JPH01230366A - Cell aggressive medical material - Google Patents
Cell aggressive medical materialInfo
- Publication number
- JPH01230366A JPH01230366A JP63053837A JP5383788A JPH01230366A JP H01230366 A JPH01230366 A JP H01230366A JP 63053837 A JP63053837 A JP 63053837A JP 5383788 A JP5383788 A JP 5383788A JP H01230366 A JPH01230366 A JP H01230366A
- Authority
- JP
- Japan
- Prior art keywords
- collagen
- helices
- denatured
- medical material
- denatured collagen
- 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 abstract description 25
- 108010035532 Collagen Proteins 0.000 claims abstract description 68
- 102000008186 Collagen Human genes 0.000 claims abstract description 68
- 229920001436 collagen Polymers 0.000 claims abstract description 68
- 239000000463 material Substances 0.000 claims description 8
- 239000011248 coating agent Substances 0.000 claims description 4
- 238000000576 coating method Methods 0.000 claims description 4
- 239000000969 carrier Substances 0.000 abstract description 4
- 102000004190 Enzymes Human genes 0.000 abstract description 3
- 108090000790 Enzymes Proteins 0.000 abstract description 3
- 238000004925 denaturation Methods 0.000 abstract description 2
- 230000036425 denaturation Effects 0.000 abstract description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 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
- 238000000354 decomposition reaction Methods 0.000 abstract 1
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- 108010045569 atelocollagen Proteins 0.000 description 24
- 210000001519 tissue Anatomy 0.000 description 24
- 239000000243 solution Substances 0.000 description 23
- 210000004027 cell Anatomy 0.000 description 22
- 238000004132 cross linking Methods 0.000 description 20
- 238000012360 testing method Methods 0.000 description 18
- 239000011159 matrix material Substances 0.000 description 13
- 241000700159 Rattus Species 0.000 description 11
- 230000000052 comparative effect Effects 0.000 description 10
- 238000000034 method Methods 0.000 description 9
- 206010052428 Wound Diseases 0.000 description 8
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- 230000008595 infiltration Effects 0.000 description 8
- 238000001764 infiltration Methods 0.000 description 8
- 238000002360 preparation method Methods 0.000 description 8
- 238000010382 chemical cross-linking Methods 0.000 description 7
- 230000018044 dehydration Effects 0.000 description 7
- 238000006297 dehydration reaction Methods 0.000 description 7
- 230000001575 pathological effect Effects 0.000 description 7
- 210000003491 skin Anatomy 0.000 description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 6
- 238000002513 implantation Methods 0.000 description 6
- 238000000338 in vitro Methods 0.000 description 6
- 238000010186 staining Methods 0.000 description 6
- 210000002950 fibroblast Anatomy 0.000 description 5
- 238000010438 heat treatment Methods 0.000 description 5
- 230000000704 physical effect Effects 0.000 description 5
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- 102000029816 Collagenase Human genes 0.000 description 4
- 108060005980 Collagenase Proteins 0.000 description 4
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 4
- 239000005057 Hexamethylene diisocyanate Substances 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 4
- 238000004108 freeze drying Methods 0.000 description 4
- RRAMGCGOFNQTLD-UHFFFAOYSA-N hexamethylene diisocyanate Chemical compound O=C=NCCCCCCN=C=O RRAMGCGOFNQTLD-UHFFFAOYSA-N 0.000 description 4
- 210000000440 neutrophil Anatomy 0.000 description 4
- 239000012188 paraffin wax Substances 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 238000002054 transplantation Methods 0.000 description 4
- 206010063560 Excessive granulation tissue Diseases 0.000 description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 3
- 241001465754 Metazoa Species 0.000 description 3
- 239000007864 aqueous solution Substances 0.000 description 3
- 230000004709 cell invasion Effects 0.000 description 3
- 229960002424 collagenase Drugs 0.000 description 3
- 230000007547 defect Effects 0.000 description 3
- 238000011156 evaluation Methods 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 210000001126 granulation tissue Anatomy 0.000 description 3
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 3
- 210000000056 organ Anatomy 0.000 description 3
- CPKVUHPKYQGHMW-UHFFFAOYSA-N 1-ethenylpyrrolidin-2-one;molecular iodine Chemical compound II.C=CN1CCCC1=O CPKVUHPKYQGHMW-UHFFFAOYSA-N 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 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
- 230000001413 cellular effect Effects 0.000 description 2
- 239000000515 collagen sponge Substances 0.000 description 2
- 230000000593 degrading effect Effects 0.000 description 2
- 210000004207 dermis Anatomy 0.000 description 2
- 229940088598 enzyme Drugs 0.000 description 2
- 230000028993 immune response Effects 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 239000012948 isocyanate Substances 0.000 description 2
- 150000002513 isocyanates Chemical class 0.000 description 2
- 230000014759 maintenance of location Effects 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 229940105631 nembutal Drugs 0.000 description 2
- 230000007935 neutral effect Effects 0.000 description 2
- 230000035515 penetration Effects 0.000 description 2
- WEXRUCMBJFQVBZ-UHFFFAOYSA-N pentobarbital Chemical compound CCCC(C)C1(CC)C(=O)NC(=O)NC1=O WEXRUCMBJFQVBZ-UHFFFAOYSA-N 0.000 description 2
- 239000004814 polyurethane Substances 0.000 description 2
- 229920002635 polyurethane Polymers 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 238000000807 solvent casting Methods 0.000 description 2
- 206010003694 Atrophy Diseases 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 208000019300 CLIPPERS Diseases 0.000 description 1
- 108010022355 Fibroins Proteins 0.000 description 1
- 208000005422 Foreign-Body reaction Diseases 0.000 description 1
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 description 1
- 206010062717 Increased upper airway secretion Diseases 0.000 description 1
- 102000011722 Matrix Metalloproteinase 13 Human genes 0.000 description 1
- 108010076503 Matrix Metalloproteinase 13 Proteins 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 102000057297 Pepsin A Human genes 0.000 description 1
- 108090000284 Pepsin A Proteins 0.000 description 1
- 239000004809 Teflon Substances 0.000 description 1
- 229920006362 Teflon® Polymers 0.000 description 1
- BZHJMEDXRYGGRV-UHFFFAOYSA-N Vinyl chloride Chemical compound ClC=C BZHJMEDXRYGGRV-UHFFFAOYSA-N 0.000 description 1
- 230000005856 abnormality Effects 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 150000001299 aldehydes Chemical class 0.000 description 1
- 230000000735 allogeneic effect Effects 0.000 description 1
- 230000037444 atrophy Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 208000021930 chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids Diseases 0.000 description 1
- 238000002983 circular dichroism Methods 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000008094 contradictory effect Effects 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
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- 238000004090 dissolution Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000002500 effect on skin Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 210000003195 fascia Anatomy 0.000 description 1
- DCPMPXBYPZGNDC-UHFFFAOYSA-N hydron;methanediimine;chloride Chemical compound Cl.N=C=N DCPMPXBYPZGNDC-UHFFFAOYSA-N 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 238000007654 immersion Methods 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000005847 immunogenicity Effects 0.000 description 1
- 239000007943 implant Substances 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 230000002757 inflammatory effect Effects 0.000 description 1
- 230000009545 invasion Effects 0.000 description 1
- 239000010410 layer Substances 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 230000004660 morphological change Effects 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 229940111202 pepsin Drugs 0.000 description 1
- 230000000737 periodic effect Effects 0.000 description 1
- 208000026435 phlegm Diseases 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 229920000747 poly(lactic acid) Polymers 0.000 description 1
- 229920000728 polyester Polymers 0.000 description 1
- 239000004626 polylactic acid Substances 0.000 description 1
- 229920001296 polysiloxane Polymers 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000003307 reticuloendothelial effect Effects 0.000 description 1
- 239000013464 silicone adhesive Substances 0.000 description 1
- 229920006268 silicone film Polymers 0.000 description 1
- 210000001626 skin fibroblast Anatomy 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 238000007447 staining method Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000002344 surface layer Substances 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 229920001059 synthetic polymer Polymers 0.000 description 1
- 229920003002 synthetic resin Polymers 0.000 description 1
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- 230000017423 tissue regeneration Effects 0.000 description 1
- 239000011800 void material Substances 0.000 description 1
- 230000010388 wound contraction Effects 0.000 description 1
Landscapes
- Materials For Medical Uses (AREA)
Abstract
Description
【発明の詳細な説明】 ［産業上の利用分野コ 本発明は細胞侵入性医用材料に関する。[Detailed description of the invention] [Industrial application fields] The present invention relates to cell-invasive medical materials.
さらに詳しくは本発明は担体にヘリックス含量が０〜８
０％である変性コラーゲンを結合あるいは被覆した細胞
侵入性医用材料に関する。More specifically, in the present invention, the carrier has a helical content of 0 to 8.
This invention relates to a cell-invasive medical material bound or coated with 0% denatured collagen.
本発明の医用月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. Ru.
［従来の技術］
生体組織に何らかの異常が生じた場合、自己の他部位の
組織あるいは、親族なと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様な供給か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が多
く、そのため組織や免疫系細胞から不感作であるような
、いイっゆる組織反応か低い物質を求める努力か続けら
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とはその一例である。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
やかに物質か組織と同化し７てしまうことにより器官と
しての機能をイ」与するという考え方がある。人工物と
（７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目的の組織をおおわ
せ免疫反応を免れる考え方で、後者の方がより理想に近
い形である。[Prior art] When some abnormality occurs in living tissue, it is preferable to transplant tissues from other parts of the body or allogeneic transplants from relatives with less immunogenicity, but if such supply is difficult, artificial transplantation is preferable. The idea of replacing something with something has existed for a long time. However, naturally there are many cases where they are subject to immune rejection reactions, and therefore efforts are being made to find substances that are insensitive to tissues and immune system cells and have a low tissue reaction rate. One example is research into making synthetic polymers, such as polyurethane, more hydrophobic. On the contrary, there is the idea that organs can function as organs by quickly assimilating them into substances or tissues before triggering an immune response. Artificial materials (7) are selected from bio-derived materials such as collagen, and cells with tissue repair functions such as fibroblasts are allowed to invade at an early stage, avoiding the construction of connective tissue-like tissue and covering the target tissue. The latter idea is closer to the ideal, as it avoids the immune response.
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。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. 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
する耐性が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ｊ１ｉ独で用い
る手段か選択される。これを実施すると、物性面での性
質向上か著しい。例えば、１１０°Ｃの温度で真空下に
２４時時間−て熱的な架橋を導入した場合には、コラゲ
ナーゼ３ ｕ ｎ ｉ ｔ ／ ｍｌ ＋：ｌ−＋に３
７℃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ケナーセ゛
１ｏＯｕｎｉｔ／ｍｌ中に３７°Ｃで７日経過しても形
態に変化か見られない。ところが、かかる強固な架橋を
導入すると、導入前にコラーゲンか有していた細胞、組
織に対する親和性か大幅に低下し、細胞侵入か阻止され
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上とは、両立が
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 adopted. Among these, thermal dehydration crosslinking is safer than chemical treatment, or physical resistance to collagenase enzymes is lower than chemical crosslinking. Therefore, a method is selected to use chemical crosslinking in combination with thermal crosslinking or to use chemical crosslinking alone. When this is done, the physical properties are significantly improved. For example, when thermal cross-linking is introduced at a temperature of 110°C under vacuum for 24 hours, collagenase 3 units/ml +:l−+ is
When left standing at 7°C, it dissolves within one day, whereas when only isocyanate-based crosslinking is applied, no change in form is observed even after 7 days at 37°C in 1 unit/ml of collagenase. . However, when such strong crosslinks are introduced, the affinity that collagen had for cells and tissues before introduction is significantly reduced, and cell invasion tends to be inhibited. In other words, strengthening physical properties and improving biological performance such as affinity for cells and tissues are contradictory phenomena that are difficult to reconcile, and it has not been possible to find a material that satisfies them.
［問題点を解決するための手段］
本発明の目的は生体内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何し、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か容易にその内部に入り込みや
すい医用材料を提供することにある。[Means for Solving the Problems] The purpose of the present invention is to provide resistance to degrading enzymes in the living body when implanted in the living body or coated on the wound surface, and to maintain the necessary mechanical strength for a certain period of time. It is an object of the present invention to provide a medical material that retains cells, has good affinity for cells and tissues, and allows proliferated cells to easily enter therein.
かかる本発明のｉ」的は以下の構成によって達成される
。The objective of the present invention is achieved by the following configuration.
１）担体にヘリックス含量が０〜８０％である変性コラ
ーゲンを結合または被覆したことを特徴とする細胞侵入
性医用材料。1) A cell-invasive medical material characterized by having a carrier bound or coated with denatured collagen having a helical content of 0 to 80%.
２）担体か生体吸収利料であるコ項の医用材料。2) Medical materials in item C that are carriers or bioabsorbable materials.
３）生体吸収旧材かコラーゲンである］または２項の医
用月利。3) Bio-absorbable old material or collagen] or the medical monthly benefit of item 2.
４）コラーゲンが熱脱水架橋あるいは化学架橋されてい
る１〜・３項のいずれかの項に記載の医用材料。4) The medical material according to any one of items 1 to 3, wherein the collagen is thermally dehydrated crosslinked or chemically crosslinked.
５）コラーゲンおよびヘリックス含量が０〜８０％であ
る変性コラーゲンを混合し、フィルムまたは多孔体を形
成させた後架橋されたことを特徴とする細胞侵入性医用
材料。5) A cell-invasive medical material characterized in that collagen and denatured collagen having a helix content of 0 to 80% are mixed to form a film or porous body, which is then crosslinked.
−り−
６）架橋されたコラーゲンフィルムあるいは多孔体をヘ
リックス含量０〜８０％である変性コラーゲン溶液で被
覆したことを特徴とする細胞侵入性医用材料。-ri- 6) A cell-invasive medical material characterized by coating a crosslinked collagen film or porous body with a denatured collagen solution having a helix content of 0 to 80%.
本発明のへソックス含量０〜８０％の変性コラーゲンは
、牛真皮由来のコラーゲンを酸またはアルカリ処理し、
得られた三重鎖へソックスを有するコラーゲンを水の存
在下で５０〜１２５℃好ましくは９０〜１２１℃で２０
分〜２４時間加熱することによって得られる。ヘリック
ス含量とはコラーゲン特有の三重鎖へソックスの含量を
意味し、変性コラーゲンてはこのへソックスがランダム
コイル化しているためへソックス含量が変性度に対応す
る。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、 Ｌ、 Ｇｏｒｄｅｎ ｅｔａｌ、 Ｍａｃｒｏｍ
ｏｌｅｃｕｌｅｓ、 １（Ｂ） ９５４ （１，９７４
）） 。The denatured collagen with a hesox content of 0 to 80% of the present invention is obtained by treating collagen derived from bovine dermis with acid or alkali, and
The resulting triple-stranded collagen was heated in the presence of water at 50-125°C, preferably at 90-121°C for 20
Obtained by heating for minutes to 24 hours. The helix content refers to the content of triple-stranded socks unique to collagen, and since the helical socks of denatured collagen are randomly coiled, the helical content corresponds to the degree of denaturation. This helix content is measured using a circular dichroism spectrometer (CD).
It can be measured with an infrared spectrophotometer (IR) [:
P, L, Gorden etal, Macrom
olecules, 1(B) 954 (1,974
)).
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０〜５０％である。The hesox content of the denatured collagen of the present invention is 0 to 80%
and more preferably 0 to 50%.
原料コラーゲンは酸またはアルカリ処理したコラーゲン
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
が望ましい。The raw material collagen is preferably acid- or alkali-treated collagen and further digested with protase or pepsin to remove the telopeptide at the molecular end to eliminate antigenicity.
」二記変性コラーゲンが結合または被覆される担体は、
生体内の分解酵素によって分解されず一定の期間、機械
的強度を有し、生体に受容されるものか使用される。こ
のような担体の例としてポリエステル、ポリウレタン、
塩化ビニルのような合成樹脂をあげることができるが好
適には、コラーゲン、フィブロイン、ポリ乳酸のような
生体吸収旧材か使用される。The carrier to which the denatured collagen is bound or coated is
It is used if it is not degraded by degrading enzymes in the body, has mechanical strength for a certain period of time, and is accepted by the body. Examples of such carriers include polyester, polyurethane,
Synthetic resins such as vinyl chloride can be used, but bioabsorbable old materials such as collagen, fibroin, and polylactic acid are preferably used.
最も好ましくは、コラーゲンを加熱処理または架橋剤で
処理した架橋コラーゲンが使用される。Most preferably, cross-linked collagen is used, which is obtained by heat-treating collagen or treating it with a cross-linking agent.
コラ−ケ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 in a conventional manner.
加熱処理による場合は、コラーゲンを真空下で１１．０
℃に２時間以上保持して脱水するのが望ましい。In the case of heat treatment, the collagen is heated to 11.0 under vacuum.
It is desirable to dehydrate by holding at ℃ for 2 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, and examples include aldehyde crosslinking agents such as glutaraldehyde, isocyanate crosslinking agents such as hexamethylene diisocyanate, and 1-ethyl-3-(3-dimethylaminopropyl). Carbosite crosslinking agents such as carbodiimide hydrochloride are used.
架橋度か低すぎると医用材料と（２ての十分な物理的強
度が得られず、逆に高ずぎるとコラーゲンの１１４造、
性質か損われるので避けるべきである。If the degree of crosslinking is too low, sufficient physical strength of the medical material (2) cannot be obtained; on the other hand, if the degree of crosslinking is too high, collagen
It should be avoided as it will damage its properties.
０．０１〜５％（ｗ／ｖ） 、好ましくは１〜３％（Ｗ
／の架橋剤濃度で架橋させると適当な架橋度のコラーゲ
ンか得られる。0.01-5% (w/v), preferably 1-3% (W/v)
When crosslinking is carried out at a crosslinking agent concentration of /, collagen with an appropriate degree of crosslinking can be obtained.
架橋が導入されるべきコラーゲ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があま
り向上［７ないので分散状コラーゲンを３７℃でりん酸
系の緩衝液を用い゛Ｃ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
果て物性が飛躍的に向上する。If the collagen to be cross-linked is a dispersed water-soluble one with double-stranded helices, the physical properties will not improve much even if cross-linked [7], so the dispersed collagen was heated at 37°C in a phosphate buffer solution. It is preferable to perform C neutralization treatment to form reconstituted fibrous collagen having a periodic fibrous structure similar to that found in living organisms. This results in a synergistic effect with the crosslinking treatment, resulting in a dramatic improvement in physical properties.
本発明において担体としてコラーゲンを使用する場合細
胞侵入性医用材料は次の方法によって作製される。In the present invention, when collagen is used as a carrier, the cell-invasive medical material is produced by the following method.
１）コラーゲン水溶液を調製し、これを２分して一方は
そのまま放置し他方はこのコラーゲン水溶液を加熱処理
して変性させることにより変性コラーゲンとする。両溶
液を混合した後、ソルベントキャスティング法によりフ
ィルムを、凍結乾燥法により多孔性スポンジをそれぞれ
作製した後、該フィルムあるいは該多孔性スポンジに熱
架橋あるいは化学架橋を形成させることにより本発明の
医用材料を作製する。1) Prepare a collagen aqueous solution, divide it into two parts, leave one part as it is, and heat the other part to denature it to obtain denatured collagen. After mixing both solutions, a film is produced by a solvent casting method and a porous sponge is produced by a freeze-drying method, and then thermal crosslinking or chemical crosslinking is formed in the film or the porous sponge to produce the medical material of the present invention. Create.
２）コラーゲン水溶液を作製した後、ソルベントキャス
ティング法によりフィルムあるいは凍結乾燥法により多
孔性スポンジを作製する。その後、熱架橋反応あるいは
化学架橋反応を行なう。2) After preparing an aqueous collagen solution, a film is prepared using a solvent casting method or a porous sponge is prepared using a freeze-drying method. Thereafter, a thermal crosslinking reaction or a chemical crosslinking reaction is performed.
一方、変性コラーゲンはコラーゲン水溶液を加熱処理に
よって変性させることにより別途作製する。該架橋コラ
ーゲンフィルムあるいは多孔体スポンジを該変性コラー
ゲン溶液に浸漬した後、取り出し自然あるいは真空ある
いは凍結乾燥することにより本発明の医用材料を作製す
る。On the other hand, denatured collagen is separately produced by denaturing an aqueous collagen solution by heat treatment. The medical material of the present invention is produced by immersing the crosslinked collagen film or porous sponge in the denatured collagen solution, then taking it out and drying it naturally, under vacuum, or by freeze-drying.
コラーゲン以外の担体を使用する場合にも」二記と同様
の方法により変性コラーゲンの水溶液に浸漬して担体に
変性コラーゲンを被覆または結合させる。When using a carrier other than collagen, the carrier is immersed in an aqueous solution of denatured collagen to coat or bond the denatured collagen to the carrier in the same manner as described in Section 2 above.
担体に対する変性コラーゲンの組成はおよそ５〜８０％
（ｗ／ｗ）であり、より好ましくは１０〜５０％（ｗ／
ｗ）である。Composition of denatured collagen to carrier is approximately 5-80%
(w/w), more preferably 10 to 50% (w/w).
w).
次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。Next, the present invention will be explained in more detail with reference to Examples and Test Examples.
実施例 ］ ］アテロコラーゲンー変性アテロコラー
ゲンマトリックの調製
アテロコラーゲン（ＡＣ） ｉ、ＯｇをｐＨ３，０の
希塩酸に溶解させた。Examples ] Atelocollagen - Preparation of modified atelocollagen matrix Atelocollagen (AC) i, Og was dissolved in dilute hydrochloric acid at pH 3.0.
この溶液を６０℃の恒温槽て３０分間保持したのち、室
温下で２時間放置して変性アテロコラーゲン（ＨＡＣ）
の溶ン夜を得た。このよう１こして得られた変性アテロ
コラ−ケンのへリックス含量は約４０％であ−〕だ。Ｄ
、３ｖ／ｖ％アテロコラーゲンー ］〇 −
（ｐＨ３，０）溶液を撹拌しなから、０．３ｗ／ｖ％変
性アテロコラーゲン溶液を添加し混合した。この溶液を
ステンレスバットに注入し、そのまま−３０°Ｃに急速
凍結し、十分凍結した後、−４０°Ｃ１０，１トール未
満の真空下で凍結乾燥した。さらに生成物を５０ミリト
ール未昼の真空下１１０°Ｃ１２４時間処理して熱脱水
架橋した。This solution was kept in a constant temperature bath at 60°C for 30 minutes, and then left at room temperature for 2 hours to produce denatured atelocollagen (HAC).
Got a melting night. The helical content of the modified atelocolakene thus obtained was about 40%. D
, 3v/v% atelocollagen] - (pH 3,0) While stirring the solution, 0.3w/v% denatured atelocollagen solution was added and mixed. This solution was poured into a stainless steel vat, rapidly frozen as it was at -30°C, sufficiently frozen, and then freeze-dried at -40°C under a vacuum of less than 10.1 Torr. Further, the product was subjected to thermal dehydration crosslinking by treating it under a vacuum of 50 mTorr at 110°C for 124 hours.
比較例 １ アテロコラーゲンマトリックスの調製
アテロコラーゲン（ＡＣ）１．０ ｇを０．３ｗ／ｖ％
の濃度になるようにｐＨ３，０の希塩酸に溶解させた。Comparative Example 1 Preparation of atelocollagen matrix 1.0 g of atelocollagen (AC) was added to 0.3 w/v%
It was dissolved in dilute hydrochloric acid at pH 3.0 to a concentration of .
この溶液を上記の方法で凍結乾燥し、さらに熱脱水架橋
した。This solution was freeze-dried by the method described above and further cross-linked by thermal dehydration.
試験例１ アテロコラーゲン−変性アテロコラーゲン
マトリックスのｉｎ ｖｔｔｒ。Test Example 1 Atelocollagen-denatured atelocollagen matrix in vttr.
細胞侵入性試験
上記実施例１、および比較例１て得られたマトリックス
について、ラットの皮膚線維芽細胞を用いてｉｎ ｖｉ
ｔｒｏで培養実験を行ない細胞侵入性の評価を行なった
。Cell penetration test The matrices obtained in Example 1 and Comparative Example 1 were tested in vitro using rat skin fibroblasts.
A culture experiment was conducted using tro to evaluate cell invasiveness.
一］］−
６０ｍｌ１１滅菌シヤーレ〔テルモ（製）〕に直径３５
ｃｍ Ｊｇのコラーゲンスポンジを置き、線維芽細胞を
ｌＸｌ０”個／ｍｌの濃度で１ｍｌ、スポンジ上に滴下
し、２４４時間３フ
含むＤＭＥ培地を３ｍｌ入れ、３７°Ｃドで６日間培養
した。1] - 60 ml 11 sterile chalet [manufactured by Terumo] with a diameter of 35
A collagen sponge of cm Jg was placed, 1 ml of fibroblasts at a concentration of 1 x 10'' cells/ml was dropped onto the sponge, 3 ml of DME medium containing 3 ml of fibroblasts was added for 244 hours, and cultured at 37°C for 6 days.
１０％中性緩衝ホルマリン液で固定後染色を施し、光学
顕微鏡で観察［７評価した。評価の結果を表−１に示し
た。After fixation with 10% neutral buffered formalin solution, staining was performed and observed with an optical microscope [7 evaluations were made]. The results of the evaluation are shown in Table-1.
（以下余白）
表 − １
コラーゲンマトリックスへの
ｉｎ ｖｉｔｒｏ細胞侵入性試験
注）細 胞 侵入 性 スポンジの形態保持性− 全
く なし 消失（溶解）
± 軽微に侵入 はとんど溶解
＋ 小規模に侵入 検体は残存しているが著しい形態
変化
廿 中程度に侵入 小規模の収縮・溶解帯顕著に侵入
形態不変
表−１から、ＡＣ単独のマトリックスに対し、ＨＡＣを
混合したマトリックスでは、大幅に細胞の侵入性が向上
することか判った。但し、スポンジの形状維持の観点か
らは、ＨＡＣの重量％か８０％未満であることが好まし
いといえる。(Leaving space below) Table 1 In vitro cell invasiveness test into collagen matrix Note) Cell invasiveness Sponge shape retention - Not at all Disappeared (dissolved) ± Slightly invaded Mostly dissolved + Small scale invaded Specimen cells remain, but there is a significant morphological change.Moderate invasion Small-scale shrinkage/dissolution zones noticeably invade From the morphological unchanged Table 1, cells in the matrix mixed with HAC significantly invaded compared to the matrix containing AC alone. I found out that my sexuality improved. However, from the viewpoint of maintaining the shape of the sponge, it is preferable that the weight percent of HAC is less than 80%.
比較例 ２ 線維化アテロコラーゲンの調製アテロコ
ラーゲン１．０ｇをｐ＋＋３．ｏの希塩酸に溶解して０
．３ｗ／ｖ％にした。この溶液を４℃の恒温槽に入れ撹
拌しなから、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、３０ｍ Ｍりん酸−２−す
トリウム、 １００ｍＭ ＮａＣｆ１であるコラーゲ
ン溶液を調製した。ついて３７℃の恒温槽に１日浸漬し
、線維化コラ−ケン（Ｆ Ｃ）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（Ｆ
Ｃ）溶液を調製した。この溶液を一３０°Ｃで急速凍結
した後、凍結乾燥を行ないスポンジを作製した。その後
このスポンジを真空下１１０℃、２時間処理し熱脱水架
橋した。Comparative Example 2 Preparation of fibrotic atelocollagen 1.0 g of atelocollagen was mixed with p++3. Dissolved in dilute hydrochloric acid of 0
．． It was set to 3w/v%. This solution was placed in a constant temperature bath at 4°C and stirred, then phosphate buffer was added to bring the final concentration to 0.1% (
A collagen solution was prepared: w/v) atelocollagen, 30mM 2-storium phosphate, 100mM NaCf1. Then, it was immersed in a constant temperature bath at 37°C for one day to obtain a fibrous collagen (FC) solution. This solution was centrifuged (5000 rpm, 1.0 min) to concentrate and 0.3% (W/V) fibrotic atelocollagen (F
C) A solution was prepared. This solution was quickly frozen at -30°C and then freeze-dried to produce a sponge. Thereafter, this sponge was treated under vacuum at 110° C. for 2 hours to carry out thermal dehydration crosslinking.
実施例 ２ 線維化アテロコラーゲン−変性アテロコ
ラーゲンマトリックスの
調製
上記で調製した０．３％（ｗ／ｖ）線維化アテロコラー
ゲン（Ｆ Ｃ）と１％（ｗ／ｖ）変性アテロコラ−ケン
（ＨＡＣ）を３７℃で混合し、１時間撹拌した。Example 2 Preparation of fibrotic atelocollagen-denatured atelocollagen matrix 0.3% (w/v) fibrotic atelocollagen (FC) and 1% (w/v) denatured atelocollagen (HAC) prepared above were incubated at 37°C. and stirred for 1 hour.
この溶液を一３０℃で急速凍結した後、凍結乾燥を行な
いスポンジを作製した。その後、このスポンジを真空下
１１０°Ｃ，２時間処理し、熱脱水架橋した。This solution was quickly frozen at -30°C and then freeze-dried to produce a sponge. Thereafter, this sponge was treated under vacuum at 110° C. for 2 hours to carry out thermal dehydration crosslinking.
試験例 ２ 線維化アテロコラーゲン−変性アテロコ
ラーゲンマトリックスの
ｉｎ ｖｔｔｒｏ細胞侵入性試験
上記実施例２、および比較例２で得られたマトリックス
について、試験例１と同様の操作でラットの線維芽細胞
を用いてｉｎ ｖ＋ｔｒｏで培養実験を行ない、細胞侵
入性を評価した。Test Example 2 In vitro cell invasiveness test of fibrotic atelocollagen-denatured atelocollagen matrix The matrices obtained in Example 2 and Comparative Example 2 were tested in vitro using rat fibroblasts in the same manner as in Test Example 1. A culture experiment was conducted using v+tro to evaluate cell invasiveness.
評価の結果を表−２に示した。The evaluation results are shown in Table-2.
（以下余白）
表 −２
コラーゲンマトリックスへの
ｉｎ ｖｉｔｒｏ細胞侵入性試験
表−２から、ＦＣを基材とするマトリックスは、全てス
ポンジの形態維持が良く、安定性に優れていた。細胞の
侵入では、ＦＣ単独でも若干の偏在的細胞侵入が見られ
たものの、ＨＡＣ添加系では非常に多くの細胞が、しか
も均一に分散して侵入しており、スポンジの形状もｉｎ
ｖｉｔｒｏ培養実験系でありながらｉｎ ｖｉｖｏの
生体組織に近い様相を呈していた。(The following is a blank space) Table 2 In vitro cell penetration test into collagen matrix From Table 2, all the matrices based on FC had good retention of sponge form and excellent stability. Regarding cell invasion, although some unevenly distributed cell invasion was observed with FC alone, in the HAC addition system, a large number of cells invaded in a uniformly dispersed manner, and the shape of the sponge also changed.
Although it was an in vitro culture experiment system, it resembled an in vivo biological tissue.
試験例 ３ 線維化アテロコラーゲン−変性アテロコ
ラーゲンマトリックスの
ｉｎ ｖｉｖｏ皮下埋入試験
実施例２、および比較例２で作製したマトリツクスをラ
ット皮下に埋入し、病理学的に組織像を検索する。Test Example 3 In vivo subcutaneous implantation test of fibrotic atelocollagen-denatured atelocollagen matrix The matrices prepared in Example 2 and Comparative Example 2 are implanted subcutaneously in rats, and the histological images are examined pathologically.
皮下埋植（埋入）には、約２００ｇのＷｉＳｔａｒ −
ＫＹ系、雌性ラットを用いる。埋入前に、５倍希釈ネン
ブタールで麻酔後ラットの背面を手術用のイソジン液（
明治製菓■製）で漏らし、毛刈り用カミソリで毛の刈り
残しがないように、背面を注意深く剃毛する。その後、
剃られた背面をイソジンとエタノールで消毒する。各々
の切り込みから、ラットの皮部下の疎性結合鉱内に空隙
を作るように切り込みを広げる（ただし、隣接する切り
込み同志は連絡しないように配慮する。）。この空隙に
検体をさし込み、検体全体か平らに横たわるようにする
。角針付六イロン糸で切り口を縫合する。Approximately 200 g of WiStar − for subcutaneous implantation (implantation)
KY strain female rats are used. Before implantation, the back of the rat was anesthetized with 5-fold diluted Nembutal and surgical Isodine solution (
Use a hair clipper (made by Meiji Seika ■) and carefully shave the back of the hair using a razor to make sure no hair is left uncut. after that,
Disinfect the shaved back with isodine and ethanol. From each incision, widen the incision so as to create a void in the loosely bound ore under the rat's skin (however, take care not to contact adjacent incisions). Insert the specimen into this gap so that the entire specimen lies flat. Suture the incision with hexagonal thread with a square needle.
切り口は３針縫う。同じ検体を別のラットにも同様にし
て埋入する。Stitch the incision with 3 stitches. The same specimen is implanted in another rat in the same manner.
埋入後３，２８日目に動物をエーテルあるいは２倍希釈
ネンブタールを用いて殺す。埋入検体が組織中に留まっ
ているようにして、ラットの背筋」二の皮膚組織を８
ｃｍ Ｘ １２ｃｍあるいはそれ以上の太きさに切り取
る。この組織を１０％中性緩衝ホルマリン溶液中に置き
、−昼夜放置し固定後、病理組織検索を施す。Animals are sacrificed using ether or 2x diluted Nembutal on days 3 and 28 after implantation. The skin tissue on the rat's back muscle was removed by 8 minutes, making sure that the implanted specimen remained in the tissue.
Cut into pieces 12cm x 12cm or thicker. This tissue is placed in a 10% neutral buffered formalin solution, left to stand day and night for fixation, and then subjected to pathological examination.
病理組織検索は組織からの検体の切り出しに始まる。検
体が確実に含まれるように、組織を０，５ｃｍＸ２．５
ｃｍ程度のたんさく状に切り出す。これをエタノール、
次にキンレンで透徹し、最後にパラフィンに置換する。Pathological tissue retrieval begins with cutting out a specimen from tissue. To ensure that the specimen is included, the tissue is placed in a 0.5 cm x 2.5
Cut into strips about cm in size. This is ethanol,
Next, it is cleared with phlegm, and finally it is replaced with paraffin.
置換後、固型パラフィンの加熱溶解液に、検体を含む組
織を置き、急冷してパラフィン包埋を完了する。包埋さ
れた組織は、ヤマト■製回転式ミクロトームにて薄切を
行ない、厚さ４μｍのパラフィン切片とする。これを脱
パラフィンした後、任意の染色法で病理組織染色を行な
い、プレパラートを完成する。病理組織染色として、ヘ
マトキシリン−エオシン（Ｈ−Ｅ）染色、アサン染色、
レゾルシン−フクンン染色等を採用できる。結果を表−
３に示した。After replacement, the tissue containing the specimen is placed in a heated solution of solid paraffin and rapidly cooled to complete paraffin embedding. The embedded tissue was sliced into paraffin sections with a thickness of 4 μm using a rotary microtome manufactured by Yamato ■. After deparaffinizing this, pathological tissue staining is performed using an arbitrary staining method to complete the preparation. As pathological tissue staining, hematoxylin-eosin (H-E) staining, Asan staining,
Resorcinol-Fukunun staining etc. can be employed. Display the results -
Shown in 3.
（以下余白）
表 −３
皮下埋入試験の病理組織変化
ＦＣだけでは、３日日においては好中球浸潤が強くかつ
線維芽細胞の侵入は中）度であり、２８［］目において
は、出来上かった肉芽組織が萎縮している。それに対し
、ＨＡＣが１０ないし２０重量％はいる事により３日日
における好中球浸潤は弱く、逆に線維等細胞侵入は、−
層良好となる。さらに２８日日日おける肉芽組織の萎縮
も著しく緩和される事が明らかである。(Margins below) Table 3: Pathological Histopathological Changes in Subcutaneous Implant Test In FC alone, neutrophil infiltration was strong and fibroblast infiltration was moderate at day 3, and at day 28, The resulting granulation tissue is atrophied. On the other hand, with 10 to 20% by weight of HAC, neutrophil infiltration on day 3 was weak, and conversely, fibrous cell infiltration was -
The layer becomes better. Furthermore, it is clear that the atrophy of the granulation tissue after 28 days is also significantly alleviated.
実施例 ３ 変性コラーゲンを被覆した架橋コラーゲ
ンの調製
比較例２で得た線維化アテロコラーゲン（ＦＣ）凍結乾
燥スポンジを０．０１％および１％へキサメチレンージ
ーイソシアネート（ＨＤＩ）＝エタノール溶液に一昼夜
浸漬し、化学架橋を導入した。それぞれのスポンジに実
施例コ−で得られた変性コラーゲン（ＨＡＣ）水溶液を
３０ｍ１添加し、十分浸漬後再び凍結乾燥してスポンジ
化１７、それぞれを真空下、１１０℃で２時間加熱処理
、および２４時間加熱処理を施し、熱脱水架橋を導入し
た。こうして、変性コラーゲン（ＨＡＣ）を被覆したコ
ラーゲンマトリックスを得た。最終的な組成化はＨＡＣ
が１０重量％となるようにした。Example 3 Preparation of crosslinked collagen coated with denatured collagen The fibrotic atelocollagen (FC) freeze-dried sponge obtained in Comparative Example 2 was immersed in 0.01% and 1% hexamethylene diisocyanate (HDI) = ethanol solutions overnight. Then, chemical crosslinking was introduced. 30 ml of the denatured collagen (HAC) aqueous solution obtained in Example 1 was added to each sponge, and after thorough immersion, freeze-drying was performed again to form a sponge. Heat treatment was performed for a period of time to introduce thermal dehydration crosslinking. In this way, a collagen matrix coated with denatured collagen (HAC) was obtained. The final composition is HAC
was adjusted to be 10% by weight.
比較例 ３ 架橋コラーゲンの調製
実施例３のうぢ、変性コラーゲン（ＨＡＣ）水溶液の添
加の過程を省いた、単独の線維化アテロコラーゲン（Ｆ
Ｃ）のみの凍結乾燥スポンジ（架橋の導入は実施例３
と同一）を比較例３として用意した。Comparative Example 3 Preparation of Crosslinked Collagen In Example 3, only fibrotic atelocollagen (F
C) only freeze-dried sponge (crosslinking was introduced in Example 3)
) was prepared as Comparative Example 3.
試験例 ４ 変性コラ−ケン被覆架橋コラーゲンマト
リックスのｉｎνｉｖｏ皮下
埋入試験
実施例３および比較例３で作製したマトリックスを、試
験例３の手法に準じてラット皮下に埋入し、病理・組織
学的検索に付す。但し、試料は７日後、１４日後に取り
出した。結果を表−４に示した。Test Example 4 Invivo Subcutaneous Implantation Test of Modified Kolaken Coated Crosslinked Collagen Matrix The matrices prepared in Example 3 and Comparative Example 3 were implanted subcutaneously in rats according to the method of Test Example 3, and pathological and histological tests were performed. Add to search. However, the samples were taken out after 7 days and 14 days. The results are shown in Table-4.
（以下余白）
それぞれ比較例として置いたＦＣは、一部好中球等の浸
潤もあるものの、細胞成分自体の浸潤が、炎症性および
網内系細胞も含め、極めて悪い。それに比し２て、それ
ぞれＨＡ Ｃを被覆したＦＣは、細胞成分の浸潤が甚だ
良好で、それに伴って一部自己組織化も行な４つねてお
り、中には異物反応がやや強いものもあるものの、特に
ＨＡＣ被覆ＦＣＯ旧％ＨＤＩ架橋」−熱架橋２時間の試
料に至っては好中球浸潤すら既に少ない極めて真皮に近
い構造を試料の部位に再構築しており、当発明の１」的
等を考えても最も理想に近いマトリックスであると言え
る。(Margins below) The FCs used as comparative examples have some infiltration of neutrophils, but the infiltration of cellular components themselves, including inflammatory and reticuloendothelial cells, is extremely poor. In contrast, FCs coated with HAC showed extremely good infiltration of cellular components and also partially self-organized, and some of them had a slightly stronger foreign body reaction. However, especially in the case of the HAC-coated FCO former %HDI cross-linked sample, which was thermally cross-linked for 2 hours, a structure very similar to the dermis with even little neutrophil infiltration was reconstructed at the sample site, which is one of the features of the present invention. It can be said that this is the matrix that is closest to the ideal considering the target and other factors.
実施例 ４ シリコーン膜含有コラーゲンスポンジの
調製
テフロン上に５０％５ｉｌａｓｔｉｃシリコ一ン接着剤
型Ａ （Ｄｏｗ Ｃｏｒｎｉｎｇ社製）のヘキサン溶液
を精密被覆用具（アプリケーター）を用いて塗布し製膜
した。塗布した直後に実施例３によって製造したスポン
ジをのせ、室温で１ｏ分程放置した後、Ｂ Ｏ’Ｃで少
なくとも１時間オーブンで硬化させた。Example 4 Preparation of collagen sponge containing silicone film A film was formed by applying a hexane solution of 50% 5ilastic silicone adhesive type A (manufactured by Dow Corning) onto Teflon using a precision coating tool (applicator). Immediately after coating, the sponge prepared according to Example 3 was placed on it, left at room temperature for about 10 minutes, and then cured in an oven at B O'C for at least 1 hour.
試験例 ５ 皮膚欠損創−・の移植試験実施例４によ
り製造したスポンジを使用して、ラットの皮膚欠損創へ
の移植試験を行なった。Test Example 5 Transplantation Test on Skin Defect Wounds Using the sponge prepared in Example 4, a transplantation test on skin defect wounds in rats was conducted.
ラット背部皮膚に皮下筋膜を創面とする全創皮膚欠損創
（２ｃｍＸ２ｃｍ）を作製し、シリコーン膜を表層に（
＝Ｊ与した検体を結紮縫合した。動物は移植後４週「ｊ
に殺し、移植物と傷床を切り取り、病理検索を施した。A total skin defect wound (2 cm x 2 cm) was created on the rat's back skin with the subcutaneous fascia as the wound surface, and a silicone membrane was placed on the surface layer (
=J The given specimen was ligated and sutured. Animals were harvested 4 weeks after transplantation.
The animal was sacrificed, the graft and wound bed were excised, and a pathological examination was performed.
４週１」では創収縮はあまり見られず、良好な肉芽組織
が形成［７、表皮再生が見られた。At 4 weeks 1'', there was not much wound contraction, good granulation tissue was formed [7, and epidermal regeneration was observed.
［発明の効果］
本発明の医用材料は、担体にヘリックス含量が０〜８０
％である変性コラーゲンを結合または被覆したものから
なるため、４１；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かできるとともに
、生体適合性に優れ、その内部に増殖した細胞か容易に
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 helical content of 0 to 80 in the carrier.
41%, it is resistant to collagenase when implanted in the body or coated on a wound surface, and has the required mechanical strength for a certain period of time. It has excellent biocompatibility, and cells that have proliferated inside can easily enter it. Medical materials obtained using atelocollagen as a raw material are particularly desirable because they do not have antigenicity.
従って本発明の医用材料は埋入型人工臓器例えば生体内
留置人工心臓、人工血管等や深度熱傷時の人工被覆材と
して利用される。Therefore, the medical material of the present invention can be used as an implantable artificial organ such as an indwelling artificial heart, an artificial blood vessel, etc., or as an artificial covering material for deep burns.
Claims (1)
ーゲンを結合または被覆したことを特徴とする細胞侵入
性医用材料。 ２）担体が生体吸収材料である請求項１の医用材料。 ３）生体吸収材料がコラーゲンである請求項１または２
の医用材料。 ４）コラーゲンが熱脱水架橋あるいは化学架橋されてい
る請求項１〜３のいずれかの項に記載の医用材料。 ５）コラーゲンおよびヘリックス含量が０〜８０％であ
る変性コラーゲンを混合し、フィルムまたは多孔体を形
成させた後架橋されたことを特徴とする細胞侵入性医用
材料。 ６）架橋されたコラーゲンフィルムあるいは多孔体をヘ
リックス含量０〜８０％である変性コラーゲン溶液で被
覆したことを特徴とする細胞侵入性医用材料。[Scope of Claims] 1) A cell-invasive medical material characterized in that a carrier is bound or coated with denatured collagen having a helical content of 0 to 80%. 2) The medical material according to claim 1, wherein the carrier is a bioabsorbable material. 3) Claim 1 or 2, wherein the bioabsorbable material is collagen.
medical materials. 4) The medical material according to any one of claims 1 to 3, wherein the collagen is thermally dehydrated crosslinked or chemically crosslinked. 5) A cell-invasive medical material characterized in that collagen and denatured collagen having a helix content of 0 to 80% are mixed to form a film or porous body, which is then crosslinked. 6) A cell-invasive medical material characterized by coating a crosslinked collagen film or porous body with a denatured collagen solution having a helix content of 0 to 80%.
Priority Applications (6)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63053837A JPH0622579B2 (en) | 1988-03-09 | 1988-03-09 | Cell invasive medical materials |
| AU32126/89A AU632273B2 (en) | 1988-03-09 | 1989-03-09 | Medical material permitting cells to enter thereinto and artificial skin |
| DE68915540T DE68915540T2 (en) | 1988-03-09 | 1989-03-09 | FOR CELL-PULLABLE MEDICAL MATERIAL AND ARTIFICIAL SKIN. |
| EP89903232A EP0403650B1 (en) | 1988-03-09 | 1989-03-09 | Medical material permitting cells to enter thereinto and artificial skin |
| PCT/JP1989/000257 WO1989008465A1 (en) | 1988-03-09 | 1989-03-09 | Medical material permitting cells to enter thereinto and artificial skin |
| US07/576,493 US5263983A (en) | 1988-03-09 | 1989-03-09 | Medical material and prosthetic skin in which cells can invade |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63053837A JPH0622579B2 (en) | 1988-03-09 | 1988-03-09 | Cell invasive medical materials |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH01230366A true JPH01230366A (en) | 1989-09-13 |
| JPH0622579B2 JPH0622579B2 (en) | 1994-03-30 |
Family
ID=12953898
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63053837A Expired - Lifetime JPH0622579B2 (en) | 1988-03-09 | 1988-03-09 | Cell invasive medical materials |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0622579B2 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPWO2003094985A1 (en) * | 2002-05-14 | 2005-09-08 | 北海道ティー・エル・オー株式会社 | Artificial extracellular matrix and method for producing the same |
| US8372433B2 (en) | 2007-01-18 | 2013-02-12 | Gunze Limited | Substrate for culture of cardiovascular tissue |
| US8748142B2 (en) | 1999-09-09 | 2014-06-10 | Gunze Limited | Culture of cardiovascular cells on a matrix and method for regenerating cardiovascular tissue |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP6100702B2 (en) | 2012-01-12 | 2017-03-22 | 株式会社ニッピ | Collagen structure and method for producing collagen structure |
Citations (7)
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|---|---|---|---|---|
| JPS5230885A (en) * | 1975-07-15 | 1977-03-08 | Massachusetts Inst Technology | Crosslinked polymer consisting of collagen and mucopolysaccharide |
| JPS5558163A (en) * | 1978-10-24 | 1980-04-30 | Unitika Ltd | Material for treating wounded portion |
| JPS61191364A (en) * | 1985-02-21 | 1986-08-26 | 工業技術院長 | Anti-thrombotic material |
| JPS6226230A (en) * | 1985-07-25 | 1987-02-04 | Koken:Kk | Crosslinked medical material |
| JPS62127056A (en) * | 1985-11-27 | 1987-06-09 | 宇部興産株式会社 | Wound cover material |
| JPS62194854A (en) * | 1985-11-19 | 1987-08-27 | メリドン・サ−ビセズ・プロプライエタリ−・リミテツド | Biocompatible long-term biodegradable reticulated cloth |
| JPS62157524U (en) * | 1986-03-27 | 1987-10-06 |
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1988
- 1988-03-09 JP JP63053837A patent/JPH0622579B2/en not_active Expired - Lifetime
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5230885A (en) * | 1975-07-15 | 1977-03-08 | Massachusetts Inst Technology | Crosslinked polymer consisting of collagen and mucopolysaccharide |
| JPS5558163A (en) * | 1978-10-24 | 1980-04-30 | Unitika Ltd | Material for treating wounded portion |
| JPS61191364A (en) * | 1985-02-21 | 1986-08-26 | 工業技術院長 | Anti-thrombotic material |
| JPS6226230A (en) * | 1985-07-25 | 1987-02-04 | Koken:Kk | Crosslinked medical material |
| JPS62194854A (en) * | 1985-11-19 | 1987-08-27 | メリドン・サ−ビセズ・プロプライエタリ−・リミテツド | Biocompatible long-term biodegradable reticulated cloth |
| JPS62127056A (en) * | 1985-11-27 | 1987-06-09 | 宇部興産株式会社 | Wound cover material |
| JPS62157524U (en) * | 1986-03-27 | 1987-10-06 |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8748142B2 (en) | 1999-09-09 | 2014-06-10 | Gunze Limited | Culture of cardiovascular cells on a matrix and method for regenerating cardiovascular tissue |
| JPWO2003094985A1 (en) * | 2002-05-14 | 2005-09-08 | 北海道ティー・エル・オー株式会社 | Artificial extracellular matrix and method for producing the same |
| US8372433B2 (en) | 2007-01-18 | 2013-02-12 | Gunze Limited | Substrate for culture of cardiovascular tissue |
Also Published As
| Publication number | Publication date |
|---|---|
| JPH0622579B2 (en) | 1994-03-30 |
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