JPWO2006093313A1 - T細胞アポトーシス誘導剤 - Google Patents
T細胞アポトーシス誘導剤 Download PDFInfo
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- JPWO2006093313A1 JPWO2006093313A1 JP2007506047A JP2007506047A JPWO2006093313A1 JP WO2006093313 A1 JPWO2006093313 A1 JP WO2006093313A1 JP 2007506047 A JP2007506047 A JP 2007506047A JP 2007506047 A JP2007506047 A JP 2007506047A JP WO2006093313 A1 JPWO2006093313 A1 JP WO2006093313A1
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- lactic acid
- strain
- ferm
- bifidobacterium
- cells
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Abstract
Description
(形態学的性質)
1)細胞の形:桿菌、2)運動性の有無:無、3)胞子の有無:無、4)グラム染色:陽性
(生理学的性質)
1)カタラーゼ:陰性、2)インドールの生成:陰性、3)硝酸塩の還元:陰性、4)酸素に対する態度:通性嫌気性、5)15℃で生育:無、6)グルコースからホモ乳酸発酵によりDL乳酸生成、ガス産生無
7)各種糖類から酸生成の有無
グルコース + メリビオース −
ラクトース + ラフィノース +
マンノース + マンニトール −
フラクトース + ソルビトール −
ガラクトース + エスクリン +
シュークロース + サリシン +
アラビノース − N−アセチルグルコサミン +
マルトース + アミグダリン +
キシロース − ゲンチオビオース +
ラムノース − メレチトース −
セロビオース + デキストリン −
トレハロース + スターチ −
ラクトバチルス属の乳酸菌はMRS培地、ビフィドバクテリウム属の乳酸菌はGAM培地を用いて、37℃で18時間培養した。培養後、遠心により回収した乳酸菌菌体を洗浄して凍結乾燥を行い、乾燥菌体をPBS溶液中に懸濁し、100℃、10分間加熱処理したものを実験に供した。
細胞の調製:卵白アルブミン(以下OVA)の323残基から339残基領域をI-Ad拘束的に認識するT細胞クローンDO11.10由来のαβ-T細胞レセプター(TCR)遺伝子が導入されているDO11.10 TCR-トランスジェニックマウスから脾臓を摘出し、単細胞浮遊液を調製した。これをMACS緩衝液(0.5%ウシ血清アルブミン、2mM EDTAを含むPBS溶液)に懸濁したCD4マイクロビーズ(Militenyi Biotec)を4℃、15分作用させ、細胞を洗浄後、磁気分離カラム(Militenyi Biotec)を用い、ポジティブセレクションを回収して、CD4陽性T細胞を調製した。BALB/cマウスから脾臓を摘出して、細胞浮遊液を調製し、MACS緩衝液に懸濁したThy1.2マイクロビーズ(Militenyi Biotec)を4℃、15分作用させ、細胞を洗浄後、磁気分離カラムを用い、ネガティブセレクションを回収し、これを抗原提示細胞とした。
細胞の調製、アポトーシスの検出は実施例2と同じ方法で行った。細胞の培養:実施例2に準じて行い、添加する乳酸菌加熱死菌体として、ラクトバチルス・アシドフィルス L−92株を0.1、1.0、10μg/mlで用いた。
Th2細胞の調製:DO11.10マウス脾臓由来CD4陽性T細胞を5×105/ml、50μg/mlのマイトマイシンC(Sigma)で37℃、30分間処理したBALB/cマウス脾臓由来抗原提示細胞を1.5×106/mlに調製し、5μg/mlの抗マウスIL-12抗体(クローンC17.8)、2ng/mlのリコンビナントマウスIL-4を培地中に添加し、1mg/mlのOVA存在下、7日間培養後、回収されてきた細胞をTh2細胞として使用した。細胞培養方法及びアポトーシス検出法は実施例2と同じである。
0.1M Na2HPO4溶液で1μg/mlになるように希釈した抗IL-4抗体(Clone:11B11、BD Pharmingen)溶液をイムノプレート(Nunc)に50μl添加し、4℃で一晩静置して抗体をプレートにコーティングした。0.05% Tween20を含むPBS溶液(PBS-Tween)でウェルを洗浄後、1% BSA/PBS-Tween溶液を100μl添加し、室温で2時間静置してブロッキングを行った。PBS-Tweenでウェルを洗浄後、1% BSA/PBS-Tween溶液で希釈した標準検体の希釈列または培養上清をウェルに50μl添加して、室温で2時間静置した。PBS-Tweenでウェルを洗浄後、1% BSA/PBS-Tween溶液で0.25μg/mlになるように希釈したビオチン化抗IL-4抗体(Clone: BVD4-1D11、BD Pharmingen)溶液を50μlウェルに添加し、室温で2時間静置した。PBS-Tweenでウェルを洗浄後、1% BSA/PBS-Tween溶液で1.5μg/mlになるように希釈したアルカリフォスファターゼ−ストレプトアビジン(Zymed)溶液を50μlウェルに添加し、室温で1時間静置した。PBS-Tweenでウェルを洗浄後、4-ニトロフェニルりん酸二ナトリウム(東京化成工業)をジエタノールアミン−塩酸緩衝液(pH8.9)に1mg/mlになるように溶解させ、ウェルに50μlウェルに添加し、405nmの吸光度を測定した。
Th1細胞の誘導:DO11.10マウス脾臓由来CD4陽性T細胞を5×105/ml、50μg/mlのマイトマイシンC(Sigma)で37℃、30分間処理したBALB/cマウス脾臓由来抗原提示細胞を1.5×106/mlに調製し、5μg/mlの抗マウスIL-4抗体、2ng/mlのリコンビナントマウスIL-12を培地中に添加し、1mg/mlのOVA存在下、7日間培養後、回収されてきた細胞をTh1細胞として使用した。細胞培養方法及びアポトーシス検出法は、実施例2と同じである。
OVA特異的T細胞レセプタートランスジェニックマウス(DO11.10マウス)に対して2%のOVA(和光純薬 Cat. No.012−09885)溶液を飲水として与え、これに通常のCE-2食を与えた群をコントロールとし、0.05%のL-92加熱死菌体を含むCE-2食を与えた群をL-92群とした。また、通常の飲水とCE-2食を与えた群を無処理群(NT群)とした。飼育7日後、脾臓と腸間膜リンパ節を摘出し、OVA抗原特異的T細胞の割合を、FITCラベルされた抗CD4抗体と、PEラベルされたKJ1.26抗体(OVA-TCR特異的抗体)を用いた二重染色により、フローサイトメーターによって測定した。
Claims (9)
- ラクトバチルス(Lactobacillus)属又はビフィドバクテリウム(Bifidobacterium)属に属する乳酸菌を有効成分とするT細胞アポトーシス誘導剤。
- ラクトバチルス属に属する乳酸菌がラクトバチルス・アシドフィルス(Lactobacillus acidophilus)、ラクトバチルス・アミロボラス(Lactobacillus amylovorus)からなる群より選ばれた乳酸菌である、請求項1記載のT細胞アポトーシス誘導剤。
- ビフィドバクテリウム属に属する乳酸菌がビフィドバクテリウム・カテニュラータム(Bifidobacterium catenulatum)、ビフィドバクテリウム・ロンガム(Bifidobacterium longum)からなる群より選ばれた乳酸菌である、請求項1記載のT細胞アポトーシス誘導剤。
- 乳酸菌が、ラクトバチルス・アシドフィルス L-92株(FERM BP-4981)、ラクトバチルス・アミロボラス CP1750株(FERM BP-10532)、ビフィドバクテリウム・カテニュラータム CP2829株(FERM BP-10533)、ビフィドバクテリウム・ロンガム CP760株(FERM BP-10531)からなる群より選ばれた乳酸菌である、請求項1記載のT細胞アポトーシス誘導剤。
- ラクトバチルス(Lactobacillus)属又はビフィドバクテリウム(Bifidobacterium)属に属する乳酸菌を含有する、T細胞のアポトーシスを誘導するための健康食品。
- 乳酸菌が、ラクトバチルス・アシドフィルス L-92株(FERM BP-4981)、ラクトバチルス・アミロボラス CP1750株(FERM BP-10532)、ビフィドバクテリウム・カテニュラータム CP2829株(FERM BP-10533)、ビフィドバクテリウム・ロンガム CP760株(FERM BP-10531)からなる群より選ばれた乳酸菌である、請求項5記載のT細胞のアポトーシスを誘導するための健康食品。
- 新規乳酸菌ラクトバチルス・アミロボラス CP1750株(FERM BP-10532)。
- 新規乳酸菌ビフィドバクテリウム・カテニュラータム CP2829株(FERM BP-10533)。
- 新規乳酸菌ビフィドバクテリウム・ロンガム CP760株(FERM BP-10531)。
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EA016734B1 (ru) * | 2006-12-21 | 2012-07-30 | Калпис Ко., Лтд. | СРЕДСТВА ДЛЯ СТИМУЛЯЦИИ ПРОДУЦИРОВАНИЯ IgA НА ОСНОВЕ БАКТЕРИЙ LACTOBACILLUS AMYLOVORUS И ШТАММ БАКТЕРИЙ LACTOBACILLUS AMYLOVORUS CP1750 FERM BP-10532 |
TWI356680B (en) * | 2007-01-05 | 2012-01-21 | Promd Biotech Co Ltd | Anti-allergy lactic acid bacteria |
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US9439449B2 (en) * | 2009-08-18 | 2016-09-13 | Nestec S.A. | Nutritional composition comprising Bifidobacterium longum strains and reducing food allergy symptoms, especially in infants and children |
JP5737646B2 (ja) * | 2010-03-24 | 2015-06-17 | 森下仁丹株式会社 | 抗アレルギー剤 |
FR2962134B1 (fr) * | 2010-07-05 | 2014-12-19 | Bifinove | Procede de preparation d'un complexe macromoleculaire non separe (totum), et utilisations dudit complexe dans la prevention de troubles arthritiques |
JP5840368B2 (ja) | 2011-02-02 | 2016-01-06 | カルピス株式会社 | 関節炎予防改善用物質 |
CN104780932A (zh) | 2012-02-29 | 2015-07-15 | 伊西康内外科公司 | 微生物区系的组合物及与其相关的方法 |
EP3212001A4 (en) | 2014-10-31 | 2018-04-25 | Whole Biome Inc. | Methods and compositions relating to microbial treatment and diagnosis of disorders |
CN111372596A (zh) | 2017-08-30 | 2020-07-03 | 潘德勒姆治疗公司 | 用于治疗微生物组相关病症的方法和组合物 |
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JPH07265064A (ja) * | 1993-11-23 | 1995-10-17 | Taketoshi Yamada | 腸内細菌叢改善組成物 |
JP2992945B2 (ja) * | 1994-03-11 | 1999-12-20 | カルピス株式会社 | ラクトバチルス・アシドフィルス乳酸菌 |
JP3400282B2 (ja) * | 1997-02-21 | 2003-04-28 | 株式会社ヤクルト本社 | 脂質代謝改善剤およびそれを含有する食品 |
JPH10309178A (ja) * | 1997-05-09 | 1998-11-24 | Wakamoto Pharmaceut Co Ltd | ビフィズス菌を有効成分とする抗アレルギー剤および醗酵食品 |
IT1298918B1 (it) * | 1998-02-20 | 2000-02-07 | Mendes Srl | Uso di batteri dotati di arginina deiminasi per indurre apoptosi e/o ridurre una reazione infiammatoria e composizioni farmaceutiche |
US7125698B2 (en) * | 1999-08-09 | 2006-10-24 | Matthew Glenn | Polynucleotides, materials incorporating them, and methods for using them |
JP2002306125A (ja) * | 2001-04-12 | 2002-10-22 | Morinaga Milk Ind Co Ltd | プロバイオティクスを含有する包装容器入り乳幼児用栄養組成物 |
US20030092163A1 (en) * | 2001-07-26 | 2003-05-15 | Collins John Kevin | Probiotic bifidobacterium strains |
CA2364249A1 (fr) | 2001-11-27 | 2003-05-27 | Unknown-Inconnu | Effet de bacteries lactiques sur l'apoptose cellulaire de tumeur |
JP4212838B2 (ja) * | 2002-06-26 | 2009-01-21 | カルピス株式会社 | 抗アレルギー剤 |
TWI241912B (en) * | 2002-10-30 | 2005-10-21 | Food Industry Res & Dev Inst | Novel Acid-and bile salt-resistant Lactobacillus isolates having the ability to lower and assimilate cholesterol |
US20040208863A1 (en) | 2003-01-30 | 2004-10-21 | James Versalovic | Anti-inflammatory activity from lactic acid bacteria |
WO2004076615A2 (en) | 2003-02-27 | 2004-09-10 | Bioneer A/S | Immunomodulating probiotic compounds |
JP4479162B2 (ja) * | 2003-03-18 | 2010-06-09 | 高梨乳業株式会社 | 免疫機能調節剤および免疫機能調節食品 |
JP2005060285A (ja) | 2003-08-11 | 2005-03-10 | Masako Ito | 口腔清拭法 |
JP3862687B2 (ja) | 2003-09-09 | 2006-12-27 | 沖電気工業株式会社 | レベルシフタ回路 |
TW200637908A (en) | 2005-01-04 | 2006-11-01 | Calpis Co Ltd | Method for preparation of lactic acid bacterium having anti-allergic activity |
TW200700074A (en) | 2005-03-04 | 2007-01-01 | Calpis Co Ltd | Inducer of t cell apoptosis |
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US20110250189A1 (en) | 2011-10-13 |
US9339055B2 (en) | 2016-05-17 |
EP1854468A4 (en) | 2012-02-08 |
WO2006093313A1 (ja) | 2006-09-08 |
US20080166787A1 (en) | 2008-07-10 |
TW200700074A (en) | 2007-01-01 |
EP1854468A1 (en) | 2007-11-14 |
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EP1854468B1 (en) | 2013-04-24 |
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