical material and method for producing the same |
Publications (2)
Publication Number | Publication Date |
---|---|
JPH01230365A true JPH01230365A (en) | 1989-09-13 |
JP2610471B2 JP2610471B2 (en) | 1997-05-14 |
Family
ID=12953872
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Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP63053836A Expired - Fee Related JP2610471B2 (en) | 1988-03-09 | 1988-03-09 | Cell-invasive medical material and method for producing the same |
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Country | Link |
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JP (1) | JP2610471B2 (en) |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2002531181A (en) * | 1998-12-01 | 2002-09-24 | クック・バイオテック・インコーポレーテッド | Polymorphic collagen biomaterial medical device |
WO2005105165A1 (en) * | 2004-04-28 | 2005-11-10 | Ihara & Company Ltd. | Stretchable collagen material and manufacturing method and use thereof |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPS57168920A (en) * | 1980-11-13 | 1982-10-18 | Heyl & Co | Collagen blend, manufacture and use |
US4522753A (en) * | 1980-07-17 | 1985-06-11 | Massachusetts Institute Of Technology | Method for preserving porosity in porous materials |
-
1988
- 1988-03-09 JP JP63053836A patent/JP2610471B2/en not_active Expired - Fee Related
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4522753A (en) * | 1980-07-17 | 1985-06-11 | Massachusetts Institute Of Technology | Method for preserving porosity in porous materials |
JPS57168920A (en) * | 1980-11-13 | 1982-10-18 | Heyl & Co | Collagen blend, manufacture and use |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2002531181A (en) * | 1998-12-01 | 2002-09-24 | クック・バイオテック・インコーポレーテッド | Polymorphic collagen biomaterial medical device |
WO2005105165A1 (en) * | 2004-04-28 | 2005-11-10 | Ihara & Company Ltd. | Stretchable collagen material and manufacturing method and use thereof |
Also Published As
Publication number | Publication date |
---|---|
JP2610471B2 (en) | 1997-05-14 |
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