201219573 六、發明說明： 【發明所屬之技術領域】 本發明有關於生物相雜域物，其係使躲獅或盘 研究對象中的植體材料或輸送系統有關之纖維化組織反應的方 法。發明亦有關於用以治療或預防與研究對象中之植體材料 輸送系統有關的纖維化組織缺陷之方法。 一 【先前技術】 諸如髖關節、膝關節及肩關節義體(以及所有活動關節）、乳 房、肌肉及牙齒植體、心瓣膜、血管移植物及皮膚移植物的植體 應用已自不同類型之包含鈦、聚合物之生物材料及如膠原蛋白、 玻尿酸、幾丁聚醣、豬小腸、肌腱等之生物成分及組織成功地發 展。然而，即使至今所使用的材料具有惰性本質，但仍由於與周 圍細胞及組織缺乏建設性交互作用（生物相容性)而常有植體^力能 不良。此外，用以促進細胞再生的人工裝置植入或細胞移植已遭 到受限的臨床成功。 因此，為了改善人體健康及病患生命品質，而對處理位於植 體組織界面之植體裝置及細胞輸送系統的生物接受度問題有条迫 需求。 ^ 組織受傷後，可能發生二明顯現象：1.受傷細胞由相同類型的 健康細胞所取代的正常再生過程；或2.結締組織以細胞外基質之 未X控沉積取代正常組織的慢性纖維化反應。可將已開發世界中 的所有死亡中約45%歸因為一些類型的慢性纖維增生疾病。關於 植體材料的纖維化反應經常變成植體鬆弛及器官功能減低(即髖 部、肩部、及頜骨）。 WO 98/54301 (MICKLE DONALD A G et al.)揭露用以在心臟 組織中形成移植物的方法，包含移植選自心肌細胞、纖維母細胞、 平滑肌細胞、内皮細胞及骨骼肌母細胞的細胞。移植物在治療心 臟上的傷疤組織方面尤其有用。亦提供用於此移植物的分離及培 201219573 養心肌細胞的方法。 WO 01/32129 (GERIGENE MEDICAL CORP)揭露藉由注射或 直接手術放置/植入源自結締組織、真皮、肌膜、固有層^啦^^ propria)、基質(stroma)、脂肪組織、肌肉、肌腱、韌帶或毛囊的自 體培養細胞及/或培養細胞產生的細胞外基質，而長期增^及/ 或修復皮膚缺陷(傷疤、皮膚鬆弛、皮膚變薄、及皮膚增生）、皮下 脂，團、乳房組織、創傷及灼傷、泌屎道及腸胃道的括約肌結構、 疝氣)牙周疾病及失調、肌腱及韌帶撕裂及禿頭的方法。改善性 應用係進行於鄰近或在缺陷區域内的組織上。該方法涉及自嬰兒 或人類胎兒的主體取出能發育的細胞。亦或是，改善性應用涉及 在置入關於研究對象缺陷的位置之前，放置於較佳地由作為三維 結構或作為懸浮物的自體細胞外基質成份所組成之基質中的細 胞。在進一步實施例中，自培養物收集較佳的自體細胞外基質成 份並放置在關於研究對象缺陷的位置。 US^2010/129414 (Medtronic Vascular，Inc.)描述用以治療動脈 瘤、血管阻塞、及血管受損的方法。該方法包含使用可植入的醫 療裝置，其包含與其表面結合的生物活性劑基質。微脂體儀 封裝生物活性劑並全身性或局部性地輸送至血流。一旦選定適當 位置且微脂體已經由血管分佈將自身分散，便使用自微脂體釋放 生物活性劑的方法。一經釋放，則可藉由與可植入的醫療裝置結 合的生物活性劑基質隔離生物活性劑，並緩慢地釋放以給予g 組織治療效果。 未分化的胎兒細胞已揭露於WO 03/068287 (Neocutis SA)中。 亦揭露設計成用以治療罹患皮膚症狀、失調或疾病之研究對象的 方法及組成物。組成物包含與膠原蛋白基質或與載體結合之未分 化的胎兒皮膚細胞。 對於續保可回復失去的組織功能及克服導致全面整體性降低 的纖維化不利反應的安全具作用之植體材料具有急迫需求。迄 今’尚未有藥物被核准為抗纖維化療法(s. Schultze_M〇sgau et al. "Principles and mechanisms of peri-implant soft tissue healing» 4 201219573201219573 VI. Description of the Invention: [Technical Field] The present invention relates to a biological phase hybrid which is a method for reacting a plant material or a delivery system-related fibrotic tissue in a lion or a disk research object. The invention also relates to methods for treating or preventing fibrotic tissue defects associated with an implant material delivery system in a subject. [Prior Art] Implant applications such as hip, knee and shoulder joints (and all active joints), breast, muscle and dental implants, heart valves, vascular grafts and skin grafts have been used in different types. Biological materials including titanium, polymers, and biological components such as collagen, hyaluronic acid, chitosan, pig small intestine, tendon, and the like have been successfully developed. However, even though the materials used to date have an inert nature, they often have poor implantability due to lack of constructive interaction (biocompatibility) with surrounding cells and tissues. In addition, artificial device implantation or cell transplantation to promote cell regeneration has been limited in clinical success. Therefore, in order to improve human health and the quality of life of patients, there is a strong need to deal with the biological acceptability of implant devices and cell delivery systems located at the interface of plant tissues. ^ After tissue injury, two obvious phenomena may occur: 1. normal regeneration process in which injured cells are replaced by healthy cells of the same type; or 2. connective tissue replaces chronic fibrosis with normal tissue by uncontrolled deposition of extracellular matrix . About 45% of all deaths in the developed world can be attributed to some types of chronic fibroproliferative diseases. The fibrotic response to implant materials often becomes loss of implants and reduced organ function (ie, hips, shoulders, and jaw bones). WO 98/54301 (MICKLE DONALD A G et al.) discloses a method for forming a graft in cardiac tissue comprising transplanting cells selected from the group consisting of cardiomyocytes, fibroblasts, smooth muscle cells, endothelial cells, and skeletal muscle mother cells. Grafts are especially useful in treating scar tissue on the heart. Methods for isolating and culturing 201219573 cardiomyocytes are also provided. WO 01/32129 (GERIGENE MEDICAL CORP) discloses that by injection or direct surgical placement/implantation originating from connective tissue, dermis, sarcolemma, lamina propria, propria, stroma, adipose tissue, muscle, tendon Autologous culture cells of ligaments or hair follicles and/or extracellular matrices produced by cultured cells, and long-term increase and / or repair of skin defects (scarred, skin loosening, skin thinning, and skin hyperplasia), subcutaneous fat, mass, Breast tissue, trauma and burns, sphincter structure of the gastrointestinal and gastrointestinal tract, hernia) periodontal disease and disorders, tendon and ligament tears and baldness. The improvement application is performed on tissue adjacent to or within the defect area. The method involves withdrawing a developing cell from the body of an infant or human fetus. Alternatively, the improved application involves placing the cells in a matrix preferably composed of autologous extracellular matrix components as a three-dimensional structure or as a suspension prior to placement of the defect in the subject. In a further embodiment, a preferred autologous extracellular matrix component is collected from the culture and placed at a location relative to the defect of the subject. US^2010/129414 (Medtronic Vascular, Inc.) describes a method for treating aneurysms, vascular occlusion, and vascular damage. The method comprises the use of an implantable medical device comprising a bioactive agent matrix bound to its surface. The liposome encapsulates the bioactive agent and delivers it to the bloodstream systemically or locally. Once the appropriate location is selected and the liposomes have been dispersed by the vascular distribution, a method of releasing the bioactive agent from the liposomes is used. Once released, the bioactive agent can be isolated by a bioactive agent matrix associated with the implantable medical device and slowly released to impart a therapeutic effect to the g tissue. Undifferentiated fetal cells have been disclosed in WO 03/068287 (Neocutis SA). Methods and compositions designed to treat subjects suffering from skin symptoms, disorders or diseases are also disclosed. The composition comprises undiluted fetal skin cells in combination with a collagen matrix or with a carrier. There is an urgent need for renewal of implant materials that can restore lost tissue function and overcome the safety effects of fibrotic adverse reactions that result in overall overall reduction. To date, no drugs have been approved for anti-fibrotic therapy (s. Schultze_M〇sgau et al. "Principles and mechanisms of peri-implant soft tissue healing» 4 201219573
Quintessence International, Vol. 36, Number 10, November/December i〇〇5-pp759_769)，且因此協助結合再生組織及克服植體媒介(輸送 系統媒介)之纖維化反應的能力之方法將具有重大益處。 •如將自前述内容顯而易見的該等及其他目標已由本發明達 成0 【發明内容】 .在第一貫施例中，本發明有關於生物相容性組成物，使用於 預防或治療與研究對象中之植體材料或輸送系統有關的纖維組織 反應的方法。本發明之生物相容性組成物包含自懷孕9至16週、 較佳地自10至16週、且更佳地自懷孕12_14週之已分化所得到的 胎兒細胞產物。 本發明亦有關於用以治療與研究對象中之植體材料或輸送系 統有關的纖維化組織缺陷的方法，該方法包含將本發明之生物相 谷性組成物置入位於該纖維化組織缺陷内或鄰近該纖維組織缺陷 處的該植體材料或輸送系統。尤其，該方法有關於受培養之三維 ^分化胎兒細胞產物或其衍生物或碎片的注射、植入、及/或附 著。 〆 士，明之另一目標有關於用以預防與研究對象中之植體材料 或輸送祕有_纖維組織反應之方法，該方法包含咖本發明 之生物相谷性組成物塗覆該植體材料或輸送系統。 亦揭路治療或預防研究對象罹患纖維化組織反應失調，該方 法包含將本發明之生物相容性組成物施加至該研究對象。 見 而、隹ίϊίϋί術領域者而言，藉由檢閱參考以下例示性圖式 ，進订的__續畴糊，其他目標及優鱗變得顯而易 【實施方式】 申請人已出乎意料地魏朗胎兒 助使得生物材料適用於成人細胞及幹細胞輸送(見f下實 201219573 體或基置人體㈣’軟_，尤其是硬纽軟骨，伴 成纖維組織的反應。申請人已說明自懷孕9至16週、較佳地 至16週且更佳地自週取得的已分化胎兒細胞產物藉由 及/或>肖除紐/輸送系、關®的纖維纟職形成秘得外來物更 具可接受度。 此外’多半使用巧理上自宿主分離細胞且偈限細胞的封裝程 序，以呈現用於臨床環境的胚胎幹細胞進而使得其免疫上可為 且預防腫瘤職。實際上這些植體仍在植體周圍u般性ς 化且未排列的組織形成。在這些情況中，提出使用得自懷孕9至 較^地自1G至16週且更佳地自1244週的已分化胎兒細 胞產物，平行地亦限制及/或消除這有害的纖維化反應。 如同此處所使用，為了增進本發明之理解，提供;列定義。 「一」意為「至少一個」或「一或更多」。 用語-「包含」通常用於包括的概念，亦即是允許出現一或更 多特徵或元件。 用語「生物材料」意為當與細胞或生物組織接觸時不具有害 作用的包含金屬、陶瓷及聚合物之自然或合成材料。通常^物& 料支持物係選自由聚合支持物、生物相容金屬支持物、及/或生 物相容陶瓷、及其混合物所組成的群組，該聚合支持物包含烯烴 ♦合物、氟聚合物、聚苯乙烯、聚丙浠聚合物、聚酯聚合物、聚 氨醋聚合物、石夕聚合物、、纖維素聚合物、氧樹脂聚合^、石夕樹 脂基聚合物、合成水凝膠、聚碳酸酯；該生物相容金屬支持物包 含鈦及鈦合金、鎳鈦形狀記憶合金、鍅土、不鏽鋼及鈷鉻合金、 矽土一錯土組成物；且該生物相容陶瓷包含瓷、氫氧基鱗灰石。 此處所使用的用語「生物相容」應意為不造成動物傷害或死 亡、或當安置成與動物組織緊密接觸時不在動物中引發有害反應 的任何材料。有害反應包含但不侷限於：發炎、感染、X纖士組^ 形成、細胞死亡及血栓形成。 201219573 ϋ斤，用=「生物可降解」意指生物相容 途狀作用麵_受到分解的聚合體 =吊生化 換使用，然而它們不具有制顧^^ 聚合物為生物可降解，並因此可經ί化催itlt 切割成生物相容性職物。 ⑽水解作用 此處所使用的「非生物可降解意指生 經由正常生化麵之作較解㈣合體内不 ϋ所使用的「無實質上毒性」意指全身或局性， 理，術領域具有通常技能的醫師及藥理學家所判定4 接文者的利益超過治療的生理上有害作用。 與質上I接受^指於活體内的有效劑量無 只貝上:性的所有化合物、前驅藥物、衍生物、及鹽類。 用語「已分化胎兒細胞」意為相較=依wo 03/068287_)〇^ SA)之未分化胎兒細胞的已分化細胞。^本發 明相反I用於WO_68287的用語「未分化」描述未成熟^^ 細胞:舉，言，未分化的敎皮膚細胞包含可分化成真皮纖維 母細胞及表皮肖質細胞者。已分化細胞為當放置在特定針對另一 細胞麵的分化培養基中或至不同微環境中時，將不輕易去分化 成不同細胞譜系者。糊來說’如果將胎兒皮膚_母細胞放置 到成骨性分化培養基中，則其將不會變成整群的造骨細胞；或如 果將其放置在成脂性分化培養基中’則其將不會變成整群的脂肪 細胞，並且如果將相同細胞放置到與硬骨結合的3D基質中，則除 非經過不合理時期’其將不會由於環境改變而去分化成整群的造 骨細胞。如少於9週之胎兒細胞的未分化細胞族群及間質幹細胞 (Mesend^nal stem cell，MSC)可藉由添加14_21天的特定生長因子 而誘導成諸如成骨性或成脂性的其他細胞譜系。 /用語「得自懷孕9_16週的已分化胎兒細胞產物」意為細胞族 群係源自特定組織，亦即皮膚、硬骨、軟骨、肌肉，且具有特定 組織類型的型態及表面標記。低於9週之妊娠的細胞按照定義為. 201219573 MSC且構成未分化細胞族群。 在許多國家，懷孕16週之後，針對移植目的而使用胎兒組織 /細胞在法律上及倫理上不可允許。（例如，見“Bioengineering: Principles, Methodologies and Applications, ISBN: 978-1-60741-0; 2009 Nova Science Publisher, Inc. Chapter 4 Bioengineering of Human Fetal Tissues For Clinical Use, Lee Ann Applegate et al; see also Verklan，MT. (1993), “The ethical use of fetal tissue for transplantation and research”. J Advanced Nursing, 18, 1172-1177; or Rahman A. et al. (1998) “A global review of laws on induced abortion’’，1985-1997. Intern Family Planning Perspec, 24, 56-64) 用語「適當的培養條件」為包含促進增殖之養分的用於培養 細胞之培養基。營養培養基可以適當組合及以適當濃度而包含下 列之任何一者：等張食鹽溶液、緩衝液、胺基酸、血清或血清取 代物、及其他外來添加因子。熟悉本技藝者將察覺可使用任何一 般性使用的培養條件。 用語「細胞系」意指：給予適當的新鮮培養基及充足空間而 將無限增殖的長期不變地建立之細胞培養物。吾人並非生產已建 立的細胞系，而生產初代細胞系。 初代細胞糸之用語意指具有受限傳代數的已建立細胞培養 物。 用語「純株」意指通常發展自單一選定細胞的次群集細胞。 ^用語「細胞寄存」意指：自捐贈者胎兒組織獲得檢體；在適 當培養條件下，使胎兒組織生長並將胎兒細胞增殖至高濃度；使 組織及產生的培養物之細胞受到胰蛋白酶作用來允許其懸浮； 合懸浮細胞以自培養物製成大致均⑽細麟浮體；與^東劑^ 緩混合；將等分之細麟浮體贿在魏巾；及鮮分結 由以1°C /min降低安瓿溫度直到書c，而後約24小時後 ^ 已鮮的WFf㈣作_輯進行完整 本超低溫庫贿細胞使其停止老化，藉此允許聽有在其被 201219573 採集當天所具有的功能及活性。 用語「治療」（如在「治療纖維化組織缺陷、症狀、失調或疾 病」中）包含：（1)預防症狀，即避免任何症狀之臨床徵候；（2)抑制 症狀，即停止臨床徵候之發展或進展；及/或⑶緩和、修復或逆 轉症狀’即造成臨床徵候之復原。 少 用叩症狀」、「缺陷」、「失調」、及「疾病」於此處可互相交 換使用，以意指可藉由施用敘述於此的本發明之包含胎兒細胞產 物的生物相容性組成物而加以預防或治療的生理狀態。 …f語「研究對象」（如於「研究對象」之治療中）或「病患」 係欲思指涉受到折磨、易於或罹患症狀、缺陷、失調、及疾病(如 ^處所指明)的哺乳類健。本聽包含人類及動物兩者。舉例而 吕’研究對象可為如人類、非人類的靈長類、野生動物、狗、猶、 馬牛、豬、緯羊、兔、大鼠、或小鼠。如此處所使用，野生動 任何非•丨翻哺乳類、鳥類、兩棲類或魚類。此野 生動物之:r例包含但不舰H雜、獅、虎、熊、鷹 二維基質意為選自膠原蛋白基質或PLA、PLGA、PE(}、 ί& HM_Hyal_ie add，必)的水凝膠、 =二f:?喊其組成物的任何基質。基質提供三維空間 絲胞或胎兒產物適當覆蓋及輸送至或亦與額外植體材 妝化=「ϊ原蛋白」意指受到料酵素降解之师上親水的多 受到徹底研究，因此許多關鍵參數可加以 =方3 itif原’藉此導致最小的排斥可能。用於本發 月之τ法及用途中的較佳膠原蛋白為馬膠原蛋白。 「植體」可被視為欲替換欠缺的生物 如療植體為人造裝置，且接觸身體的植體表面可』 如鈦、魏樹脂、聚合物、鱗灰石、生物 成。—些植體可具有如可植入膠囊或塗藥血管竿的 —式生物活性溶出麵。频材料可針對抗齡化反應而= 201219573 定組織類型的已分化胎兒細胞結合。 .產生纖維化組織反應的植體材料之具體實例包含但不侷限 於.： 齒植體:最頻繁的材料為作為決定性_的鈦及並混人 合金。植體表面可藉由電浆喷塗、陽極處理、_或喷沙^加二 修改來增加植體之表面積及整合可能性。 、 -齒增大物 硬骨：包含PLA、PLGA、氫·磷灰石、碟酸三灿 phosphate，TCP)的多種聚合物 -軟組織：膠原發泡體、水凝膠、針對植體 膜及牙齦增料合物絲 瓷 料 體 建 _競關節、肩關節、膝關節、活動關節植體：材料 健’ A體可包含如喊頭絲乙職關蛛的其他材 -乳房植體：㈣包含各種轉性f的鄉性體 膠原蛋白、水凝膠。 γ安…日乂尥 量重 -肌植體：使用如矽氧樹脂及膠原蛋白的材料之軟組織大 -角膜及視網膜植體包含石夕氧樹脂、凝膠、pla -皮膚植體包含HA凝膠、膠原蛋白、矽氧樹脂。 -或熟悉本技藝者已知的其絲何可獻醫縣置。例 植入醫療裝置包含但不偈限於：企管支架、導尿管、一般 縫線、可植入的心律調節器、微粒、探針及血管移 醫療裝置可單獨或組合使用。 」植入 用語「輸送系統」意為提供轉移已分化胎兒細胞或依 明之已分化胎兒細胞產物本身或與植體結合之手段以治 ^ 使得植體的任何金屬性或常用的整形外科、創傷、顎面之自、= 合成植體材料、水凝膠、矽氧樹脂或移植物。 “'、2 此伴隨得自懷孕9至16週、較佳地自1〇至16週且 12-14週的已分化减細胞輸送之抗_化活性的新發現丄有決 201219573 定性的重要性。再者，用 16週且更佳地自12_14 代孕9至16週、較佳地自1〇至 助使得生物材料相容於成人細月產物(存活或死亡)，協 植體及裝置可取決於 輸达(見以下實例）。 殺菌程序而與組織不佳的 ^置材料、表面降解性質及終端 體/裝置周圍的快出現有關。植 孕9至16週、較佳地自1〇V=敦/决疋取佳功能結果。得自懷 胎兒細胞可協助植體/裝置 规自12·14週的已分化 炎反應。在懷孕早期的(ff 修復及適應以避免發 在12-14警八斗週較佳地自10至16週且更佳地 纖維母細胞形成刀5為在組織修復_抑制肌 /麥置妹人日士 Βπ^^/ 並猎此抑制纖維化，尤其當與植體 肌^香^丨Ϊ義母細胞在以細胞外基質過量產生及平滑肌 ii 重要角色。對抗與逐步組織破壞有關之纖維化的可能 ϋ應針對局雜眺組織修復區域及/或與植體表面結合的 肌纖維母細胞之常駐族群。間f幹細胞及胎躺f幹細胞呈現指 不肌纖維母細胞類型之細胞的alpha_ SMA陽性。這些細胞為形成 傷疲及纖維組織中的關鍵要素(Estes et al，Differentiati〇n，Quintessence International, Vol. 36, Number 10, November/December i〇〇5-pp759_769), and thus the method of assisting in the ability to combine regenerative tissue and overcome the fibrotic response of the implant medium (transport system media) would be of great benefit. • These and other objects as apparent from the foregoing have been achieved by the present invention. [Invention] In the first embodiment, the present invention relates to a biocompatible composition for use in prevention or treatment and research subjects. A method of fiber tissue reaction associated with an implant material or delivery system. The biocompatible composition of the present invention comprises fetal cell products obtained from the differentiation of 9 to 16 weeks of pregnancy, preferably from 10 to 16 weeks, and more preferably from 12 to 14 weeks of pregnancy. The invention also relates to a method for treating a fibrotic tissue defect associated with an implant material or delivery system in a subject, the method comprising placing a biological phase gluten composition of the invention within the fibrotic tissue defect or The implant material or delivery system adjacent to the fibrous tissue defect. In particular, the method pertains to the injection, implantation, and/or attachment of cultured three-dimensionally differentiated fetal cell products or derivatives or fragments thereof. Gentleman, another target of the invention relates to a method for preventing the reaction of the implant material or the secret tissue in the research object, the method comprising the bio-phase gluten composition of the invention coating the implant material Or delivery system. The subject of the treatment or prevention of the subject is also suffering from a disorder of fibrotic tissue reaction, which method comprises applying the biocompatible composition of the present invention to the subject. In the field of seeing, 隹 ϊ ϊ ϋ , , , , , , , 检 检 检 检 检 检 检 检 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考 参考Diwei's fetal help makes biomaterials suitable for adult cell and stem cell delivery (see f sturdy 201219573 or base human (4) 'soft _, especially hard neonatal cartilage, with fibroblastic tissue response. Applicant has stated that self-pregnancy The differentiated fetal cell product obtained from 9 to 16 weeks, preferably to 16 weeks, and more preferably from the week, is formed by the and/or > It is more acceptable. In addition, most of the packaging procedures used to isolate cells from the host and limit cells are presented to present embryonic stem cells for use in the clinical environment, thereby making them immune and preventing oncology. In fact, these implants Tissues that are still ubiquitous and unaligned around the implant are formed. In these cases, it is proposed to use differentiated fetal cell products from pregnancy 9 to more than 1G to 16 weeks and more preferably from 1244 weeks. Parallel and limited / or eliminate this harmful fibrotic reaction. As used herein, in order to enhance the understanding of the present invention, a column definition is provided. "One" means "at least one" or "one or more". The term - "include" is usually The concept of inclusion, that is, the admission of one or more features or elements. The term "biological material" means a natural or synthetic material comprising metals, ceramics and polymers that are not harmful when in contact with cells or biological tissues. Typically, the support & support is selected from the group consisting of polymeric supports, biocompatible metal supports, and/or biocompatible ceramics, and mixtures thereof, the polymeric support comprising an olefin conjugate, Fluoropolymer, polystyrene, polypropylene polymer, polyester polymer, polyurethane polymer, Shixia polymer, cellulose polymer, oxygen resin polymerization, Shishi resin-based polymer, synthetic water condensation Glue, polycarbonate; the biocompatible metal support comprises titanium and titanium alloy, nickel titanium shape memory alloy, alumina, stainless steel and cobalt chrome alloy, bauxite-type soil composition; and the biocompatible Porcelain contains porcelain, hydroxyapatite. The term "biocompatible" as used herein shall mean any material that does not cause injury or death to an animal, or that does not cause an adverse reaction in an animal when placed in close contact with animal tissue. Harmful reactions include, but are not limited to, inflammation, infection, X-ray formation, cell death, and thrombosis. 201219573 ϋ, with = "biodegradable" means biocompatible pathways _ subject to decomposition Polymer = hang biochemical use, however, they do not have the technical properties of the polymer is biodegradable, and therefore can be cut into biocompatible jobs by stimulating itlt. (10) Hydrolysis The "non-biological" used here Degradable means that the normal biochemical surface is used to solve the problem. (4) The "no substantial toxicity" used in the body is determined by the physician or pharmacologist with general skills in the whole body or in the field. The interests of the recipient exceed the physiologically harmful effects of the treatment. The effective dose of the substance I accepts in vivo is not all of the compounds, precursor drugs, derivatives, and salts. The term "differentiated fetal cells" means differentiated cells of undifferentiated fetal cells compared to ==03/068287_)〇^ SA). In contrast, the term "undifferentiated" used in WO_68287 describes immature cells. In other words, undifferentiated sputum skin cells contain those that can differentiate into dermal fibroblasts and epidermal tract cells. The differentiated cells will not readily differentiate into different cell lineages when placed in a differentiation medium specific to another cell surface or into a different microenvironment. For the paste, 'If the fetal skin_mother cells are placed in the osteogenic differentiation medium, they will not become a whole group of osteoblasts; or if they are placed in the adipogenic differentiation medium, then it will not It becomes a whole group of fat cells, and if the same cells are placed in a 3D matrix that binds to the hard bone, unless it passes through an irrational period, it will not dedifferentiate into a whole group of osteoblasts due to environmental changes. For example, undifferentiated cell populations and mesenchymal stem cells (MSCs) of fetal cells less than 9 weeks can be induced into other cell lineages such as osteogenic or adipogenic by adding 14-21 days of specific growth factors. . / The phrase "differentiated fetal cell product from 9-16 weeks of pregnancy" means that the cell family is derived from a specific tissue, namely skin, hard bone, cartilage, muscle, and has a type and surface marker of a particular tissue type. Cells less than 9 weeks of gestation are defined as .201219573 MSC and constitute an undifferentiated cell population. In many countries, the use of fetal tissue/cells for transplantation purposes is not legally and ethically permitted after 16 weeks of pregnancy. (For example, see "Bioengineering: Principles, Methodologies and Applications, ISBN: 978-1-60741-0; 2009 Nova Science Publisher, Inc. Chapter 4 Bioengineering of Human Fetal Tissues For Clinical Use, Lee Ann Applegate et al; see also Verklan , MT. (1993), “The ethical use of fetal tissue for transplantation and research”. J Advanced Nursing, 18, 1172-1177; or Rahman A. et al. (1998) “A global review of laws on induced abortion” ', 1985-1997. Intern Family Planning Perspec, 24, 56-64) The term "appropriate culture conditions" is a medium for culturing cells containing nutrients for promoting proliferation. The nutrient medium may be suitably combined and contained in an appropriate concentration to include any of the following: an isotonic saline solution, a buffer, an amino acid, a serum or serum substitute, and other external addition factors. Those skilled in the art will be aware of the culture conditions in which any general use can be used. The term "cell line" means a long-term, invariably established cell culture that is given an appropriate fresh medium and sufficient space for immortalization. We do not produce established cell lines, but produce primary cell lines. The term "primary cell" means an established cell culture with a restricted number of passages. The term "pure strain" means a sub-clustered cell that is usually developed from a single selected cell. ^ The term "cell storage" means: obtaining a specimen from the fetal tissue of the donor; under appropriate culture conditions, the fetal tissue is grown and the fetal cells are proliferated to a high concentration; and the cells of the tissue and the culture produced are subjected to trypsin action. Allow the suspension to be suspended; the suspension cells are made into a substantially uniform (10) fine lining floating body from the culture; and the mixture is slowly mixed with the ^dong agent; the aliquot of the lining float is bribed in the Wei towel; and the fresh branch is 1°. C / min lowers the ampoule temperature until the book c, and after about 24 hours, the fresh WFf (four) is used to complete the ultra-low temperature library to stop the aging, thereby allowing the listener to have the function on the day of its collection by 201219573. And activity. The term "treatment" (eg, in "treatment of fibrotic tissue defects, symptoms, disorders, or diseases") includes: (1) prevention of symptoms, ie, avoiding any clinical signs of symptoms; (2) inhibition of symptoms, that is, stopping the development of clinical signs Or progress; and/or (3) mitigating, repairing or reversing the symptoms' causes recovery of clinical signs. "Use of sputum symptoms", "defect", "disorder", and "disease" are used interchangeably herein to mean a biocompatible composition comprising a fetal cell product of the invention as described herein. The physiological state of prevention or treatment. ...f "study object" (such as in the treatment of "subjects") or "patient" is a mammal that is afflicted with affliction, susceptibility or suffering from symptoms, defects, disorders, and diseases (as indicated by ^where) Jian. This listens to both humans and animals. For example, Lu's subjects may be humans, non-human primates, wild animals, dogs, baboons, horses, pigs, lats, rabbits, rats, or mice. As used herein, wildly move any mammal, bird, amphibious or fish. This wild animal: r case contains but not H, lion, tiger, bear, eagle two-dimensional matrix means selected from collagen matrix or PLA, PLGA, PE (}, ί & HM_Hyal_ie add, must) Glue, = two f:? Shout any substrate of its composition. The matrix provides three-dimensional space for the cell or fetal product to be properly covered and transported to or also with additional implants. "Plutonium protein" means that the hydrophilicity of the material that is degraded by the enzyme is thoroughly studied, so many key parameters can be Add = square 3 itif original 'by taking this to cause minimal rejection possible. A preferred collagen for use in the τ method and use of this month is horse collagen. "Insulin" can be considered as a replacement for a missing organism. For example, the implant is an artificial device, and the surface of the implant that contacts the body can be made of titanium, Wei resin, polymer, ash, or bio. - Some implants may have a biologically active dissolution surface such as an implantable capsule or a coated vasospasm. The frequency material can be targeted to the ageing response = 201219573 to determine the tissue type of differentiated fetal cells. Specific examples of implant materials that produce a fibrotic tissue reaction include, but are not limited to,: Dental implants: The most frequent materials are titanium and miscible alloys that are decisive. The surface of the implant can be modified by plasma spraying, anodizing, _ or sandblasting to increase the surface area and integration possibilities of the implant. - Tooth enlargement hard bone: a variety of polymers including PLA, PLGA, hydrogen apatite, tributyl phosphate, TCP) - soft tissue: collagen foam, hydrogel, implant film and gum feeding线丝瓷料体体_Competition joints, shoulder joints, knee joints, active joint implants: material health 'A body can contain other materials such as shouting silk heads and spiders - breast implants: (four) contain various transformations f homeopathic collagen, hydrogel. γ安...日乂尥量量-muscle implants: soft tissue using materials such as enamel resin and collagen large-corneal and retinal implants containing oxime, gel, pla-skin implants containing HA gel , collagen, epoxy resin. - or familiar with the know-how of the artisan. Examples Implantable medical devices include, but are not limited to, esthetic stents, catheters, general sutures, implantable rhythm regulators, microparticles, probes, and vascular grafts. Medical devices can be used alone or in combination. The term "delivery system" is used to provide any means of transferring the differentiated fetal cells or the differentiated fetal cell products themselves or to the implants to treat any metallic or commonly used orthopedics, wounds,颚面的, = synthetic implant material, hydrogel, silicone resin or graft. "', 2 This is a new finding that has been associated with the anti-chemical activity of differentiated and reduced cell transport from 9 to 16 weeks of pregnancy, preferably from 1 to 16 weeks and 12-14 weeks. Furthermore, with 16 weeks and more preferably from 9 to 16 weeks of pregnancy, preferably from 1 to 16 weeks, the biomaterial is compatible with adult fine-moon products (survival or death), and the co-plants and devices may depend on For delivery (see example below). Sterilization procedures are associated with poorly organized materials, surface degradation properties, and rapid appearance around the terminal/device. Implantation 9 to 16 weeks, preferably from 1〇V= The results of good function are obtained from the patient. The fetal cells can help the implant/device to regulate the differentiated inflammation from 12 to 14 weeks. In the early stage of pregnancy (ff repair and adaptation to avoid the 12-14 police Preferably, the period from 10 to 16 weeks and more preferably the fibroblast formation knife 5 is in the tissue repair _ inhibition muscle / Mai set sister Nis Β ^ Β Β 并 并 并 并 并 并 抑制 抑制 抑制 抑制 抑制 抑制 抑制 抑制 抑制 , , , , , , , , , Muscles and sputum cells are overextended in the extracellular matrix and play an important role in smooth muscle ii. The possibility of fibrosis should be directed to the resident area of the mycoplasma tissue repair area and/or the myofibroblasts that bind to the surface of the implant. The inter-f stem cells and the placenta stem cells represent the alpha_SMA of the cells of the non-myofibroblast type. Positive. These cells are key elements in the formation of fatigue and fibrous tissue (Estes et al, Differentiati〇n,
J Ι"，56。73-：!8”。由糾人纖維母細胞去分化成肌纖維母細胞的’ 型態及其在收縮能力方面的内在差異，胎兒及成人纖維母細胞的 差異係胎兒纖維母細胞為何不能產生纖維化反應及在胎兒受傷治 癒期間沒有傷疤組織的主要機制之一的關鍵要素(M〇ulin et al.， Cell Physiology, 188:211-222, 2001) ° 得自懷孕9至16週、較佳地自10至16週且更佳地自12-14 週的已分化胎兒皮膚細胞不輕易去分化，且不具有肌纖維母細胞 表型的型態（圖11)。已在特定專用的細胞庫中發展的這些細胞族 群與以上機制中的胎兒間質皮膚細胞及間質幹細胞非常不同（見實 例4)。alpha-SMA之表現顯示出表現於早期間質細胞族群中的胚 胎發展(C16ment et al·，J of Cell Science，120:229-238,2006)。 當12-14週或10-16週之間或9-16週的胎兒皮膚細胞在只包 11 201219573 各=作伴隨生物材料、基質及植體(即pLA、勝原蛋白泡棉、 凝膠、塑膠)之生長因子的胎兒血清的培養基中培養時，其不去分 化成肌纖維母細胞，亦不表現alpha_SMA(見圖1〇)。這些細胞不造 成材料周圍的收縮或硬化。成人間質幹細胞不與基質、植體材料 結合或合併’且去分化成在基質/植體界面形成累積的肌纖維母 細胞表型。這些細胞亦具有肌纖維母細胞之表型，且在其生長方 =叉到改變(本質上凋亡）。具體與細胞類型及細胞寄存方式有關的 這些巧鍵差異，顯示細胞選擇及懷孕時期的重要性(見如圖9)。 當隨著時間培養於2D培養物中時，如胚胎幹細胞的間質細胞 及其他未分化族群的細胞去分化，並顯示肌纖維母細胞表型。肌 纖維母細胞係惡名昭彰地涉及產生實心體植體周圍的組織收縮及 因此涉及纖維化組織反應(Siggeik〇w et ai，Bi〇materials，24: 1101-1109, 2003)。吾人一致同意肌纖維母細胞為涉及間隙基質累 積、結構變形、及逐步纖維化情況的細胞類型(Bames and G〇rin， Kidney Int., 79: 944-956, 2011 ° 當植體或基質被置入體内時，具有尤其伴隨軟組織、但亦伴 隨硬骨及軟骨的形成纖維組織之反應。申請人已見到得自懷孕9 至16週、較佳地自1〇至16週且更佳地自12_14週的已分化胎兒 皮膚細胞藉由消除或限制植體(或輸送系統)周圍的纖維組織形成 而使得外來物更具可接受性，見以下實例1_4。 申請人已發展出不侷限於此的下列應用： -由以各種劑量施用在各種輸送系統中的已分化胎兒細胞自 身引起的抗纖維化活性； -利用已分化胎兒細胞、純株或其碎片的植體及/或輸送系統 之塗層； -在( 褒孕9至16週，較佳地在1〇至16週且更佳地在12-14 週的已分化胎兒細胞與基質的組合，用以取代傳統飼養層來製備 用於病患自體移植的「生物活性的輸送系統」（用以自體移植的皮 膚移植物輸送之實例在以下實例1中顯示）。這亦已於如硬骨、軟 骨、肌肉、盤、肌腱及肺臟的其他組織類型之情形中顯示。 12 ⑧ 201219573 16週I 東乾燥的懷孕9至16週(較佳地在懷孕10至 • 4週)之已分化胎兒細胞(見以下實例3及圖8 if性4效性）。經由冷;東乾燥處理的已分化軌細胞之安 項“對於「傳統藥物」而言提供用於植體塗層的令人關注 在⑺療性理由方面，使用已分化胎兒細胞之重要優點 $ H滩(p&immunoincompe㈣且與此細胞之接受 者中引么免疫反應的能力降低有關。 ㈣ϋΐΐ施例中，可藉由在谬原蛋白或水凝膠内混合、組合、 =s反覆吸注、植覆(seeding)、平板培養(plating)、或放置細 胞而發生懷孕9至16週、較佳地在10至16週且 週的已，化胎兒細胞與膠原蛋白或水凝膠之整合。 用香「整合」或「整合於」用以描述任何混合的手段豆 有關巧細胞添加至基f。_語包含但純限於]合、植合： 以吸罝官反覆吸注、植覆、平板培養、或放置。 、、 本技藝者將察覺可運用任何結合手段 原蛋白賴三維基質)為馬膠原蛋白基質， κ把例中為HA組成物之水凝膝。 Λ另包含依據本發明的得自9至16週、較佳地自 更佳地自m4週姑娘的已分化胎兒細胞產物的生物 f可藉由下列方式加以製備：自捐贈者胎兒組織獲 兒組織發展細胞系；使胎兒組織生長並將胎兒細胞 “基物的細胞庫；及使移植物與膠原 & ^ ^又另、悲樣中，包含依據本發明的在懷孕9至16週、較佳 相六wfiL週且更佳地在12-14週之已分化胎兒細胞產物的生物 ΪΓίΐί可藉由下列方式加以製備：獲得胎兒細胞；使胎 蛋田胞或已分化胎兒細胞產物與朦原 在基本目的方面，本發明有關使用於預防、限制或治療有關 13 201219573 研究對^中之植體材料或輪送系統的纖維化組織反應之方法中的 生物相容性組成物。本發明之生物相容性組成物包含得自懷孕 9-16週的已分化胎兒細胞產物。較佳而言，本發明之該生物相容 性組成物在於得自懷孕9-16週’較佳地自懷孕ι〇_16週的已分化 的胎兒細胞產物’及更佳地得自懷孕12_14週的已分化胎兒細胞產 物。 、尤其’「已分化胎兒細胞產物」由已分化胎兒細胞、其衍生物 或碎片所組成。已分化胎兒細胞之衍生.物可包含生長因子、細胞 激素、蛋自質、DNA、RNA、細缝、或全細胞產物。已分化胎 兒細胞產物之碎片係定義為如細胞胞器、細胞壁及細胞膜。 該胎兒細胞為已分化的存活或死亡細胞。已分化胎兒細胞來 自特定組織，例如：來自胎兒皮膚真皮的胎兒皮膚纖維母細胞、 來自胎兒長骨的胎兒硬骨細胞、來自硬骨終板或活動關節的胎兒 軟骨細胞、來自胎兒肌腱的胎兒肌腱細胞等。當展 胞族群置於細齡化專用的鱗基憎，特定細胞類型佔^優 勢^將相同的胎兒細胞族群置於非適用於其分化的另一培養基 中時’並無整健群之妓化。置於成纽或成雜培養基中的 胎兒皮膚纖騎細胞以整體族群來說將不去分化成造#細胞 肪細胎.。 .在本發明之第一實施例中，得自懷孕9至16週、較佳地自十 孕10至16週的已分化胎兒細胞且更佳地得自懷孕12_14週的已j 化胎兒細胞係源自單一器官捐贈。抑或是’本發明之已分化 兒細胞可源自體外培養。 典本發明之已分化胎兒細胞或其碎片較佳地係選自皮膚、^ 骨、肌肉、盤、肺臟或肌腱細胞。該已分化胎兒細胞通$ 為新鮮或冷凍乾燥的胎兒細胞。 _ 關中，包含得自鮮9至16週的已分化的胎u 物’及較佳地得自懷孕1()至16週且更佳地自12_14週的 化胎兒細胞產物的生物相容性組成物，進一步包含由任何合邊 自然材料所製成的三絲質(如上述）。健而言，其巾整合依據々J Ι",56.73-:!8". The type of dedifferentiation of myofibroblasts into myofibroblasts and their intrinsic differences in contractile capacity, the difference between fetal and adult fibroblasts is fetal fiber The key element of why mother cells fail to produce fibrosis and one of the main mechanisms of scar tissue during fetal wound healing (M〇ulin et al., Cell Physiology, 188:211-222, 2001) ° from pregnancy 9 to The differentiated fetal skin cells at 16 weeks, preferably from 10 to 16 weeks and more preferably from 12 to 14 weeks, do not readily dedifferentiate and do not have a myofibroblast phenotype (Figure 11). These cell populations developed in dedicated cell banks are very different from fetal interstitial skin cells and mesenchymal stem cells in the above mechanisms (see Example 4). The performance of alpha-SMA shows embryo development in early mesenchymal cell populations. (C16ment et al., J of Cell Science, 120: 229-238, 2006). When 12-14 weeks or 10-16 weeks or 9-16 weeks of fetal skin cells are only included in the package 11 201219573 Biomaterials, substrates and implants (ie pLA, Shengyuan) When cultured in fetal serum of white foam, gel, plastic) growth factor, it does not dedifferentiate into myofibroblasts and does not exhibit alpha_SMA (see Figure 1〇). These cells do not cause contraction around the material or Hardening. Adult mesenchymal stem cells do not bind or merge with matrix, implant material and dedifferentiate into a myofibroblast phenotype that accumulates at the matrix/implant interface. These cells also have a phenotype of myofibroblasts, and Growth side = fork to change (essentially apoptotic). These differences in cell type and cell registration patterns indicate the importance of cell selection and pregnancy (see Figure 9). In 2D cultures, cells such as interstitial cells of embryonic stem cells and other undifferentiated populations dedifferentiate and display a myofibroblast phenotype. The myofibroblastic cell line is notoriously involved in producing tissue contractions around solid implants and thus Involving fibrotic tissue response (Siggeik〇w et ai, Bi〇materials, 24: 1101-1109, 2003). We agree that myofibroblasts are involved in gap-based Cell types of mass accumulation, structural deformation, and progressive fibrosis (Bames and G〇rin, Kidney Int., 79: 944-956, 2011 ° When implants or stroma are placed in the body, especially with soft tissue, However, it is also accompanied by the formation of fibrous tissue by hard bones and cartilage. Applicants have seen differentiated fetal skin cells from 9 to 16 weeks of pregnancy, preferably from 1 to 16 weeks, and more preferably from 12 to 14 weeks. Elimination or restriction of fibrous tissue formation around the implant (or delivery system) makes foreign objects more acceptable, see Example 1-4 below. Applicants have developed the following applications not limited to this: - anti-fibrotic activity caused by the differentiated fetal cells themselves administered in various delivery systems at various doses; - utilization of differentiated fetal cells, pure strains or fragments thereof a coating of the implant and/or delivery system; - in combination with a differentiated fetal cell and matrix at 9 to 16 weeks, preferably between 1 and 16 weeks, and more preferably between 12 and 14 weeks A "biologically active delivery system" for autologous transplantation of patients is prepared in place of a conventional feeder layer (an example of skin graft delivery for autologous transplantation is shown in Example 1 below). It is shown in the case of cartilage, muscle, disc, tendon and other tissue types of the lungs. 12 8 201219573 16 weeks I Divided fetal cells from 9 to 16 weeks of pregnancy in the east (preferably 10 to 4 weeks of pregnancy) See Example 3 below and Figure 8 if the 4th effect). The safety of the differentiated orbital cells via cold; East drying treatment "for the "traditional drugs" to provide attention for implant coatings in (7) treatment For sexual reasons, use differentiated fetal cells The important advantage of $H Beach (p&immunoincompe (4) is related to the reduced ability of the recipient of this cell to induce an immune response. (4) In the case of the application, it can be mixed, combined, and repeated in the prion protein or hydrogel. By sucking, seeding, plating, or placing cells for 9 to 16 weeks, preferably 10 to 16 weeks and weeks, the fetal cells are combined with collagen or hydrogel. Integration. Use "integration" or "integration" to describe any mixed means of adding beans to the base f. _ Included but purely limited to, combined, planted: repeated sucking and mulching , plate culture, or placement., the skilled artisan will perceive the use of any combination means the original protein to the three-dimensional matrix) for the horse collagen matrix, κ in the case of the HA composition of the water condensation knee. The inventive organism f from 9 to 16 weeks, preferably from the differentiated fetal cell product of the m4 week girl, can be prepared by obtaining a tissue development cell line from the donor fetal tissue; Fetal tissue growth and fine fetus a cell bank of the substrate; and the graft and collagen & ^^, in another sad form, comprising 9 to 16 weeks of pregnancy, preferably 6 weeks of wfiL weeks, and more preferably 12- The biological process of the differentiated fetal cell product of 14 weeks can be prepared by obtaining fetal cells; making the fetal egg cell or the differentiated fetal cell product and the progenitor in terms of basic purpose, the invention relates to prevention, restriction Or treating a biocompatible composition in a method for studying the fibrotic tissue response of an implant material or a delivery system of 13 201219573. The biocompatible composition of the invention comprises from 9-16 weeks of pregnancy. The differentiated fetal cell product. Preferably, the biocompatible composition of the present invention resides in a differentiated fetal cell product obtained from 9-16 weeks of pregnancy, preferably from pregnancy 〇 16 weeks, and more preferably from pregnancy 12-14. Weekly differentiated fetal cell product. In particular, the "differentiated fetal cell product" consists of differentiated fetal cells, derivatives or fragments thereof. Derivatives of differentiated fetal cells may comprise growth factors, cytokines, egg mass, DNA, RNA, nicks, or whole cell products. Fragments of differentiated fetal cell products are defined as, for example, cell organelles, cell walls, and cell membranes. The fetal cells are differentiated viable or dead cells. The differentiated fetal cells are derived from specific tissues, such as fetal skin fibroblasts from fetal skin dermis, fetal hard bone cells from fetal long bones, fetal chondrocytes from hard bone endplates or moving joints, fetal tendon cells from fetal tendons, etc. . When the population of the squamous cell is placed in a squamous squamous squamous cell, the specific cell type is dominant. When the same fetal cell population is placed in another medium that is not suitable for its differentiation, . Fetal skin fibrosus cells placed in adult or adult medium will not differentiate into adult cells in the overall population. In a first embodiment of the invention, the differentiated fetal cells obtained from 9 to 16 weeks of pregnancy, preferably from 10 to 16 weeks of pregnancy, and more preferably from 12 to 14 weeks of pregnancy, have been obtained. From a single organ donation. Or, the differentiated babies of the invention may be derived from in vitro culture. The differentiated fetal cells or fragments thereof of the invention are preferably selected from the group consisting of skin, bone, muscle, disc, lung or tendon cells. The differentiated fetal cells pass $ for fresh or freeze-dried fetal cells. _ Guanzhong, comprising a biocompatible composition of differentiated fetal urinary material from fresh 9 to 16 weeks and preferably obtained from pregnant 1 () to 16 weeks and more preferably from 12 to 14 weeks. And further comprising a trifilament made of any natural material of the edging (as described above). In terms of health, its towel integration basis
14 201219573 杳明的已分化胎兒細胞產物的三維基質係選自膠原蛋白基質 匕LA、PLGA、PEG、雜蛋白、包含ha的水凝膠、石夕氧樹脂： 4丁聚料其混合物。本發明中所用的三絲質可取決於其應用 „為可生物賴或非可生物轉。顯而m敎的 為無實質上毒性，且應為藥物上可接受。 可藉由三維組織及/或細胞及相關的生長因子而表現的生物 性質包含但不侷限於預防及/或減少組織纖維化反應、組織重 塑、促進上皮形成、組織生長、血管形成及/或血管新生。因此， 對研究對象提供用以促進組織重塑、上皮形成、組織生長、血管 形成及/或血管新生之方法亦為本發明之目的，該方法包含將如 上述的本發明之生物相容性組成物施加至該研究對象。若具有纖 ，化^應的組織修復獲得預防，則因血管可遷移，故正常組織修 復不文阻礙反而在關於上皮形成、.組織生長、血管形成及/戋血 管新生方面獲得提昇。 通常而言，產生纖維化組織反應的植體材料係選自牙齒植 體、牙齒增大物、髖關節、肩關節、膝關節、活動關節植體、乳 房植體、肌肉植體、可植人的f療裝置、或包含皮膚及肌肉的任 何軟組織。其它軟組織可為心臟、肺臟、腦部、腎臟、胰 以上定義）。 v A在另一方面，產生纖維化組織反應的輸送系統係選自金屬性 或常用整形外科、鑛、麵的自然或合成植體材料、水凝膠、 幾丁聚醣、石夕氧樹脂或移植物。 如此處所使用的本發明之「生物相容性組成物」可含有來自 懷孕9-16週^較佳地自10_16週的一或更多已分化胎兒細胞，且 更佳地源自懷孕12-14週的已分化的胎兒細胞；伴隨其他化學成分 包含但不侷限於：傳統藥物、生理上適合的載體及辅藥。抑或是， 本發明之生物相容性組成物可包含得自懷孕9至％週及較佳地自 10至16週的一或更多已分化胎兒細胞，且更佳地得自懷孕12_14 巧的已分化胎兒細胞及/或已受到安定化的一或更多胎兒蛋白， 伴隨其他化學成分包含但不侷限於：傳統藥物、生理上適合的载 15 201219573 體及辅藥。 此成分協助促進對研究對象施用蛋白質及/或細胞。本發明 之生物相容性組成物可藉由本麟領域所熟知練序加以製造， ^利用習用的混合、溶解、粒化、裹糖衣、磨細、乳化、膠 ，封裝、包覆或冷;東乾騎理。潍成分的配製及施用的技術可 於併於，匕處作為參考的"Remingt〇n· The §cience and practice 〇f14 201219573 The three-dimensional matrix of differentiated fetal cell products of the present invention is selected from the group consisting of collagen matrix 匕LA, PLGA, PEG, hybrid proteins, hydrogels containing ha, and diarrhea: 4 butyl aggregates. The trifilament used in the present invention may be biologically or non-biotransducible depending on its application. It is substantially non-toxic and should be pharmaceutically acceptable. It can be composed of three-dimensional tissue and/or Biological properties expressed by cells or associated growth factors include, but are not limited to, prevention and/or reduction of tissue fibrosis, tissue remodeling, promotion of epithelialization, tissue growth, angiogenesis, and/or angiogenesis. It is also an object of the invention to provide a method for promoting tissue remodeling, epithelialization, tissue growth, angiogenesis and/or angiogenesis, the method comprising applying to the biocompatible composition of the invention as described above The research object. If the tissue repair with fiber and the chemical is prevented, the blood vessels can migrate, so the normal tissue repair is not hindered, but it is improved in terms of epithelial formation, tissue growth, blood vessel formation and/or angiogenesis. In general, the implant material that produces the fibrotic tissue response is selected from the group consisting of dental implants, tooth enlargements, hip joints, shoulder joints, knee joints, and active joints. Implants, breast implants, muscle implants, implantable devices, or any soft tissue containing skin and muscles. Other soft tissues can be defined for heart, lung, brain, kidney, and pancreas.) In one aspect, the delivery system that produces the fibrotic tissue response is selected from metallic or conventional orthopedic, mineral, facial natural or synthetic implant materials, hydrogels, chitosan, lycopene or grafts. The "biocompatible composition" of the present invention used in the field may contain one or more differentiated fetal cells from 9-16 weeks of pregnancy, preferably from 10-16 weeks, and more preferably from 12-14 weeks of pregnancy. The differentiated fetal cells; with other chemical components including but not limited to: traditional drugs, physiologically suitable carriers and adjuvants. Or, the biocompatible composition of the present invention may comprise one or more differentiated fetal cells obtained from 9 to % weeks of pregnancy and preferably from 10 to 16 weeks, and more preferably from pregnancy 12-14. Differentiated fetal cells and/or one or more fetal proteins that have been stabilized, along with other chemical components including, but not limited to, traditional drugs, physiologically suitable carriers, and adjuvants. This component assists in the administration of proteins and/or cells to the subject. The biocompatible composition of the present invention can be produced by the well-known practice in the field of the present invention, using conventional mixing, dissolving, granulating, sugar coating, grinding, emulsifying, gluing, encapsulating, coating or cold; Dry riding. The technique for the preparation and application of bismuth ingredients can be used as a reference. "Remingt〇n· The §cience and practice 〇f
Pharmacy" Lippincott Williams & Wilkins Publishing Co., 20th edition。 其他活性劑亦可被包含於本發明之「生物相容性組成物」中， 例?：抗發㈣、止鋪、抗微生侧、抗黴_、抗生素、維 他命、抗氧化劑。 生物相容性組成物之「有效」或「治療上有效」劑量意指在 伴Ik任何醫學棘的之合理效以驗比例下提供鮮效果的無毒 ，但充分之魅。舰效果可紐和或驗疾病的症候、徵候、 或原因、或生物系統的其他任何期望改變。 热悉本技藝者將理解，用來製成本發明之三維生物相容性組 成物的得自懷孕9至16週及較佳地自10至16週的已分化胎兒細 胞、且更佳地源自懷孕12-14週的已分化胎兒細胞，可包含：用於 皮膚移植的已分化胎兒皮膚細胞、用於肌肖移植的已分化胎兒肌 肉細胞、用於硬骨移植的已分化胎兒硬骨細胞、用於肌鍵移植的 已分化胎兒肌腱細胞、及用於軟骨移植的已分化胎兒軟骨細胞且 更具體之=於關節軟骨移植的已分化的月台兒關節軟骨細胞。 在一態樣中，本發明提供包含與膠原蛋白基質(如馬膠原蛋白) 整合之得自懷孕9-16週及較佳地自懷孕1〇_16週的已分化胎兒細 胞’且更佳地得自懷孕12-14週的已分化胎兒細胞的三維皮膚組織 異體移植建構物。舉例而言，得自懷孕9_16週及較佳地自1〇至 16週的已分化胎兒細胞，且更佳地得自懷孕12_14週的已分化胎 兒細胞可為例如在適當培養條件下可分化成真皮纖維母細胞及上 皮角質細胞者的胎兒皮膚細胞。整合可藉由熟悉本技藝者所熟知 的包含但不侷限於··混合、組合、以吸量管反覆吸注、植覆了平 201219573 ίΐί祕f放置的倾方法加以達成。本建難而射用以治療 罹桃域反應雜、失贼赫之研究縣。 ’、 产理if 之已分化胎兒細胞的細胞庫，在依據CHUV的 二i 及政策的鮮中止之後立即取得來自胎兒組織 ，分成在複數個1G em2的組織培養皿中則H Dulbecco, MEM (DMEM)^,,^ ^ 培4皿。#細胞生長進行時，如約-週後，組織及細 立^騰St酶作用。而後將-些培養皿如麵態氮巾冰殊成獨 =胞ΚΓ兒細胞離心並重新懸浮以自培養物產生大略均 其次’將細胞和緩地與防凍劑(如DMEM5ml+胎兒小牛血清 4ml+二甲基亞砜(dimethylsulf〇xide，DMS〇) i禪昆合。然後將胎 ，細胞懸#體㈣於等分巾，絲例如液態氮巾财。在一較佳 貫施例中’等分係j^C/min之速率、纟綠，朗其達到_8()()c之溫 度’而後在約24小時後轉變至_160〇c。藉纟開啟胎兒細胞庫的安 ，、融化内容物、並轉移至一般細胞培養基内，同質儲存的細胞 等^而後可用於如三維組織異體移植建構物及/或本發明之生物 相谷性組成物之生產的任何預期目的，或用於製且苴 求之研究對象中的纖維化組織反應缺陷的藥物。Ί、南 為了製成本發明之生物相容性組成物，可取得來自胎兒捐贈 者的組織之檢體，並如上述内容發展細胞系。舉例而言，可將胎 兒組織分成複數個1〇 cm2的組織培養皿中的小碎片，該等組織培 養皿已製備成具有利用手術刀產生之切口。將具有麩醯胺酸及 1^2胎兒血清的DMEM組織培養基加至培養皿◊胎兒細胞擴張成 咼濃度以產生細胞庫。而後將胎兒細胞冷凍於DMS〇、DMEM& 胎兒血清之混合物中直到需要時。 來自第5-10代的胎兒細胞係製備成約5 3 χ 1〇3細胞/毫升的濃 度。此濃度可取決於纖維組織反應缺陷的類型及病患是否為成人 或小孩而加以改變。舉例而言，細胞濃度可在自約9.3 χ 1〇2細胞/ 17 201219573 細胞/毫升的範圍。而後使胎兒蛋白質安定且以 早蜀^式或與月口兒細胞結合而併入載體(三維基質）中。 仔自懷孕9-16週及較佳地自懷孕1〇_16週的已分化胎兒細 更佳2地得自懷孕m4週的已分倾兒細胞係由來自腹部粗 織的lcm2之皮膚加以製備。 股丨、，且 平例而言，將胎兒組織分成在複數個1〇 cm2的組織培養皿中 =小碎片，且於具有麩醯胺酸的Duibec⑺，①廳以)組 基中生長。而後將胎兒血清加至培養皿。當細胞生長進行 如約-週後’組織及細胞受到胰蛋白酶作用。而後將一些培養瓜 如在液態氮中冰凍成獨立單元。然後將胎兒細胞離心並重新懸 以自培養物產生大略均勻的細胞懸浮體。 其次，將細胞和緩地與防凍劑(如DMEM5ml+胎兒小牛血清 4ml+ 二甲基亞颯(dimethyisulf0xide，DMS〇)匕喊合。而後二 兒細胞懸浮體密封於等分巾，並如在雜氮巾財。在—較佳二 施例中’等分以TC/min之速率結來，直到其達到_8〇〇c之溫产、， 而後在約24小時後轉變至_160eC。藉由開啟胎兒細胞庫的安 融化内容物、並轉移至一般細胞培養基内，同質儲存的細胞等分 而後可用於如三維組織異體移植建構物及〆或本發明之生物相容 性組成物之生產的任何預期目的，或用於製造治療具有其需求 研究對象中的纖維化組織反應缺陷的藥物。 八 對於本發明之目的而言，各種施用生物相容性組成物之 徑均有可能。 本發明之另一目的為k供用以治療與研究對象中之植體材料 或輸送系統有關的纖維化組織缺陷的方法，該方法包含將如上述 的本發明之生物相容性組成物置入位於該纖維組織缺陷内或鄰近 於該纖維組織缺陷的該植體材料或輸送系統。 本發明之進一步目的為用以限制及/或預防與研究對象中之 植體材料或輸送系統有關的纖維化組織反應的方法，該方法包含 具有本發明之生物相容性組成物的該植體材料或輸送系統之^ ⑧ 層0 18 201219573 ㈣ίϋ之i—目標為治療或預防研究對象罹患纖維組織反應 研究對氣女t方ϋ含將本發明之生物相容性組成物施加至該 或輸送系統纖維化組織反應失調係較佳地源自植體材料 之生物相容性經成物使用在製造用於治療或預 材料或輪送系統或移植物的纖維化組織反應失 十，fun的ΐ法提供數種技術性及生物性的優點。舉例而 i，且更刀佳地即得自懷孕9_16週及較佳地自懷孕1(Μ6 ί主接i t=12·14獅__細齡衫疫上可受 乂义=個$s*贈對於細胞庫產生及檢驗有所必要。 Φ ^fr m >fS 〇 3所有此種變化及修改。本發明亦包含本說B月奎 有變化包使來自均等之意纽翻内的所 為參ΐ朗各種辦文獻，其各者健義於此處作 士旅^上描述參打列實鑛更完伽解。缺而H為杆 本發明之方法的例示，且並非意欲舰I明之^圍此貝例為心 實例 實例1 在「敎細胞塗層」基質上附著及運送的成人角質細胞移植 19 201219573 物: 、角質細胞的自體移植物傳統上作為必須利用約12個鈦夾及/ 戴^蛋白_著至紗布以能將這些配製轉移至病患的薄層细 t 為飼養層㈣丽€絲械狀械蚊纖維母細 胞或包皮纖維母細胞上)(圖2)而加以輸送。更有甚者，如 配製物非常脆弱，且與供細胞形成輸朝之整體細 肊層的培養時間有關。成人細胞通常不輕易地黏附至如膠原蛋白 =棉這就是為何迄今尚未將翻來安定角質細胞 込養。月口兒細胞輕易與如膠原蛋白的植體材料 黏合^見® 3，PLA表面上之胎兒細胞的SEM圖顯示完全^著口)物 藉由使用具杨兒皮膚細胞(不論存活或死 ίίΪΪ，、或聚合物’這使得基質能生物相容於成人 角質、·田胞生長並貫施對於病患更穩定的自體移植物輸送。 結論： 产ϋ胎ΐί”胞之基質塗層提供所有類型的利用諸如皮 膚、軟月、硬月、角膜之細胞的自體移植技術之輸送。 的太Ϊ兒「生物相容」於成人細胞並具有抗纖維化 的本質。、k並非早獨基質的情況(即成人角質細胞不輕易附著）。 -成人細胞附著 -受控制的經由表面之—致分布(無法彻本f上黏稠的喷 務） -當病患傷口基部準備好時，移植物就緒 實例2 應 由月。兒細胞及/或碎片所提供之龍材料的抗纖維化反 处生Ϊ多=維組織並產生如植體鬆他及不穩定的機 月b φ 疋；體/、軟組織的交互作用及纖维化反;s。纖维 組織可形成如圍繞植體材料的歸m 4 化反f W念隹八♦展(圖親察_關圍之實心 白色饴市刀帶及圓圈内之非常少至無硬骨生長）0 20 ⑧ 201219573 士當將單獨生物材料(PLA)插入綿羊股骨頭中的硬骨缺陷中 組織回應外來物質而形成於鋪周圍。若胎兒細胞與進 帛手杈型中之硬骨缺陷的相同植體材料結合，則沒有觀察到纖 維組織定位雜復(圖5，整體财硬骨形成，且無麟組織形= 實例3 在軟組織中的基質中所輸送的胎兒細胞之抗纖維化反應： 、許多植體材料、裝置、骨架在如皮膚及肌肉的軟組織中產生 纖巧化組織累積。當如皮膚的軟組織受傷時，於修復過程中可具 有壞死性、纖維化組織反應。當前期的纖維化組織已於皮膚移植 法^前出現時，吾人已見到與基質/植體結合的胎兒細胞可降低 或消除纖維化組織反應。這可見於肌肉及皮膚軟組織中（見圖6)及 大型表面皮膚重建中（見圖7)。重要地，就比較血管新生活性而言， 胎兒細胞之生物活性可藉由將細胞及/或碎片冷凍/冷凍乾燥(見 圖8)來加以安定化。 實例4 特定細胞類型之間的差異為伴隨懷孕12-14週(或9-16週、較 佳地10-16週)的已分化胎兒皮膚細胞所見的抗纖維化反應的原 因。未分化之特定細胞的胎兒間質皮膚細胞(MSC)可輕易地分化成 複數個細胞譜系，且具有與alpha- SMA表現有關的肌纖維母細胞 表型(Hinz，JBiomechanics，43: 146-155, 2010)。MSC 細胞族群亦具 有alpha- SMA，且與傷疤及纖維組織反應中之關鍵要素的肌纖維 母細胞有關(Comut et al·，Biomaterials，21: 1887-1896, 2000)。懷孕 12- 14週(或9-16週)的已分化胎兒皮膚細胞為難以去分化的已分 化細胞，且當與基質或與植體材料結合時不造成纖維化反應。當 懷孕12-14週或9-16週之間的胎兒皮膚細胞與生物材料、基質及 植體(即PLA、膠原蛋白海綿、HA凝膠、塑膠)一起培養時，其不 去分化成肌纖維母細胞，亦不表現alpha- SMA。這些細胞不造成 材料周圍的收縮或硬化。成人間質幹細胞不與基質、植體材料結 21 201219573 合或整合’且去分化成在基質//顧介面形成?、積的肌纖維母細 胞表型。具體與細胞麵及細胞寄存方式有關的這些瞻差異， 顯示細胞選擇及懷孕時期之重要性。 當隨著時間培養於2D培養中時，間質細胞及如胚胎幹細胞的 其他細胞之未分化族群去分化，並顯示肌_母細胞表型。肌纖 維母細胞惡名崎地涉及產生實心體植體周_組敵縮及因此 涉及纖維化組織反應(Siggelkow et al.，Bi〇materials，2令 11〇1_11〇9, 2003)。吾人一致同意肌纖維母細胞為涉及間隙基質累積、結構變 形、及逐步，纖維化情況的細胞類型(Bames _ G〇rin，⑺如办Μ 79: 944-956, 2011； Valentin et aL, J Bone Joint Surgery 88* 2673-2686,2006)。 g ^ ' 肌纖維母細胞型態及其在收縮能力方面的固有差異，係為何 .胎兒纖維母細胞不能產錢靴反應及在胎兒創絲細間沒有 傷疤組織的主要機制之一的關鍵要素^河⑽丨化et此，了 cellPharmacy" Lippincott Williams & Wilkins Publishing Co., 20th edition. Other active agents may also be included in the "biocompatible composition" of the present invention, for example: anti-hair (four), stop-stop, anti-microbial side, anti-mildew, antibiotics, vitamins, antioxidants. The "effective" or "therapeutically effective" dose of a biocompatible composition means a non-toxic, but full-feeling effect that provides a fresh effect at a reasonable rate with any medical spine of Ik. The ship's effect can be used to determine the symptoms, signs, or causes of the disease, or any other desired changes in the biological system. It will be appreciated by those skilled in the art that differentiated fetal cells from 9 to 16 weeks of pregnancy, preferably from 10 to 16 weeks, and more preferably from the three-dimensional biocompatible composition of the present invention are derived. Divided fetal cells of 12-14 weeks of gestation may include: differentiated fetal skin cells for skin transplantation, differentiated fetal muscle cells for myocardium transplantation, differentiated fetal hard bone cells for hard bone transplantation, for Muscle-transplanted differentiated fetal tendon cells, and differentiated fetal chondrocytes for cartilage transplantation and more specifically = differentiated platform articular chondrocytes of articular cartilage transplantation. In one aspect, the invention provides a differentiated fetal cell comprising from a pregnant 9-16 weeks of pregnancy and preferably from 1 to 16 weeks of pregnancy integrated with a collagen matrix (eg, horse collagen) and more preferably A three-dimensional skin tissue allograft construct from differentiated fetal cells from 12-14 weeks of gestation. For example, differentiated fetal cells derived from 9-16 weeks of gestation and preferably from 1 〇 to 16 weeks, and more preferably from 12-14 weeks of gestation, can be differentiated, for example, under appropriate culture conditions. Fetal skin cells of dermal fibroblasts and epithelial keratinocytes. Integration can be achieved by familiar methods known to those skilled in the art including, but not limited to, mixing, combining, pipetting with a pipette, and planting a flattening method. This construction is difficult to shoot and used to treat the research area of the Taotao domain. ', the cell bank of the differentiated fetal cells of the production of if, immediately after the fresh stop according to the CHUV II and policy, from the fetal tissue, divided into a number of 1G em2 tissue culture dish H Dulbecco, MEM (DMEM ) ^,, ^ ^ 4 dishes. # When cell growth is carried out, for example, after about-week, the tissue is organized and the St-Enzyme is acted upon. Then, some of the culture dishes, such as the surface nitrogen mask, are made into a single cell, and the cells are centrifuged and resuspended to produce roughly the same amount from the culture. Secondly, the cells are gently mixed with antifreeze (such as DMEM 5ml + fetal calf serum 4ml + dimethyl Dimethylsulfoxime (DMS〇) i zen-kun. Then the fetus, cell suspension #4 (4) in the aliquot, silk, such as liquid nitrogen towel. In a preferred embodiment, 'equal system j The rate of ^C/min, 纟 green, lang reaches the temperature of _8()()c and then changes to _160〇c after about 24 hours. By opening the fetal cell bank, melting the contents, And transfer to a general cell culture medium, homogenously stored cells, etc., and then used for any intended purpose, such as the production of a three-dimensional tissue allograft construct and/or the biological phase cereal composition of the present invention, or for A drug having a defect in fibrotic tissue reaction in a subject. In order to prepare a biocompatible composition of the present invention, a specimen from a tissue of a fetus donor can be obtained, and a cell line can be developed as described above. In other words, the fetal tissue can be divided into a plurality of 1〇 Small pieces in a tissue culture dish of cm2, which have been prepared to have an incision produced by a scalpel. DMEM tissue culture medium containing glutamic acid and 1^2 fetal serum is added to the culture dish and the fetal cells are expanded. The concentration of the sputum is increased to produce a cell bank. The fetal cells are then frozen in a mixture of DMS 〇, DMEM & fetal serum until needed. The fetal cell line from passage 5-10 is prepared to be about 5 3 χ 1 〇 3 cells/ml. Concentration. This concentration may vary depending on the type of fibrous tissue reaction defect and whether the patient is an adult or a child. For example, the cell concentration may range from about 9.3 χ 1〇2 cells / 17 201219573 cells/ml. The fetal protein is then stabilized and incorporated into the vector (three-dimensional matrix) either early or in combination with the cells of the month. Aberdeen has differentiated from 9-16 weeks of pregnancy and preferably from 1 to 16 weeks of pregnancy. Fetal finer 2 The differentiated cell line obtained from the m4 week of pregnancy is prepared from the skin of lcm2 from the abdomen coarse weaving. The femoral hernia, and, in the ordinary case, the fetal tissue is divided into a plurality of 1〇cm2 Tissue culture In the dish, small fragments were grown and grown in Duibec (7), chamber 1 with glutamic acid. The fetal serum is then added to the culture dish. When the cells grow for about about week, the tissues and cells are subjected to trypsinization. Some cultured melons are then frozen into individual units in liquid nitrogen. The fetal cells are then centrifuged and resuspended to produce a substantially uniform cell suspension from the culture. Secondly, the cells are gently combined with antifreeze (such as DMEM 5ml + fetal calf serum 4ml + dimethyisulf0xide (DMS〇). Then the second cell suspension is sealed in an aliquot, and as in the aza towel In the preferred embodiment, the 'equal score is TC/min until it reaches the temperature of _8〇〇c, and then changes to _160eC after about 24 hours. The contents of the cell bank are thawed and transferred to a general cell culture medium, and the homogenously stored cells are aliquoted and then used for any intended purpose such as three-dimensional tissue allograft constructs and/or production of the biocompatible composition of the present invention. Or for the manufacture of a medicament for treating a defect in fibrotic tissue response in a subject in need thereof. VIII For the purposes of the present invention, various routes for administering a biocompatible composition are possible. Another object of the present invention Providing k a method for treating a fibrotic tissue defect associated with an implant material or delivery system in a subject, the method comprising placing a biocompatible composition of the invention as described above The implant material or delivery system within or adjacent to the fibrous tissue defect. A further object of the present invention is to limit and/or prevent fibrotic tissue reaction associated with the implant material or delivery system in the subject Method comprising the implant material or delivery system having the biocompatible composition of the invention. 8 18 201219573 (4) — ϋ — — — — — 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗 治疗The t-square containing the biocompatible composition of the present invention is applied to the or delivery system. The fibrotic tissue reaction disorder is preferably derived from the biocompatible composition of the implant material used in the manufacture for treatment or pre-treatment. The fibrotic tissue reaction of the material or the transfer system or the graft is lost. The method of fun provides several technical and biological advantages. For example, i, and more preferably, it is obtained from pregnancy for 9-16 weeks and preferably. Since pregnancy 1 (Μ6 主 main connection it=12·14 __ 细 衫 疫 = = = = = = $ $ $ $ $ $ $ $ $ $ $ $ $ $ $ $ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * All such changes and modifications The invention also includes the fact that the B-Kui has a change package to make the various documents from the equal meaning of the 纽 纽 内 , , , , , , , , , , , 各种 各种 各种 各种 各种 各种 各种 各种 各种 描述 描述 描述 描述 描述 描述 描述 描述 描述 描述 描述 描述 描述 描述H. H is the exemplification of the method of the present invention, and is not intended to be a case of the case. Example 1 Adult keratinocyte transplantation attached and transported on a "sputum cell coating" substrate 19 201219573 : Autologous grafts of keratinocytes have traditionally been used as a feeding layer for the feeding layer (four) of Lishi, which must use about 12 titanium clips and/or protein to the gauze to transfer these preparations to the patient. It is transported on the silkworm fibroblasts or foreskin fibroblasts (Fig. 2). What's more, such as the formulation is very fragile and related to the incubation time for the cells to form the whole fine layer. Adult cells usually do not easily adhere to such as collagen = cotton, which is why it has not yet been stabilized by keratinocytes. The cells of the moon are easily adhered to the implant material such as collagen. ^3, the SEM image of the fetal cells on the surface of the PLA shows the completeness of the skin by using the skin cells of the yang (regardless of survival or death). , or a polymer' which allows the matrix to be biocompatible with adult keratin, cell growth, and more stable autologous graft delivery to patients. Conclusion: Matrix coating of ϋ ϋ 胞 cells provides all types The use of autologous transplantation techniques such as skin, soft moon, hard moon, and corneal cells. The scorpion is "biocompatible" in adult cells and has an anti-fibrotic nature. (ie, adult keratinocytes are not easily attached.) - Adult cell attachment - controlled distribution through the surface (cannot be viscous) - When the patient's wound base is ready, the graft is ready for instance 2 The anti-fibrosis of the dragon material provided by the cells and/or fragments should be more than = 维 维 = 维 并 并 并 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植 植Role and fibrosis; s The fibrous tissue can form, for example, around the implant material, the anti-f W 隹 隹 ♦ ( (Figure _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 20 8 201219573 Shidang inserts a separate biological material (PLA) into the hard bone defect in the femoral head of the sheep. The tissue is formed around the shop in response to foreign matter. If the fetal cells are combined with the same implant material of the hard bone defect in the handcuffs No fibrous tissue localization was observed (Fig. 5, overall solid bone formation, and no skeletal organization = Example 3: Antifibrotic response of fetal cells transported in a matrix in soft tissue: , many implant materials, The device and the skeleton produce microscopic tissue accumulation in soft tissues such as skin and muscles. When the soft tissue such as the skin is injured, it may have a necrotic and fibrotic tissue reaction during the repair process. The current stage of fibrotic tissue has been applied to the skin grafting method. When we appeared before, we have seen that fetal cells combined with matrix/implant can reduce or eliminate fibrotic tissue reaction. This can be seen in muscle and skin soft tissue (see Figure 6) and large In facial skin reconstruction (see Figure 7). Importantly, in terms of comparing angiogenic activity, the biological activity of fetal cells can be stabilized by freezing/freezing the cells and/or debris (see Figure 8). Example 4 The difference between specific cell types is the cause of anti-fibrotic response seen with differentiated fetal skin cells with 12-14 weeks of gestation (or 9-16 weeks, preferably 10-16 weeks). The fetal mesenchymal skin cells (MSCs) of the cells can be easily differentiated into multiple cell lineages and have a myofibroblast phenotype associated with alpha-SMA expression (Hinz, J Biomechanics, 43: 146-155, 2010). The population also has alpha-SMA and is associated with myofibroblasts, a key element in scar and fibrous tissue responses (Comut et al., Biomaterials, 21: 1887-1896, 2000). Divided fetal skin cells of 12-14 weeks (or 9-16 weeks) of pregnancy are differentiated cells that are difficult to dedifferentiate and do not cause fibrotic reactions when combined with the matrix or with the implant material. When fetal skin cells between 12-14 weeks or 9-16 weeks of pregnancy are cultured with biological materials, matrices and implants (ie PLA, collagen sponge, HA gel, plastic), they do not differentiate into myofibroblasts. Cells also do not exhibit alpha-SMA. These cells do not cause shrinkage or hardening around the material. Adult mesenchymal stem cells do not integrate or integrate with the matrix and implant material 21 201219573 and dedifferentiate into the phenotype of the myofibroblasts formed in the matrix. These differences in specific aspects of cell surface and cell storage indicate the importance of cell selection and pregnancy. When cultured in 2D culture over time, the undifferentiated population of mesenchymal cells and other cells such as embryonic stem cells dedifferentiate and exhibit a muscle-mother cell phenotype. Myofibrillar cells are notoriously involved in the production of solid body implants, which are involved in fibrotic tissue reactions (Siggelkow et al., Bi〇materials, 2, 11〇1_11〇9, 2003). We agree that myofibroblasts are cell types involved in interstitial matrix accumulation, structural deformation, and gradual, fibrotic conditions (Bames _ G〇rin, (7) eg Μ 79: 944-956, 2011; Valentin et aL, J Bone Joint Surgery 88* 2673-2686, 2006). g ^ ' Myofibrillar cell type and its inherent differences in contractile capacity, why. Fetal fibroblasts can not produce cashmere response and a key element of one of the main mechanisms of scar tissue in fetal filaments (10) 丨化et this, the cell
Physiology，188:211-222, 2001)。在懷孕 9-16 週較佳地在 12_14 週 的已么:b胎兒皮膚細鮮㈣去絲，且*具有肌雜母細胞表 型之型態(圖11及12)。已發展於特定專用的細胞庫中的這些細胞 族群與以上補巾_躺f皮膚細胞關f幹細麟常不同。 alpha-, SMA之表現係顯示為表現於早期間質細胞族群中的胚胎發 展(Clement et al” J of Cell Science，120:229-238, 2006)。 【圖式簡單說明】 圖1顯不依據描述於此的一些實施例之提供生物相容性證據 的j由生物相容性馬膠原蛋白所組成之「胎兒細胞塗覆」基質上 附著及運送的成人角質細胞皮膚移植物。在圖丨中， 胞，而b)代表基質，即具有胎兒細胞之植體。 #自^田 —，2顯示正自塑膠組織培養瓶酵素性分離的人類角質細胞之 貫際薄層’其需要支持來如® 1巾所示㈣層形式或單獨細胞轉 移至病患。 圖3顯示依據描述於此之一些實施例的生物材料表面上的胎 ⑧ 22 201219573 兒細胞之SEM圖，其顯示出表面的生物相容性交互作用。 、圖4顯不纖維組織係形成於植體材料周圍。將單獨之生物材 料(PLA)插進硬骨缺陷中。伴隨單獨之生物材料的纖維組織形成： 觀察到圈周圍之實心白色密集分帶及圓圈内之非常少至無硬骨 生長。這描繪出依據描述於此的一些實施例之軟及硬組 體材料之纖維化反應。 圖5顯示促進硬骨形成之pLA中的胎兒細胞輸送至硬骨缺 陷。依據描述於此的一些實施例，整體僅有硬骨形成，且無纖維 組織形成，描繪出胎兒細胞及/或其碎片的抗纖維化活性。 圖6描繪出依據描述於此的一些實施例之在胎兒細胞輸送及 與膠原發泡體基質相容之後所見的自皮膚至肌肉之軟組織中的抗 纖維化活性。. 圖7顯示依據描述於此之一些實施例的在與胎兒細胞輸送及 與膠原發泡體基質相容之後的嚴重受傷之皮膚中的抗纖維化組織 累積。吾人觀察到利用胎兒皮膚細胞及膠原蛋白基質/植體 纖維化反應。 圖8描繪出依據描述於此的一些實施例之藉由冷凍乾燥/冷 凍以維持生物生產力及功能來使胎兒細胞及/或衍生物安定化。 生物活性在冷象乾燥之後受到保存。 圖9顯示展現出明顯細胞型態及染色的胎兒肌肉（Μ週)及胎 兒皮膚(14週）。在長期培養期間，未見到纖維母細胞至肌纖維母 細胞之形成。 圖10顯示出在懷孕期12-14週的已分化的胎兒皮膚纖維母細 胞並未去分化成肌纖維母細胞且為alpha· SMA陰性。在這懷孕期 的細胞以纖維母細胞型態與基質表面交互作用。未分化的胎<兒皮 . 膚纖維母細胞及間質幹細胞為alpha- SMA陽性，且分化成^纖維 母細胞並在界面伴隨著纖維化反應與基質材料交互作用。 圖11已分化的胎兒皮膚細胞，尤其在懷孕12_14週時，不分 化成肌纖維母細胞，亦不去分化成其他細胞譜系，例如：當放^ 於成骨性型態發生素中(以生長因子補充來誘發成骨分化的^養 23 201219573 ί羊因造骨細胞；當放置於成脂性型態發生素中(以 放置於分化的培養基口天時，亦不成為脂肪細 胎兒二(<9.MSim補充的培養基中時，未分化的 分化成肌_母細胞。分彳卩使細胞絲質祕體接觸仍不 之誘發受到限制。成其他細胞(即硬骨、肌肉、軟骨……) 【主要元件符號說明】 a自體細胞 J3^ 體 b基質’即具有胎兒細胞之植 24Physiology, 188:211-222, 2001). In the 9-16 weeks of pregnancy, it is better at 12-14 weeks. The fetal skin is fine (4) and the type of myofibroblastic phenotype is shown (Figures 11 and 12). These cell populations that have been developed in a particular dedicated cell bank are often different from the above-mentioned patches. The expression of alpha-, SMA is shown to be indicative of embryonic development in early interstitial cell populations (Clement et al) J of Cell Science, 120: 229-238, 2006) [Simplified Schematic] Figure 1 shows no evidence Some of the examples described herein provide evidence of biocompatibility of an adult keratinocyte skin graft attached to and transported on a "fetal cell coated" matrix of biocompatible horse collagen. In the figure, cells, and b) represent the matrix, ie the implant with fetal cells. #自^田—, 2 shows a continuous thin layer of human keratinocytes isolated from a plastic tissue culture flask. It needs to be supported to transfer to the patient as shown in the (4) layer of the 1 towel or alone. Figure 3 shows an SEM image of a fetal cell on the surface of a biomaterial according to some of the embodiments described herein, showing the biocompatibility interaction of the surface. Figure 4 shows that the fibrous tissue system is formed around the implant material. A separate biomaterial (PLA) is inserted into the hard bone defect. Fibrous tissue formation with separate biomaterials: Solid white dense zoning around the circle and very little to no hard bone growth within the circle were observed. This depicts the fibrillation reaction of soft and hard matrix materials in accordance with some of the embodiments described herein. Figure 5 shows the delivery of fetal cells in pLA that promotes bony formation to hard bone defects. According to some embodiments described herein, there is only bony formation overall and no fibrous tissue formation, depicting anti-fibrotic activity of fetal cells and/or fragments thereof. Figure 6 depicts anti-fibrotic activity in skin-to-muscle soft tissue seen after fetal cell delivery and compatibility with a collagen foam matrix, in accordance with some embodiments described herein. Figure 7 shows the accumulation of anti-fibrotic tissue in severely injured skin following fetal cell delivery and compatibility with a collagen foam matrix, in accordance with some embodiments described herein. We have observed the use of fetal skin cells and collagen matrix/implant fibrosis. Figure 8 depicts the stabilization of fetal cells and/or derivatives by freeze drying/freezing to maintain biological productivity and function in accordance with some embodiments described herein. Biological activity is preserved after cold image drying. Figure 9 shows fetal muscle (periorbital) and fetal skin (14 weeks) showing significant cell type and staining. During long-term culture, no formation of fibroblasts to myofibroblasts was observed. Figure 10 shows that differentiated fetal skin fibroblasts at 12-14 weeks of gestation did not dedifferentiate into myofibroblasts and were alpha·SMA negative. The cells in this gestation phase interact with the surface of the matrix in a fibroblast pattern. Undifferentiated Fetus <Children's Skin. Fibroblasts and mesenchymal stem cells are alpha-SMA positive and differentiate into fibroblasts and interact with matrix material at the interface with fibrosis. Figure 11. Differentiated fetal skin cells, especially at 12-14 weeks of gestation, do not differentiate into myofibroblasts, and do not differentiate into other cell lineages, for example, when placed in osteogenic morphogens (with growth factors) Supplementary to induce osteogenic differentiation of the cultivating bones of the genus 23 201219573 ί sheep due to osteoblasts; when placed in the adipogenic form of the morphogens (to be placed in the differentiation of the culture of the mouth, it does not become fat fetus 2 (<9 When MSim is supplemented in the medium, the undifferentiated differentiation into myo-mother cells. The bifurcation causes the cell silky secret body to contact and is still not restricted. It becomes other cells (ie hard bone, muscle, cartilage...) [mainly Component symbol description] a autologous cell J3^ body b matrix 'that has fetal cells plant 24