TWI476010B - A composition for promoting hair growth - Google Patents
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本發明係有關於一種促進毛髮生長的組合物,特別是關於一種攜鐵蛋白的用途,其係用於製備促進毛髮生長的組合物。The present invention relates to a composition for promoting hair growth, and more particularly to a use of a ferritin for the preparation of a composition for promoting hair growth.
攜鐵蛋白(transferrin,Tf)為一種醣蛋白,含有679個胺基酸,分子量大小約為79 kDa,主要是由肝臟細胞所合成,存在於身體的體液中,包括血漿、膽汁、羊水、腦脊髓液、淋巴液及乳汁等。攜鐵蛋白和乳鐵蛋白一樣能夠結合鐵,所不同的是攜鐵蛋白具有運送鐵離子的作用,主要功能在於將鐵由吸收中心,如十二指腸、巨噬細胞,運送至身體各組織,並於紅血球生成及細胞分裂扮演關鍵角色。Transferrin (Tf) is a glycoprotein containing 679 amino acids with a molecular weight of about 79 kDa. It is mainly synthesized by liver cells and is present in body fluids, including plasma, bile, amniotic fluid, and brain. Spinal fluid, lymph and milk. Ferritin can bind iron like lactoferrin, except that ferritin has the function of transporting iron ions. Its main function is to transport iron from absorption centers, such as duodenum and macrophages, to various tissues of the body. Red blood cell production and cell division play a key role.
此外,攜鐵蛋白亦與先天性免疫(innate immunity)有關,研究發現黏膜中含有攜鐵蛋白,因為其可結合鐵的特性故於黏膜中創造了低鐵離子濃度的環境,讓細菌無法獲得鐵而達到抑制細菌生長作用,另於發炎時攜鐵蛋白濃度亦會降低。In addition, ferritin is also associated with innate immunity. It has been found that the mucosa contains ferritin, which binds to iron and creates an environment of low iron ion concentration in the mucosa, preventing bacteria from obtaining iron. In order to inhibit the growth of bacteria, the concentration of ferritin is also reduced when inflamed.
血漿中攜鐵蛋白的濃度從出生後就很穩定,每公升含量約2-3克,在體內的半衰期可達8天。攜鐵蛋白的濃度和生長有極大的關係,濃度低於每公升0.1克時會增加感染的機率,生長遲緩及貧血。根據研究,攜鐵蛋白和細胞的生長、分化和細胞保護有關。一些重要的結果顯示,攜鐵蛋白會和其他生長因子協同作用,調節細胞生長和分化。不同生長因子的組合,對相同細胞有可能呈現刺激或抑制細胞生長的兩極化反應。The concentration of ferritin in plasma is stable from birth, about 2-3 grams per liter, and has a half-life of up to 8 days in the body. There is a great relationship between the concentration of ferritin and growth. Concentrations below 0.1 gram per liter increase the chance of infection, growth retardation and anemia. According to the study, ferritin is involved in cell growth, differentiation and cell protection. Some important results show that ferritin synergizes with other growth factors to regulate cell growth and differentiation. The combination of different growth factors may have a polarized response to the same cells that stimulate or inhibit cell growth.
在一項已公開的美國專利(US 20020072596)中揭露了一種新穎的攜鐵蛋白聚胜肽,其可用於促進傷口癒合與表皮細胞增生,然而目前並未有任何習知公開技術教示使用攜鐵蛋白來促進毛髮生長。A novel ferritin-rich peptide is disclosed in a published U.S. Patent (US Pat. Protein to promote hair growth.
造成脫髮的因素眾多,例如免疫失調、暴露於毒性化學物質、壓力、燒燙傷或者皮膚疾病,然而最常見的原因係為老化或遺傳所造 成的脫髮。典型促進毛髮生長的方法著重於控制細胞死亡或促進再生,儘管針對生髮劑進行了許多研究發展,傳統生髮劑對於毛髮養育作用,也就是避免毛髮脫落、促進毛髮生長等效果並不足,成效相當有限。There are many factors that cause hair loss, such as immune disorders, exposure to toxic chemicals, stress, burns, or skin diseases. However, the most common cause is aging or genetics. Hair loss. Typical methods for promoting hair growth focus on controlling cell death or promoting regeneration. Although many researches have been carried out on hair growth agents, the effects of traditional hair growth agents on hair growth, that is, avoiding hair loss and promoting hair growth, are insufficient. .
目前皮膚科醫師認可的治療脫髮或禿頭方法包含口服的柔沛(Propecia,Proscar)及外用的落建(Rogaine),此為美國DA目前唯一認可用於治療脫髮的兩種藥物,於女性則僅可適用落建(Rogaine)。柔沛係為唯一核准用於治療男性頭頂與前額脫髮的口服藥,但對女性使用者並無效果且可能具有損害男性性功能的副作用。落建又稱為敏諾西代(Minoxidil),可刺激男性使用者後頭部禿髮部位的生長,亦可促進女性使用者前額區域毛髮生長。然病患有發炎或不正常頭皮的情形時,會造成頭皮對敏諾西代的吸收增加,導致血壓下降、心跳速率增加及體重上升(體液滯留)等副作用。此外,落建中含醇之主要成分會刺激眼睛不適。Currently, dermatologist-approved methods for the treatment of hair loss or bald include oral Propecia (Proscar) and topical Rogaine, which is the only drug currently approved by DA in the United States for the treatment of hair loss. Applicable to Rogaine. Rope is the only oral drug approved for the treatment of male head and forehead hair loss, but has no effect on female users and may have side effects that impair male sexual function. Founded also known as Minoxidil, it stimulates the growth of the baldness of the male head and promotes hair growth in the forehead area of female users. However, when the patient has an irritated or abnormal scalp, the scalp may increase the absorption of senoxide, resulting in side effects such as decreased blood pressure, increased heart rate, and increased body weight (corporeal fluid retention). In addition, the main ingredients of alcohol in the construction will stimulate eye discomfort.
因此,仍需要一種使用毛髮生長促進劑之組合物,以促進毛髮生長且不會產生上述副作用。Therefore, there is still a need for a composition using a hair growth promoter to promote hair growth without causing the above-mentioned side effects.
本發明係提供一種促進一個體毛髮生長的組合物,其特徵是該組合物包含有效量的攜鐵蛋白。The present invention provides a composition for promoting growth of a body hair, characterized in that the composition comprises an effective amount of ferritin.
在本發明一實施例中,攜鐵蛋白的用量為1微克/毫升至200毫克/毫升。In one embodiment of the invention, the amount of ferritin is from 1 microgram/ml to 200 mg/ml.
較佳地,該組合物可透過塗抹、噴灑、蒸泡或噴射的其中之一方式施用在該個體的皮膚區域上或皮膚區域內。Preferably, the composition can be applied to or within the skin area of the individual by one of application, spraying, evaporating or spraying.
較佳地,該組合物之施用可透過將該個體浸泡或泡浴在該組成物中。Preferably, the composition is applied by soaking or bathing the individual in the composition.
較佳地,該組合物之施用可透過局部塗抹在該個體的皮膚。Preferably, the application of the composition is applied topically to the skin of the individual.
較佳地,該組合物可製作成軟膏(ointment)、洗髮精、潤髮乳、塗劑、生髮水、髮膠或慕斯(mousse)的型式。Preferably, the composition can be formulated as an ointment, shampoo, conditioner, lotion, hair tonic, hair gel or mousse.
較佳地,該組合物可進一步包含醫藥上可接受載體。Preferably, the composition may further comprise a pharmaceutically acceptable carrier.
本發明亦提供一種攜鐵蛋白用於製備促一個體毛髮生長之組合物的用途。The invention also provides a use of a ferritin for the preparation of a composition for promoting the growth of a body hair.
本發明之此等及其他態樣將可由下文之較佳具體實例敘述並結合下述圖式時明顯可見,但可在不偏離本揭示內容新穎概念之精神及範圍的情形下對其進行變化及修飾。These and other aspects of the present invention will be apparent from the following description of the preferred embodiments of the invention and Modification.
第1圖顯示攜鐵蛋白可顯著促進毛囊髮幹之生長。(A)為Wistar大鼠毛囊組織之顯微鏡圖像,顯示以濃度100 ug/ml之攜鐵蛋白處理毛囊組織後可顯著促進毛囊髮幹之生長;(B)為圖(A)之髮幹長度測量結果,處理後第5天之髮幹生長長度相較於PBS控制組增加達45%。* p<0.05。Figure 1 shows that ferritin can significantly promote the growth of hair follicles. (A) is a microscopic image of the hair follicle tissue of Wistar rats, which shows that hair follicle tissue treated with a concentration of 100 ug/ml can significantly promote the growth of hair follicles; (B) is the dry length of the hair of Figure (A) As a result of the measurement, the dry growth length on the 5th day after the treatment was increased by 45% compared with the PBS control group. * p<0.05.
第2圖為MTT測量真皮乳頭細胞增生之結果,顯示攜鐵蛋白可顯著提高真皮乳頭組織之增生,且隨著攜鐵蛋白濃度增加效果更為顯著。* p<0.05,** p<0.001。Figure 2 shows the results of MTT measurement of dermal papilla cell proliferation, showing that ferritin can significantly increase the proliferation of dermal papilla tissue, and the effect is more significant with the increase of ferritin concentration. * p<0.05, ** p<0.001.
除非另外定義,本文中所用之所有技術及科學辭彙具皆有熟習本文所屬技藝者所通常明瞭之相同意義。如有衝突,則以本文件,包括其定義為主。All technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art, unless otherwise defined. In case of conflict, this document, including its definition, is the main one.
當用於此處時,本文所使用冠詞「一」或「該」意指該冠詞文法上之受詞為一或一以上(亦即至少為一)。舉例而言,「一元件」代表一元件或多於一元件。As used herein, the articles "a" or "the" are used to mean that the grammatically accepted words are one or more (ie, at least one). For example, "an element" means one element or more than one element.
此處所使用之「毛囊」意指位於皮下的毛髮結構,其為由表皮向下深入真皮之囊狀結構,包含上皮部分與真皮部分。上皮部分含有內根鞘(inner root sheath)與外根鞘(outer root sheath),真皮部分則包含真皮乳頭(dermal papilla)與真皮鞘(dermal sheath)。毛髮的生長歷經生長期(anagen)、退化期(catagen)、終止期(telogen)和脫落期(exogen),並終生不斷重複這一循環。生長期係毛囊的活躍生長期,在毛髮根部的細胞快速分裂且增加至髮幹;退化期係指在生長期末了的短暫轉換階段,示意生長期的終結;終止期為毛髮的靜止階段,在此時毛囊完全呈現終止狀態且形成高爾夫球桿狀,意指髮根完全由角質化細胞組成的球狀擴大(bulbous enlargement)包圍。As used herein, "hair follicle" means a hair structure located under the skin that is a saclike structure that extends from the epidermis down into the dermis, including the epithelial portion and the dermal portion. The epithelial portion contains an inner root sheath and an outer root sheath, and the dermis portion contains a dermal papilla and a dermal sheath. Hair growth progresses through the anagen, catagen, telogen, and exogen, and this cycle is repeated throughout life. During the growth period, the hair follicles are active in the growth phase, and the cells at the root of the hair divide rapidly and increase to the hair shaft; the degeneration period refers to the transient transition phase at the end of the growth phase, indicating the end of the growth phase; the termination phase is the stationary phase of the hair, At this point the hair follicle is completely terminated and forms a golf club shape, meaning that the hair root is completely surrounded by a bulbous enlargement consisting of keratinocytes.
本發明提供一種促進一個體毛髮生長的組合物,包括至少對 該個體之毛囊髮幹的活化與真皮乳頭組織的增生其中之一有效量的攜鐵蛋白。在此「個體」意指人類或具有毛髮的動物,特別指具有毛囊組織的溫血動物及鳥類。The present invention provides a composition for promoting the growth of a body hair, including at least One of the effective amount of ferritin in the activation of hair follicles of the individual and the proliferation of dermal papilla tissue. As used herein, "individual" means human or animal having hair, particularly warm-blooded animals and birds having hair follicle tissue.
「有效量」意指提供促進對於個體之毛髮生長有效果的量,本技術領域之技藝人士可根據本發明揭示之內容,以其知識與慣行實驗的標準方法輕易判斷有效量以及劑量與施用頻率。By "effective amount" is meant an amount that provides an effect that promotes hair growth to an individual, and those skilled in the art can readily determine the effective amount, as well as the dosage and frequency of administration, in accordance with the teachings of the present invention, using their knowledge and standard methods of routine experimentation. .
此處所使用之「攜鐵蛋白」包含牛攜鐵蛋白、人攜鐵蛋白或由其他物種所萃取的攜鐵蛋白、人工合成的攜鐵蛋白、及上述攜鐵蛋白的衍生物。As used herein, "ferroprotein" includes bovine ferritin, human ferritin or a ferritin extracted from other species, a synthetic ferric protein, and a derivative of the above-described ferritin.
本發明發現攜鐵蛋白可促進毛囊髮幹生長,此外亦能顯著促進真皮乳頭細胞之增生,真皮乳頭細胞係毛髮生長之主要組織,因此本發明提供一種有效促進毛髮生長的組合物。The present invention has found that ferritin can promote the growth of hair follicles, and can also significantly promote the proliferation of dermal papilla cells, which are the main tissues for hair growth of dermal papilla cells. Therefore, the present invention provides a composition effective for promoting hair growth.
包含攜鐵蛋白的組合物可透過本領域所熟知的任何方式施於一個體的皮膚,包括但不侷限於局部塗抹、噴灑、蒸泡或噴射包含攜鐵蛋白的組合物至所欲促進毛髮生長之個體的皮膚上或皮膚內。此外,該個體亦可浸泡或泡浴在包含攜鐵蛋白的組合物中。根據本發明,組合物內攜鐵蛋白的含量大約可為1微克/毫升至200毫克/毫升。The composition comprising ferritin can be applied to the skin of a body by any means known in the art including, but not limited to, topical application, spraying, bubbling or spraying a composition comprising ferritin to promote hair growth. On or in the skin of an individual. In addition, the individual may also be soaked or bathed in a composition comprising ferritin. According to the present invention, the ferritin content of the composition may range from about 1 microgram/ml to 200 mg/ml.
包含攜鐵蛋白的組合物製作可以將攜鐵蛋白與其他活性或非活性成分混合成多種形式以便於使用。其他成份可包括非限制實施例如佐藥(adjuvant)、載具(vehicle)或賦形劑(excipient)、香料(perfume)、著色劑(colorant)、穩定劑(stabilizer)或其他非活性成分,或不論是否含有其他非活性成分的任何組合。較佳的非活性成分包括,例如二甲基亞碸(DMSO)、甘油(glycerol)、膠原蛋白(collagen)以及玻尿酸(hyaluronan (HA))。根據不受限制的實施例,組合物可製作成軟膏(ointment)、洗髮精、潤髮乳、塗劑、生髮水、髮膠或慕絲(mousse)的形式。本發明的組成物可包括不同的添加活性成分與平常用於局部配方的補充成分,如生長因子、荷爾蒙與熟諳本項技藝之人士所知的細胞增生因子,其含量並不會影響攜鐵蛋白促進毛髮生長效果。Compositions containing ferritin can be made by mixing ferritin with other active or inactive ingredients in a variety of forms for ease of use. Other ingredients may include, without limitation, an adjuvant such as an adjuvant, a vehicle or excipient, a perfume, a colorant, a stabilizer, or other inactive ingredients, or Whether or not it contains any combination of other inactive ingredients. Preferred inactive ingredients include, for example, dimethyl hydrazine (DMSO), glycerol, collagen, and hyaluronan (HA). According to a non-limiting embodiment, the composition can be formulated in the form of an ointment, shampoo, conditioner, lotion, hair tonic, hair gel or mousse. The compositions of the present invention may comprise different added active ingredients and supplemental ingredients commonly used in topical formulations, such as growth factors, hormones, and cell proliferating factors known to those skilled in the art, which do not affect ferritin levels. Promotes hair growth.
本發明也提供一種攜鐵蛋白用於促進個體毛髮生長的新用途。The invention also provides a novel use of ferritin for promoting hair growth in an individual.
本發明係藉由下列範例進一步說明,此僅提供而用於展現而非限制之目的。由於本揭露,本領域具有通常知識者應可理解所揭露之特定具體實施例,並對該些具體實施例進行諸多修改而獲得相似或類似的結果而仍未脫離本發明之精神與範疇。The invention is further illustrated by the following examples, which are provided for purposes of illustration and not limitation. It will be apparent to those skilled in the art that <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt;
為了研究攜鐵蛋白促進毛髮生長之效果,以腹腔注射7%水化氯醛(chloral hydrate)犧牲六週大的Wistar大鼠(購自台大醫學院動物中心),將其兩頰之皮膚剪下並以1XPBS沖洗,置入含有2.5 ug/ml fungizone的DMEM培養基並保存於4℃中,接續以眼科手術剪刀以及鑷子分離出鼠鬚的完整毛囊。將分離出的鼠鬚毛囊分別以PBS(控制組)或含有100 ug/ml攜鐵蛋白之PBS培養。在培養5天後以顯微鏡觀察測量鼠鬚毛囊髮幹之生長長度,如第1A圖與第1B圖所示,相較於未施以攜鐵蛋白之控制組,以攜鐵蛋白處理毛囊可顯著促進毛囊髮幹之增長達45%,顯示使用有效量的攜鐵蛋白可促進毛髮之生長。In order to study the effect of ferritin on hair growth, a six-week-old Wistar rat (purchased from the Animal Center of National Taiwan University of Medicine) was intraperitoneally injected with 7% chloral hydrate to cut off the skin of the cheeks. The cells were washed with 1X PBS, placed in DMEM medium containing 2.5 ug/ml fungizone and stored at 4 ° C, and the intact hair follicles of the whiskers were separated by ophthalmic surgical scissors and forceps. The isolated murine hair follicles were each cultured in PBS (control group) or PBS containing 100 ug/ml of ferritin. After 5 days of culture, the growth length of the hair follicles was measured by microscopic observation. As shown in Fig. 1A and Fig. 1B, the treatment of hair follicles with ferritin was remarkable compared with the control group without ferricin. It promotes the growth of hair follicles by 45%, indicating that the use of an effective amount of ferritin can promote hair growth.
為進一步測試攜鐵蛋白對於真皮乳頭細胞之影響,將分離出之毛囊根部剪碎,並以第一型膠原蛋白酶(2 mg/ml第一型膠原蛋白酶配置於1% FBS/DMEM溶液中)於37℃、5%二氧化碳培養箱反應2小時,直到外根鞘鬆動後以含有10% FBS之DMEM培養基終止反應,離心並吸除上清液以去除膠原蛋白酶,以含有20% FBS之DMEM培養基重新懸浮沉澱之真皮乳頭細胞進行培養,當觀察到細胞貼附並向外生長延展時,每3天更換新鮮培養液,並於培養7天後進行試驗。To further test the effect of ferritin on dermal papilla cells, the isolated hair follicle roots were shredded and labeled with type 1 collagenase (2 mg/ml type I collagenase in 1% FBS/DMEM solution). The reaction was carried out for 2 hours at 37 ° C in a 5% carbon dioxide incubator until the outer root sheath was loosened. The reaction was stopped in DMEM medium containing 10% FBS, centrifuged and the supernatant was aspirated to remove collagenase, and re-separated with DMEM containing 20% FBS. The dermal papilla cells in suspension were cultured, and when the cells were attached and spread outward, the fresh medium was replaced every 3 days, and the test was carried out 7 days after the culture.
於測試攜鐵蛋白之功效前,將真皮乳頭細胞以2×104 之細胞濃度種於24孔盤,培養16小時後吸棄孔內培養液,以PBS清洗後添加500 μl DMEM培養基(不含FBS)接續培養16小時。16小時後,吸棄孔內培養液並以PBS清洗,接著加入含有不同濃度攜鐵蛋白之DMEM培養基(不含FBS)培養16小時,控制組則以PBS取代攜鐵蛋白。以不同濃度之攜鐵 蛋白處理16小時後,吸棄孔內培養液並以PBS清洗,接著每孔加入200 μl MTT溶液反應2小時,吸棄孔內溶液並加入150 μl DMSO溶解結晶並於540 nm測量吸光值以分析細胞增生。Before testing the effect of ferritin, the dermal papilla cells were seeded in a 24-well plate at a cell concentration of 2×10 4 , and after 16 hours of culture, the culture solution in the well was discarded, and 500 μl of DMEM medium was added after washing with PBS (excluding FBS) continued to culture for 16 hours. After 16 hours, the wells were aspirated and washed with PBS, followed by addition of DMEM containing different concentrations of ferritin (without FBS) for 16 hours, and the control group was replaced with PBS for ferritin. After treatment with different concentrations of ferritin for 16 hours, the well culture solution was aspirated and washed with PBS, then 200 μl of MTT solution was added to each well for 2 hours. The solution in the well was aspirated and 150 μl of DMSO was added to dissolve and crystallize at 540. The absorbance was measured at nm to analyze cell proliferation.
如圖2所示,相較於PBS控制組,分別施以1、10、50、100、200 ug/ml攜鐵蛋白之真皮乳頭細胞可顯著觀察到細胞增生現象,其中施以50 ug/ml攜鐵蛋白之真皮乳頭細胞其增生數目為PBS控制組的1.7倍,攜鐵蛋白濃度增加至100 ug/ml時,細胞增生數目為PBS控制組的2.3倍,而當攜鐵蛋白濃度達200 ug/ml時,真皮乳頭細胞增生數目更可達3.2倍。因此結果顯示施予有效量的攜鐵蛋白能顯著地提升毛囊底部真皮乳頭組織的增生,使毛髮變粗、加速髮幹生長。As shown in Fig. 2, cell proliferation was observed in the dermal papilla cells with 1, 10, 50, 100, 200 ug/ml ferritin, respectively, compared with the PBS control group, with 50 ug/ml applied. The number of proliferating cells of ferritin-producing dermal papilla cells was 1.7 times that of the PBS-controlled group, and when the concentration of ferritin was increased to 100 ug/ml, the number of cell proliferation was 2.3 times that of the PBS-controlled group, and when the concentration of ferritin reached 200 ug. At /ml, the number of dermal papilla cells proliferates up to 3.2 times. Therefore, the results show that administration of an effective amount of ferritin can significantly increase the proliferation of the dermal papilla tissue at the bottom of the hair follicle, making the hair thicker and accelerating dry growth.
因此本發明提供一種有效促進個體毛髮生長的組合物,沒有性別或年齡的侷限。在對經分離之毛囊組織及真皮乳頭細胞施以包含有效量攜鐵蛋白的組合物後,可觀察到毛囊髮幹的顯著增長以及真皮乳頭組織顯著增生,因此該組合物可施用於個體而有效促進其毛髮生長。此外,熟習本項技藝之人士應可根據本發明的內容瞭解,攜鐵蛋白與其他衍生物也可用於本發明的組合物以促進因疾病或自然因素造成脫髮之個體的毛髮生長。The present invention therefore provides a composition effective to promote hair growth in an individual without the limitations of gender or age. After administration of the composition containing the effective amount of ferritin to the isolated hair follicle tissue and the dermal papilla cells, a significant increase in hair follicle dryness and significant proliferation of the dermal papilla tissue can be observed, so that the composition can be applied to the individual effectively Promote its hair growth. Moreover, those skilled in the art will recognize from the teachings of the present invention that ferritin and other derivatives can also be used in the compositions of the present invention to promote hair growth in individuals who develop hair loss due to disease or natural factors.
雖然上述實例的結果係以Wistar品種大鼠顯示攜鐵蛋白於促進毛髮生長的新用途,本發明的範圍不應侷限於此。熟悉本項技藝之人士可根據本發明內容瞭解本發明並應用於任何具有毛囊的溫血動物,包括人類、牛、豬、馬、羊、狗、小鼠、大鼠、兔子及禽類,如雞、鴨、鵝及火雞。Although the results of the above examples show a new use of ferritin to promote hair growth in Wistar breed rats, the scope of the present invention should not be limited thereto. Those skilled in the art will be able to understand the present invention and apply it to any warm-blooded animal having hair follicles, including humans, cows, pigs, horses, sheep, dogs, mice, rats, rabbits, and birds, such as chickens. , ducks, geese and turkeys.
咸信本發明之技藝人士基於本文之敘述,無須進一步之例示即可將本發明應用至其最廣泛之範圍。因此,應了解此處所提供之敘述及申請專利範圍係供例示目的而非以任何方式限制本發明之範疇。The present invention will be applied to its broadest scope without further elaboration, based on the description herein. Therefore, it is to be understood that the claims and claims are not intended to limit the scope of the invention.
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