JPH0827020A - Osteoclast formation and bone resorption promoter - Google Patents
Osteoclast formation and bone resorption promoterInfo
- Publication number
- JPH0827020A JPH0827020A JP6166917A JP16691794A JPH0827020A JP H0827020 A JPH0827020 A JP H0827020A JP 6166917 A JP6166917 A JP 6166917A JP 16691794 A JP16691794 A JP 16691794A JP H0827020 A JPH0827020 A JP H0827020A
- Authority
- JP
- Japan
- Prior art keywords
- growth factor
- bone resorption
- promoting
- amino acid
- seq
- 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.)
- Pending
Links
Landscapes
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Peptides Or Proteins (AREA)
Abstract
Description
【０００１】[0001]
【産業上の利用分野】本発明は、軟骨細胞増殖因子を有
効性分とする破骨細胞形成促進及び骨吸収促進剤に関す
る。FIELD OF THE INVENTION The present invention relates to an agent for promoting osteoclast formation and bone resorption, which contains chondrocyte growth factor as an effective component.
【０００２】[0002]
【従来の技術及び発明が解決しようとする課題】破骨細
胞は、造血幹細胞由来の単核細胞に由来し、血行を介し
て骨表面に運ばれた細胞が分化して形成される細胞であ
る。一方、骨芽細胞は間葉由来の未分化細胞や線維芽細
胞、間質細胞などの基質形成細胞系に属する前駆細胞か
ら分化し、骨基質を形成する。このように、破骨細胞は
骨芽細胞とは系列の異なる前駆細胞に由来する。その特
徴としては、終末分化して多核となった増殖能の乏しい
巨細胞であり、多数のカルシトニン受容体ならびに酒石
酸耐性酸性ホスファターゼ活性を発現することが挙げら
れる。また、破骨細胞は骨や象牙質などの石灰化組織を
吸収することが出来る。BACKGROUND OF THE INVENTION Osteoclasts are cells derived from hematopoietic stem cell-derived mononuclear cells, which are formed by differentiation of cells carried to the bone surface through blood circulation. . On the other hand, osteoblasts differentiate from progenitor cells belonging to a matrix-forming cell line such as mesenchymal-derived undifferentiated cells, fibroblasts, and stromal cells to form bone matrix. Thus, osteoclasts are derived from progenitor cells of a different lineage than osteoblasts. Its characteristic is that they are giant cells with poor proliferative ability, which are terminally differentiated and become multinucleated, and express a large number of calcitonin receptors and tartrate-resistant acid phosphatase activity. Further, osteoclasts can absorb calcified tissues such as bone and dentin.
【０００３】破骨細胞の形成に関しては、その前駆細胞
である造血幹細胞由来の単核細胞をマクロファージ・コ
ロニー刺激因子（M-CSF）や顆粒球・マクロファージ・
コロニー刺激因子（GM-CSF）が増殖させ、さらに、骨芽
細胞様の間質細胞との細胞接触により破骨細胞へと分化
すると考えられている。破骨細胞の活性化には、1a,25
(OH)2 ビタミン D3や副甲状腺ホルモン（PTH）、副甲状
腺ホルモン関連ペプチド（PTHrP）、各種プロスタグラ
ンジンおよびインターロイキン - 1の様な骨吸収促進因
子の添加が効果的である。しかしながら、骨芽細胞およ
びその類縁細胞の培養上清や高カルシウム血症を惹起す
る腫瘍細胞の培養上清等から破骨細胞分化因子あるいは
破骨細胞活性化因子の様な内因性の蛋白及びペプチド性
因子は未だ同定されていない。Regarding the formation of osteoclasts, mononuclear cells derived from hematopoietic stem cells, which are their progenitor cells, are treated with macrophages / colony stimulating factor (M-CSF) or granulocytes / macrophages /
It is believed that colony stimulating factor (GM-CSF) proliferates and further differentiates into osteoclasts by cell contact with osteoblast-like stromal cells. Activation of osteoclasts requires 1a, 25
Addition of bone resorption promoting factors such as (OH) 2 vitamin D 3 and parathyroid hormone (PTH), parathyroid hormone-related peptide (PTHrP), various prostaglandins and interleukin-1 is effective. However, endogenous proteins and peptides such as osteoclast differentiation factors or osteoclast activating factors are found in culture supernatants of osteoblasts and their related cells, culture supernatants of tumor cells that cause hypercalcemia, etc. The sex factor has not yet been identified.
【０００４】破骨細胞は骨のモデリング等、骨代謝に於
て重要な機能を有する細胞であり、骨粗鬆症等の代謝性
骨疾患に直接関与すると考えられている。しかし、その
細胞起源、形成機序等に関しては未だ不明な点が多く、
骨代謝及び疾患への重要な関与は予想されているが、そ
の機構解明には至っていない。破骨細胞の分化あるいは
活性化に関与する因子の一つを同定したことは、その形
成機序及び骨代謝への関与を明らかにする指針を得るこ
とにつながり、代謝性骨疾患治療の確立を可能にすると
考えられていた。Osteoclasts are cells having important functions in bone metabolism such as bone modeling, and are considered to be directly involved in metabolic bone diseases such as osteoporosis. However, there are still many unclear points regarding its cell origin, formation mechanism, etc.
Although important involvement in bone metabolism and diseases is expected, its mechanism has not yet been elucidated. The identification of one of the factors involved in osteoclast differentiation or activation leads to the elucidation of its formation mechanism and its involvement in bone metabolism, thus establishing the treatment of metabolic bone diseases. It was thought to be possible.
【０００５】[0005]
【課題を解決するための手段】本発明者らは、破骨細胞
形成促進能または破骨細胞活性化能を有する因子を得る
べく鋭意検討を重ねた結果、意外にも、先に本発明者ら
軟骨細胞増殖因子として同定した特開平５−２５５３９
８号公報に記載の分子量が約１６ＫＤａのタンパク質
（以下、「コンドロモジュリン−ＩＩ」と称することも
ある）が、破骨細胞形成促進活性及び骨吸収促進活性を
有することを見出し本発明を完成するに至った。Means for Solving the Problems As a result of intensive investigations by the present inventors to obtain a factor having an osteoclast formation-promoting ability or an osteoclast activation ability, surprisingly, the present inventors Identified as chondrocyte growth factor
It was found that the protein having a molecular weight of about 16 KDa (hereinafter, also referred to as "chondromodulin-II") described in Japanese Patent Publication No. 8 has osteoclast formation-promoting activity and bone resorption-promoting activity, thus completing the present invention. Came to do.
【０００６】即ち、本発明の要旨は、下記の理化学的性
質を有する軟骨細胞増殖因子を有効成分とする破骨細胞
形成促進及び骨吸収促進剤に存する。 ＳＤＳ−ポリアクリルアミドゲル電気泳動による分子
量が約１６，０００ドルトンである。 軟骨細胞を単独でまたは繊維芽細胞増殖因子共存下で
増殖させる活性を有する。[0006] That is, the gist of the present invention resides in an osteoclast formation promoting agent and a bone resorption promoting agent containing a chondrocyte growth factor having the following physicochemical properties as an active ingredient. The molecular weight by SDS-polyacrylamide gel electrophoresis is about 16,000 daltons. It has the activity of growing chondrocytes alone or in the coexistence of fibroblast growth factor.
【０００７】軟骨細胞の分化機能を促進させる活性を
有する。 以下、本発明をさらに詳細に説明するに、本発明の軟骨
細胞の増殖及び分化機能の誘導を促す作用を有する新規
なタンパク質は、特開平５−２５５３９８号公報に記載
のように、例えば、牛胎児軟骨を破砕し、これを遠心分
離した上清を限外濾過膜により分画、濃縮したものを、
Ｓｅｐｈａｃｒｙ１ Ｓ２００カラム（ファルマシア社
製）等による分子ふるいクロマトグラフィーによりさら
に分画したのち、ヘパリン−Ｔｏｙｏｐｅａｒｌアフィ
ニティーカラム（トーソー社製）に吸着させ、例えば
０．５ＭＮａＣｌを含む緩衝液で溶出し、次いで、ＹＭ
Ｃｐａｃｋ Ｃ８カラムクロマトグラフィー（ワイエム
シー社製）等により溶出条件を変えて繰り返し精製する
ことによって得られる。精製された蛋白質はＳＤＳ−ポ
リアクリルアミドゲル電気泳動による分子量が約１６，
０００ドルトンであり、軟骨細胞を単独または繊維芽細
胞増殖因子共存下で増殖させる活性及び軟骨細胞の分化
機能を促進する活性を有する。It has an activity of promoting the differentiation function of chondrocytes. Hereinafter, the present invention will be described in more detail. A novel protein having an action of promoting the induction of proliferation and differentiation functions of chondrocytes of the present invention is, for example, as described in JP-A-5-255398, Fetal cartilage was crushed, and the supernatant obtained by centrifuging this was fractionated and concentrated by an ultrafiltration membrane,
After further fractionation by molecular sieve chromatography using Sephacry1 S200 column (manufactured by Pharmacia), etc., it is adsorbed on a heparin-Toyopearl affinity column (manufactured by Tosoh) and eluted with a buffer containing 0.5 M NaCl, and then YM.
It can be obtained by repeatedly purifying by changing elution conditions by Cpack C8 column chromatography (manufactured by YMC) or the like. The purified protein has a molecular weight of about 16 by SDS-polyacrylamide gel electrophoresis,
It is 000 Dalton, and has the activity of proliferating chondrocytes alone or in the coexistence of fibroblast growth factor and the activity of promoting the differentiation function of chondrocytes.
【０００８】またコンドロモデュリン−IIは、配列表の
配列番号：１に示すようなＮ末端アミノ酸配列及び配列
表の配列番号：２、３及び４に示すような部分アミノ酸
配列を含む。より好ましくは、さらに部分アミノ酸配列
として配列表の配列番号：５から１４に記載のアミノ酸
配列で含むものが挙げられる。なお、軟骨細胞の増殖活
性及び分化機能の促進活性を損なわない範囲でかかるア
ミノ酸の一部を除去、置換、修飾または追加するなどの
改変を行ったものも本発明のコンドロモジュリン−ＩＩ
に含まれる。[0008] Chondromodulin-II contains an N-terminal amino acid sequence as shown in SEQ ID NO: 1 of the sequence listing and a partial amino acid sequence as shown in SEQ ID NOS: 2, 3 and 4 of the sequence listing. More preferably, the partial amino acid sequence further includes the amino acid sequences described in SEQ ID NOs: 5 to 14 in the sequence listing. It should be noted that the chondromodulin-II of the present invention may also be one that has been modified by removing, substituting, modifying or adding a part of such amino acids within a range that does not impair the activity of promoting chondrocyte proliferation and the function of differentiation.
include.
【０００９】本発明で使用されるコンドロモジュリン−
ＩＩは、生物学的にコンドロモジュリン−ＩＩと同等の
活性を有するものであれば、いかなるものも使用するこ
とができる。ヒト患者の治療を考慮した場合、ヒト由来
のものであることが望ましいが、そのアミノ酸配列中に
他の動物組織由来のコンドロモジュリン−ＩＩの配列を
含むものであってもよい。即ち、活性の上昇、安定性の
上昇を目的として、Ｎ末端から数個のアミノ酸を欠失さ
せたものや、逆にＮ末端にいくつかのアミノ酸を付加す
ることも可能である。Chondromodulin used in the present invention
Any II can be used as long as it has biologically equivalent activity to chondromodulin-II. In consideration of the treatment of human patients, human-derived ones are preferable, but a chondromodulin-II sequence derived from another animal tissue may be contained in the amino acid sequence thereof. That is, for the purpose of enhancing activity and stability, it is possible to delete several amino acids from the N-terminal, or conversely add some amino acids to the N-terminal.
【００１０】本発明において、コンドロモジュリン−Ｉ
Ｉの破骨細胞形成促進活性及び骨吸収促進活性は、例え
ば次のようにして測定される。ウサギ大腿骨から分離し
た5%ウシ胎仔血清を含む骨細胞を、滅菌処理した象牙質
上に添加しその骨吸収活性を見るアッセイ系に、ウシ胎
仔軟骨から精製したコンドロモジュリン−ＩＩを0.1- 1
00 ng/mlの濃度で添加し、その骨吸収促進活性を検討し
た。その結果、0.1ng/ml ChM-II濃度で活性が検出さ
れ、3−5 ng/mlで最大活性を示した。In the present invention, chondromodulin-I
The osteoclast formation promoting activity and bone resorption promoting activity of I are measured, for example, as follows. Bone cells containing 5% fetal bovine serum isolated from rabbit femurs were added to sterilized dentin to observe its bone resorption activity.As an assay system, chondromodulin-II purified from fetal bovine cartilage was added to 0.1- 1
It was added at a concentration of 00 ng / ml, and its bone resorption promoting activity was examined. As a result, the activity was detected at a concentration of 0.1 ng / ml ChM-II, and the maximum activity was shown at 3-5 ng / ml.
【００１１】本発明の破骨細胞形成促進及び骨吸収剤
は、例えば骨形成が盛んであり、その成長を抑える必要
があるような疾患の場合には、コンドロモジュリン−Ｉ
Ｉをそのまま、あるいは常法により製剤化、医薬組成物
化して投与することができる。具体的には、コンドロモ
ジュリン−ＩＩ１ｎｇから１００μｇを軟骨疾患部位に
コラーゲン、アテロコラーゲン、ゼラチン、ヒアルロン
酸、ポリエチレングリコール、ポリ乳酸、骨セメント、
ハイドロキシアパタイト、セラミックス、炭素繊維、フ
ィブリン糊等の外科手術用生体接着剤等の生体適合性担
体に混合、含浸、塗布することにより局所的に外科手術
によって投与する。The osteoclast formation-promoting and bone-resorbing agent of the present invention is, for example, chondromodulin-I in the case of diseases in which osteogenesis is active and the growth thereof needs to be suppressed.
I can be administered as it is or in the form of a pharmaceutical preparation or pharmaceutical composition by a conventional method. Specifically, from 1 ng of chondromodulin-II to 100 μg in a cartilage disease site, collagen, atelocollagen, gelatin, hyaluronic acid, polyethylene glycol, polylactic acid, bone cement,
It is locally administered by surgery by mixing, impregnating and applying it on a biocompatible carrier such as bioadhesive for surgery such as hydroxyapatite, ceramics, carbon fiber, and fibrin glue.
【００１２】また本発明の破骨細胞形成促進および骨吸
収剤は、新たな骨疾患治療剤を探索する上においても有
用である。すなわち同薬剤に対して阻害作用を有する物
質は、破骨細胞の形成を抑制し、骨吸収を抑制する効果
があると考えられる。従って適当な評価系を用いてコン
ドロモジュリン−ＩＩの生物活性を阻害する物質をスク
リーニングすることにより、骨粗鬆症等の骨疾患に対す
る薬剤が簡便に得られる。The osteoclast formation-promoting and bone-resorbing agent of the present invention is also useful in searching for a new therapeutic agent for bone diseases. That is, it is considered that a substance having an inhibitory effect on the same drug has an effect of suppressing the formation of osteoclasts and suppressing bone resorption. Therefore, by screening a substance that inhibits the biological activity of chondromodulin-II using an appropriate evaluation system, a drug for bone diseases such as osteoporosis can be easily obtained.
【００１３】[0013]
【発明の効果】本発明により、コンドロモジュリン−Ｉ
Ｉが破骨細胞に対してその分化あるいは骨吸収活性を促
進することが判明した。本発明によれば、現在まで有効
な治療法のなかった代謝性骨疾患に対して有効な治療法
を、さらには骨疾患に対する治療薬の新しいスクリーニ
ング法を提供することができる。According to the present invention, chondromodulin-I
It was found that I promotes the differentiation or bone resorption activity of osteoclasts. INDUSTRIAL APPLICABILITY According to the present invention, it is possible to provide an effective therapeutic method for a metabolic bone disease, which has not been effective until now, and a new screening method for a therapeutic drug for a bone disease.
【００１４】[0014]
【実施例】以下の実施例により、本発明をさらにより詳
細に説明するが、本発明は、その要旨を越えない限り以
下の実施例によって限定されるものではない。 実施例１ 150mm厚の象牙質スライスを直径6mmの円盤状に切断し、
70%アルコール中で超音波処理することにより滅菌し
た。a-MEMで洗浄した後、各スライスを96ウェルプレー
トのウェル中に移し、5%ウシ胎仔血清（FBS）と別に調
製したウサギ骨細胞1.25 x 105個を含むa-MEM 250mlを
その上に添加し、CO2インキュベーター中（5%CO2、95%
空気）37℃で2時間インキュベートした。2時間後、5%FB
Sと各濃度のコンドロモジュリン−ＩＩを含むa-MEM 100
mlを加えCO2インキュベーター中（10%CO2、90%空気）37
℃で15時間インキュベートした後、象牙質スライスから
ハンディーエンジンにより骨細胞を取り除き、酸性ヘマ
トキシリンにて3分間染色し、顕微鏡下、ピット当たり
のメッシュの数を勘定し破骨細胞の骨吸収活性とする。The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to the following examples without departing from the gist thereof. Example 1 A 150 mm thick dentin slice was cut into a disk shape having a diameter of 6 mm,
Sterilized by sonication in 70% alcohol. After washing with a-MEM, each slice was transferred into a well of a 96-well plate and 250 ml of a-MEM containing 1.25 x 10 5 separately prepared 5% fetal bovine serum (FBS) and rabbit bone cells was placed on it. was added, CO 2 incubator (5% CO 2, 95%
Incubated at 37 ° C for 2 hours. 2 hours later, 5% FB
A-MEM 100 containing S and various concentrations of chondromodulin-II
ml in a CO 2 incubator (10% CO 2 , 90% air) 37
After incubating at ℃ for 15 hours, remove the osteocyte from the dentin slice by handy engine, stain with acidic hematoxylin for 3 minutes, and count the number of meshes per pit under a microscope to determine the osteoclastic bone resorption activity. .
【００１５】図1に、本アッセイ系を用いてコンドロモ
ジュリン−ＩＩの骨吸収活性の検定を行った結果を示
す。図中、横軸はコンドロモジュリン−ＩＩの濃度を、
縦軸は象牙質スライス当たりのメッシュの数を示し、○
はコンドロモジュリン−ＩＩの結果を、△はビタミンＤ
３（ポジティブコントロール）の結果を表す。コンドロ
モジュリン−ＩＩ 0.1ng/mlから活性が検出され、濃度
依存的に活性の促進が観察され、3ng/mlで最大活性を示
した。この結果より、コンドロモジュリン−ＩＩが低能
度で破骨細胞の分化促進、あるいは活性化促進作用を有
することは明らかである。FIG. 1 shows the results of assaying the bone resorption activity of chondromodulin-II using this assay system. In the figure, the horizontal axis represents the concentration of chondromodulin-II,
The vertical axis shows the number of meshes per dentin slice, ○
Indicates the result of chondromodulin-II, △ indicates vitamin D
The result of 3 (positive control) is shown. The activity was detected from 0.1 ng / ml of chondromodulin-II, the promotion of the activity was observed in a concentration-dependent manner, and the maximum activity was shown at 3 ng / ml. From this result, it is clear that chondromodulin-II has a low-potency effect of promoting osteoclast differentiation or promoting osteoclast differentiation.
【００１６】[0016]
配列番号：１ 配列の長さ：４１ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：Ｎ末端フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Gly Pro Trp Ala Ile Ile Cys Ala Gly Lys １０ Ser Ser Asn Glu Ile Arg Thr Cys Asp Gly ２０ His Gly Cys Gly Gln Tyr Thr Ala Gln Arg ３０ Asn Gln Lys Leu His Gln Gly Val Asp Val ４０ Leu ４１ SEQ ID NO: 1 Sequence length: 41 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: N-terminal fragment Origin Biological name: Bovine Tissue type: Fetal cartilage Sequence Gly Pro Trp Ala Ile Ile Cys Ala Gly Lys 10 Ser Ser Asn Glu Ile Arg Thr Cys Asp Gly 20 His Gly Cys Gly Gln Tyr Thr Ala Gln Arg 30 Asn Gln Lys Leu His Gln Gly Val Asp Val 40 Leu 41
【００１７】配列番号：２ 配列の長さ：９ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Asn Ala Ile Asn Asn Gly Val Arg Ile ９SEQ ID NO: 2 Sequence Length: 9 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Asn Ala Ile Asn Asn Gly Val Arg Ile 9
【００１８】配列番号：３ 配列の長さ：１９ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Leu His Gln Gly Val Asp Val Leu Cys Ser １０ Asp Gly Ser Thr Val Tyr Ala Pro Phe １９SEQ ID NO: 3 Sequence Length: 19 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Leu His Gln Gly Val Asp Val Leu Cys Ser 10 Asp Gly Ser Thr Val Tyr Ala Pro Phe 19
【００１９】配列番号：４ 配列の長さ：１８ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Val Tyr Pro Gly Ile Gln Ser His Ile His １０ Ile Glu Asn Cys Asp Leu Ser Asp １８SEQ ID NO: 4 Sequence length: 18 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: Intermediate fragment Origin Biological name: Bovine tissue type: Fetal cartilage sequence Val Tyr Pro Gly Ile Gln Ser His Ile His 10 Ile Glu Asn Cys Asp Leu Ser Asp 18
【００２０】配列番号：５ 配列の長さ：６ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Ile Met Gly Gln Glu Lys ６SEQ ID NO: 5 Sequence Length: 6 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Ile Met Gly Gln Glu Lys 6
【００２１】配列番号：６ 配列の長さ：５ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Met Phe Tyr Ile Lys ５SEQ ID NO: 6 Sequence Length: 5 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Met Phe Tyr Ile Lys 5
【００２２】配列番号：７ 配列の長さ：９ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Leu Gly Thr Leu Leu Pro Leu Gln Lys ９SEQ ID NO: 7 Sequence Length: 9 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Leu Gly Thr Leu Leu Pro Leu Gln Lys 9
【００２３】配列番号：８ 配列の長さ：１１ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Asn Cys Asp Leu Ser Asp Pro Thr Val Tyr １０ Leu １１SEQ ID NO: 8 Sequence length: 11 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: Intermediate fragment Origin Biological name: Bovine tissue type: Fetal cartilage sequence Asn Cys Asp Leu Ser Asp Pro Thr Val Tyr 10 Leu 11
【００２４】配列番号：９ 配列の長さ：１４ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Phe Tyr Ile Lys Pro Ile Lys Tyr Lys Gly １０ Ser Ile Lys Lys １４SEQ ID NO: 9 Sequence Length: 14 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Phe Tyr Ile Lys Pro Ile Lys Tyr Lys Gly 10 Ser Ile Lys Lys 14
【００２５】配列番号：１０ 配列の長さ：１４ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Lys Leu Gly Thr Leu Leu Pro Leu Gln Lys １０ Val Tyr Pro Gly １４SEQ ID NO: 10 Sequence length: 14 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: Intermediate fragment Origin Biological name: Bovine tissue type: Fetal cartilage sequence Lys Leu Gly Thr Leu Leu Pro Leu Gln Lys 10 Val Tyr Pro Gly 14
【００２６】配列番号：１１ 配列の長さ：１８ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Lys Pro Tyr Lys Asn Lys Asn Ala Ile Asn １０ Asn Gly Val Arg Ile Ser Gly Gly １８SEQ ID NO: 11 Sequence length: 18 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: Intermediate fragment Origin Biological name: Bovine tissue type: Fetal cartilage sequence Lys Pro Tyr Lys Asn Lys Asn Ala Ile Asn 10 Asn Gly Val Arg Ile Ser Gly Gly 18
【００２７】配列番号：１２ 配列の長さ：１９ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Ala Pro Phe Thr Gly Lys Ile Met Gly Gln １０ Glu Lys Pro Tyr Lys Asn Lys Asn Ala １９SEQ ID NO: 12 Sequence length: 19 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: Intermediate fragment Origin Biological name: Bovine tissue type: Fetal cartilage sequence Ala Pro Phe Thr Gly Lys Ile Met Gly Gln 10 Glu Lys Pro Tyr Lys Asn Lys Asn Ala 19
【００２８】配列番号：１３ 配列の長さ：１１ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Ile Ser Gly Gly Gly Phe Cys Ile Lys Met １０ Phe １１SEQ ID NO: 13 Sequence length: 11 Sequence type: Amino acid Topology: Linear Sequence type: Protein Fragment type: Intermediate fragment Origin Biological name: Bovine tissue type: Fetal cartilage sequence Ile Ser Gly Gly Gly Phe Cys Ile Lys Met 10 Phe 11
【００２９】配列番号：１４ 配列の長さ：２７ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 フラグメント型：中間部フラグメント 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Lys Gly Ser Ile Lys Lys Gly Glu Lys Leu １０ Gly Thr Leu Leu Pro Leu Gln Lys Val Tyr ２０ Pro Gly Ile Gln Ser His Ile ２７SEQ ID NO: 14 Sequence Length: 27 Sequence Type: Amino Acid Topology: Linear Sequence Type: Protein Fragment Type: Intermediate Fragment Origin Biological Name: Bovine Tissue Type: Fetal Cartilage Sequence Lys Gly Ser Ile Lys Lys Gly Glu Lys Leu 10 Gly Thr Leu Leu Pro Leu Gln Lys Val Tyr 20 Pro Gly Ile Gln Ser His Ile 27
【００３０】配列番号：１５ 配列の長さ：１３３ 配列の型：アミノ酸 トポロジー：直鎖状 配列の種類：タンパク質 起源 生物名：ウシ 組織の種類：胎児軟骨 配列 Gly Pro Trp Ala Ile Ile Cys Ala Gly Lys １０ Ser Ser Asn Glu Ile Arg Thr Cys Asp Gly ２０ His Gly Cys Gly Gln Tyr Thr Ala Gln Arg ３０ Asn Gln Lys Leu His Gln Gly Val Asp Val ４０ Leu Cys Ser Asp Gly Ser Thr Val Tyr Ala ５０ Pro Phe Thr Gly Lys Ile Met Gly Gln Glu ６０ Lys Pro Tyr Lys Asn Lys Asn Ala Ile Asn ７０ Asn Gly Val Arg Ile Ser Gly Gly Gly Phe ８０ Cys Ile Lys Met Phe Tyr Ile Lys Pro Ile ９０ Lys Tyr Lys Gly Ser Ile Lys Lys Gly Glu １００ Lys Leu Gly Thr Leu Leu Pro Leu Gln Lys １１０ Val Tyr Pro Gly Ile Gln Ser His Ile His １２０ Ile Glu Asn Cys Asp Leu Ser Asp Pro Thr １３０ Val Tyr Leu １３３SEQ ID NO: 15 Sequence length: 133 Sequence type: Amino acid Topology: Linear Sequence type: Protein Origin of organism: Bovine tissue type: Fetal cartilage sequence Gly Pro Trp Ala Ile Ile Cys Ala Gly Lys 10 Ser Ser Asn Glu Ile Arg Thr Cys Asp Gly 20 His Gly Cys Gly Gln Tyr Thr Ala Gln Arg 30 Asn Gln Lys Leu His Gln Gly Val Asp Val 40 Leu Cys Ser Asp Gly Ser Thr Val Tyr Ala 50 Pro Phe Thr Gly Lys Ile Met Gly Gln Glu 60 Lys Pro Tyr Lys Asn Lys Asn Ala Ile Asn 70 Asn Gly Val Arg Ile Ser Gly Gly Gly Phe 80 Cys Ile Lys Met Phe Tyr Ile Lys Pro Ile 90 Lys Tyr Lys Gly Ser Ile Lys Lys Gly Glu 100 Lys Leu Gly Thr Leu Leu Pro Leu Gln Lys 110 Val Tyr Pro Gly Ile Gln Ser His Ile His 120 Ile Glu Asn Cys Asp Leu Ser Asp Pro Thr 130 Val Tyr Leu 133
【図１】実施例１のコンドロモジュリン−ＩＩの骨吸収
活性の検定を行った結果を表す図面である。FIG. 1 is a drawing showing the result of assaying bone resorption activity of chondromodulin-II in Example 1.
Claims (5)
殖因子を有効成分とする破骨細胞形成促進及び骨吸収促
進剤。 ＳＤＳ−ポリアクリルアミドゲル電気泳動による分子
量が約１６，０００ドルトンである。 軟骨細胞を単独でまたは繊維芽細胞増殖因子共存下で
増殖させる活性を有する。 軟骨細胞の分化機能を促進させる活性を有する。1. An agent for promoting osteoclast formation and bone resorption, which comprises a chondrocyte growth factor having the following physicochemical properties as an active ingredient. The molecular weight by SDS-polyacrylamide gel electrophoresis is about 16,000 daltons. It has the activity of growing chondrocytes alone or in the coexistence of fibroblast growth factor. It has an activity of promoting the differentiation function of chondrocytes.
表の配列番号：１で表わされるＮ末端アミノ酸配列及び
配列表の配列番号：２、３及び４で表わされる部分アミ
ノ酸配列を有することを特徴とする請求項１記載の破骨
細胞形成促進及び骨吸収促進剤。2. The chondrocyte growth factor according to claim 1 has an N-terminal amino acid sequence represented by SEQ ID NO: 1 in the sequence listing and a partial amino acid sequence represented by SEQ ID NOS: 2, 3 and 4 in the sequence listing. The osteoclast formation-promoting and bone-resorption promoting agent according to claim 1.
表の配列番号：１で表わされるＮ末端アミノ酸配列及び
配列表の配列番号：２から１４で表わされる部分アミノ
酸配列を有することを特徴とする請求項１記載の破骨細
胞形成促進及び骨吸収促進剤。3. The chondrocyte growth factor according to claim 1 has an N-terminal amino acid sequence represented by SEQ ID NO: 1 in Sequence Listing and a partial amino acid sequence represented by SEQ ID NOs: 2 to 14 in Sequence Listing. The osteoclast formation promoting and bone resorption promoting agent according to claim 1.
表の配列番号：１５に記載のアミノ酸配列で表されるこ
とを特徴とする請求項１記載の破骨細胞形成促進及び骨
吸収促進剤。4. The chondrocyte growth factor according to claim 1 is represented by the amino acid sequence set forth in SEQ ID NO: 15 of the sequence listing, and the promotion of osteoclast formation and the promotion of bone resorption are described. Agent.
または動物組織由来である請求項１記載の破骨細胞形成
促進及び骨吸収促進剤。5. The osteoclast formation-promoting and bone resorption-promoting agent according to claim 1, wherein the chondrocyte growth factor according to claim 1 is derived from human or animal tissue.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP6166917A JPH0827020A (en) | 1994-07-19 | 1994-07-19 | Osteoclast formation and bone resorption promoter |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP6166917A JPH0827020A (en) | 1994-07-19 | 1994-07-19 | Osteoclast formation and bone resorption promoter |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH0827020A true JPH0827020A (en) | 1996-01-30 |
Family
ID=15840048
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP6166917A Pending JPH0827020A (en) | 1994-07-19 | 1994-07-19 | Osteoclast formation and bone resorption promoter |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0827020A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000037093A1 (en) * | 1998-12-22 | 2000-06-29 | Japan As Represented By Secretary Of National Institute Of Infectious Diseases | Bone resorption inhibitors |
| EP1296706A1 (en) * | 2000-06-19 | 2003-04-02 | The Uab Research Foundation | Osteoclast secreted chemokine and uses thereof |
| US11958807B2 (en) | 2020-05-19 | 2024-04-16 | Cybin Irl Limited | Deuterated tryptamine derivatives and methods of use |
-
1994
- 1994-07-19 JP JP6166917A patent/JPH0827020A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000037093A1 (en) * | 1998-12-22 | 2000-06-29 | Japan As Represented By Secretary Of National Institute Of Infectious Diseases | Bone resorption inhibitors |
| US6723696B1 (en) | 1998-12-22 | 2004-04-20 | Japan As Represented By Secretary Of National Institute Of Infectious Diseases | Bone resorption inhibitors |
| EP1296706A1 (en) * | 2000-06-19 | 2003-04-02 | The Uab Research Foundation | Osteoclast secreted chemokine and uses thereof |
| EP1296706A4 (en) * | 2000-06-19 | 2004-06-16 | Uab Research Foundation | Osteoclast secreted chemokine and uses thereof |
| US11958807B2 (en) | 2020-05-19 | 2024-04-16 | Cybin Irl Limited | Deuterated tryptamine derivatives and methods of use |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| EP0527787B1 (en) | Method of predisposing mammals to accelerated tissue repair | |
| US5106626A (en) | Osteogenic factors | |
| CA2176205C (en) | Osteogenic product and process | |
| CA2265508C (en) | Bone morphogenetic protein-16 (bmp-16) compositions | |
| Okubo et al. | Osteoinduction by recombinant human bone morphogenetic protein‐2 at intramuscular, intermuscular, subcutaneous and intrafatty sites | |
| JP3706133B2 (en) | BMP-9 composition | |
| CA2238277C (en) | Compositions and therapeutic methods using morphogenic proteins and stimulatory factors | |
| JPH07504680A (en) | Prosthetic device with enhanced osteogenic properties | |
| KR20050010778A (en) | Connective tissue stimulating peptides | |
| WO1991005802A1 (en) | Osteogenic devices | |
| JP2009502782A (en) | Protein preparation for promoting hard tissue formation | |
| US9694047B2 (en) | Enhancement of bone morphogenic protein (BMP) retention with BMP binding peptide (BBP) | |
| JP5466017B2 (en) | Surgical application for BMP binding proteins | |
| EP0437281B1 (en) | Composition comprising acidic fibroblast growth factor (aFGF) | |
| NZ248446A (en) | Bone growth composition containing transforming growth factor beta (tgf-beta) and demineralised bone | |
| JPH0827020A (en) | Osteoclast formation and bone resorption promoter | |
| CN1209165C (en) | Method for cell adhesion and wound healing | |
| JPH10139682A (en) | Promoter of bone resorption | |
| JPH05339165A (en) | Fibroblast growth factor-containing medicine | |
| AU648997C (en) | Osteogenic devices | |
| Lullo | Source and Carrier Effect on the Bioactivity of BMP Bio-implants |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20050524 |
|
| A02 | Decision of refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A02 Effective date: 20051122 |