KR102234860B1 - Composition for prevention and treatment of muscle atrophy comprising lespedeza bicolor extract - Google Patents
Composition for prevention and treatment of muscle atrophy comprising lespedeza bicolor extract Download PDFInfo
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- KR102234860B1 KR102234860B1 KR1020190057501A KR20190057501A KR102234860B1 KR 102234860 B1 KR102234860 B1 KR 102234860B1 KR 1020190057501 A KR1020190057501 A KR 1020190057501A KR 20190057501 A KR20190057501 A KR 20190057501A KR 102234860 B1 KR102234860 B1 KR 102234860B1
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Abstract
본 발명은 싸리(Lespedeza bicolor) 추출물을 포함하는 근위축 예방 및 치료용 조성물에 관한 것이다. The present invention relates to a composition for preventing and treating muscular atrophy comprising an extract of Lespedeza bicolor.
Description
본 발명은 싸리(Lespedeza bicolor) 추출물을 포함하는 근위축 예방 및 치료용 조성물에 관한 것이다. The present invention relates to a composition for preventing and treating muscular atrophy comprising an extract of Lespedeza bicolor.
전 세계적으로 노인 인구가 급속도로 증가하고 있으며, 노화에 따른 다양한 생리적 변화는 널리 알려져 있다. 인슐린 저항성(insulin resistance)은 그 중 하나로 2형 당뇨 발병률 증가와도 연관이 있다. 이러한 인슐린 저항성은 근감소증을 비롯한 다양한 합병증을 유발한다. 인슐린 저항성 증가로 인한 근손상은 인슐린 신호전달 조절의 문제로 인해 에너지 대사가 원활하지 않게 되어 근육조직의 단백질 합성은 억제되고 분해가 증가하면서 진행된다. 이러한 근손상이 지속되면 물리적인 신체활동이 정상적으로 할 수 없게 되고, 이는 더욱 근손상을 심화시키게 된다.Worldwide, the elderly population is increasing rapidly, and various physiological changes according to aging are widely known. Insulin resistance is one of them and has been associated with an increased incidence of
이에 따라 근손상 및 근위축을 완화시킬 수 있는 치료제 개발의 필요성이 지속적으로 증가하고있다. 최근 다양한 기능성 소재의 대한 연구들이 많이 진행되고 있지만 인슐린 저항성 증가에 따른 근손실에 대한 천연물을 이용한 연구들은 부족한 실정이며, 이에 대한 개발이 더욱 요구된다. Accordingly, the need for the development of therapeutic agents that can alleviate muscle damage and muscle atrophy is constantly increasing. Recently, many studies on various functional materials have been conducted, but studies using natural products for muscle loss due to increased insulin resistance are insufficient, and further development is required.
본 발명의 목적은 싸리(Lespedeza bicolor) 추출물을 포함하는 근위축 예방 및 치료용 약학적 조성물을 제공하는 것이다.It is an object of the present invention to provide a pharmaceutical composition for preventing and treating muscular atrophy comprising an extract of Lespedeza bicolor.
본 발명의 다른 목적은 싸리 추출물을 포함하는 근위축 예방 및 개선용 식품 조성물을 제공하는 것이다. Another object of the present invention is to provide a food composition for preventing and improving muscle atrophy, comprising a salicornia extract.
본 명세서에서 사용되는 용어는 본 발명에서의 기능을 고려하면서 가능한 현재 널리 사용되는 일반적인 용어들을 선택하였으나, 이는 당 분야에 종사하는 기술자의 의도 또는 판례, 새로운 기술의 출현 등에 따라 달라질 수 있다. 또한, 특정한 경우는 출원인이 임의로 선정한 용어도 있으며, 이 경우 해당되는 발명의 설명 부분에서 상세히 그 의미를 기재할 것이다. 따라서 본 발명에서 사용되는 용어는 단순한 용어의 명칭이 아닌, 그 용어가 가지는 의미와 본 발명의 전반에 걸친 내용을 토대로 정의되어야 한다.Terms used in the present specification have selected general terms that are currently widely used as possible while taking functions of the present invention into consideration, but this may vary according to the intention or precedent of a technician working in the field, the emergence of new technologies, and the like. In addition, in certain cases, there are terms arbitrarily selected by the applicant, and in this case, the meaning of the terms will be described in detail in the description of the corresponding invention. Therefore, the terms used in the present invention should be defined based on the meaning of the term and the overall contents of the present invention, not a simple name of the term.
다르게 정의되지 않는 한, 기술적이거나 과학적인 용어를 포함해서 여기서 사용되는 모든 용어들은 본 발명이 속하는 기술 분야에서 통상의 지식을 가진 자에 의해 일반적으로 이해되는 것과 동일한 의미를 가지고 있다. 일반적으로 사용되는 사전에 정의되어 있는 것과 같은 용어들은 관련 기술의 문맥상 가지는 의미와 일치하는 의미를 가지는 것으로 해석되어야 하며, 본 출원에서 명백하게 정의하지 않는 한, 이상적이거나 과도하게 형식적인 의미로 해석되지 않는다.Unless otherwise defined, all terms used herein including technical or scientific terms have the same meaning as commonly understood by one of ordinary skill in the art to which the present invention belongs. Terms such as those defined in a commonly used dictionary should be interpreted as having a meaning consistent with the meaning in the context of the related technology, and should not be interpreted as an ideal or excessively formal meaning unless explicitly defined in the present application. Does not.
상기와 같은 목적을 달성하기 위한 본 발명의 일 측면은 싸리(Lespedeza bicolor) 추출물을 포함하는 근위축 예방 및 치료용 약학적 조성물에 관한 것이다.One aspect of the present invention for achieving the above object relates to a pharmaceutical composition for preventing and treating muscular atrophy comprising an extract of Lespedeza bicolor.
본 발명에서 “싸리(Lespedeza bicolor)”는 콩 과에 속하는 다년생 낙엽 관목으로서 잔가지는 능선이 있고 짙은 갈색이며, 목재는 연한 녹색을 띠고 골 속은 흰 식물이다. 7~8월에 채취하여 햇볕에 말리거나 신선한 것을 주로 이용한다. 싸리의 종류는 싸리, 참싸리, 물싸리, 조록싸리, 잡싸리, 괭이싸리, 꽃참싸리, 왕좀싸리, 좀싸리, 풀싸리, 해변싸리, 고양싸리, 지리산싸리, 진도싸리 등 종류가 다양하며, 모두 약으로 사용될 수 있다. 싸리 나무는 두통, 폐열로 인한 기침, 심장병, 백일해, 성병, 눈 출혈 등을 치료하며, 싸리 나무의 뿌리는 마비, 타박상, 류머티스성 관절염, 콩팥 질환 등에 효과가 있는 것으로 알려져 있다. 나아가 싸리 나무 잎은 두통에, 싸리 나무 꽃은 주근깨나 기미 등 피부 개선에 효능이 있는 것으로 알려져 있다. In the present invention, " Lespedeza bicolor " is a perennial deciduous shrub belonging to the legume family, and its fine branches have ridges and are dark brown, the wood has a light green color, and the inside is a white plant. It is collected from July to August and dried in the sun or fresh. There are a variety of types of sari, ssari, ssari, ssari, japssari, hoe ssari, flower ssari, king ssari, ssari, grass ssari, beach ssari, Goyang ssari, Jirisan ssari, Jindo ssari, etc. Can be used as a medicine. Ssari tree is known to treat headache, cough caused by lung heat, heart disease, whooping cough, sexually transmitted disease, eye bleeding, etc., and its roots are known to be effective in paralysis, bruises, rheumatoid arthritis, and kidney disease. Furthermore, it is known that the leaves of the serrata are effective for headaches and the flowers of the serrata are effective in improving skin such as freckles and spots.
한편 싸리의 다양한 치료효과에 관하여 개시한 문헌 중에 싸리 추출물이 근위축 완화에 효과가 있음을 교시 또는 개시한 문헌은 없다. 이에 본 발명자들은 본 발명에서 싸리 추출물이 근위축 및 근손상 완화에 유용한 효과가 있음을 규명하였으며, 싸리 추출물을 포함하는 약학적 조성물을 제공한다. On the other hand, among the literatures disclosed on the various therapeutic effects of Ssari, there is no document that teaches or discloses that Ssari extract is effective in relieving muscle atrophy. Accordingly, the inventors of the present invention have found that the extract is effective in relieving muscle atrophy and muscle damage in the present invention, and provides a pharmaceutical composition containing the extract.
본 발명에서, “근위축(atrophy)”는 근육 조직의 손실, 근육 자체의 병 또는 근육을 지배하는 신경의 손상으로 인한 모든 질환을 포함한다. 상기 근위축은 muscular amyotrophy), 근위축성 측삭경화증(amyotrophic lateral sclerosis, ALS), 척수구 근위축(spinobular muscular atrophy) 등을 포함할 수 있으며, 이에 제한되는 것은 아니다. In the present invention, "atrophy" includes all diseases caused by loss of muscle tissue, disease of the muscle itself, or damage to nerves that control the muscle. The muscular atrophy may include, but is not limited to, muscular amyotrophy, amyotrophic lateral sclerosis (ALS), and spinobular muscular atrophy.
구체적으로, 상기 근위축은 인슐린 저항성 증가에 의한 근위축증(amyotrophy)일 수 있다.Specifically, the muscle atrophy may be amyotrophy caused by an increase in insulin resistance.
본 발명에서, “인슐린 저항성 증가에 의한 근위축증(amyotrophy)”은 동통성 비대칭성 또는 대칭성 다발신경병증의 일종으로, 대퇴부위의 통증과 함께 근력의 약화, 근소실 및 근육 위축이 야기되는 질병을 말한다. 정확한 병리학적 소견이 밝혀진 것은 아니나, 인슐린 저항성에 의해 발생될 수 있다.In the present invention, "amyotrophy due to increased insulin resistance" is a type of painful asymmetry or symmetric polyneuropathy, and refers to a disease in which muscle weakness, muscle loss, and muscle atrophy are caused along with pain in the femur. . Although the exact pathological findings have not been established, it can be caused by insulin resistance.
더욱 구체적으로, 인슐린 저항성 증가에 의한 근위축증은 고혈당으로 인한 산화적 스트레스 및 과도한 최종 당산화물 생성으로 인해 각종 염증성 사이토카인, 성장인자 및 혈관조절인자들의 변화에 의해 유발될 수 있다. More specifically, muscular dystrophy caused by increased insulin resistance may be caused by changes in various inflammatory cytokines, growth factors, and vascular regulators due to oxidative stress due to hyperglycemia and excessive production of final sugar oxides.
본 발명에서, “인슐린 저항성”은 인슐린 분비가 정상적으로 이루어지더라도 양이 충분하지 않거나, 생성된 인슐린이 신체에 효과적으로 작용하지 않아 발생하는 것을 말한다. 인슐린 저항성은 인슐린 수용체 자체의 발현 감소 또는 인슐린 수용체의 하위 신호전달의 이상이 원인으로 알려져 있다. 인슐린 수용체 하위 신호 중에서도 IRS-1의 티로신 인산화 혹은 IRS-1 단백질 양 자체에 의해 조절되는 IRS-1(insulin receptor substrate-1)-관련(associated) PI3K 활성의 저해가 주된 원인으로 알려져 있다. In the present invention, "insulin resistance" refers to that the amount of insulin is not sufficient even if the secretion of insulin is performed normally, or that the generated insulin does not act effectively on the body. Insulin resistance is known to be caused by a decrease in the expression of the insulin receptor itself or an abnormality in sub-signaling of the insulin receptor. Among the subsignals of the insulin receptor, inhibition of IRS-1 (insulin receptor substrate-1)-associated PI3K activity, which is regulated by IRS-1's tyrosine phosphorylation or the amount of IRS-1 protein itself, is known to be the main cause.
본 발명에서는 인슐린 저항성 증가의 대표적인 모델로 마일드 (mild)한 2형 당뇨 질환 모델 쥐에 싸리 추출물을 투여한 결과 싸리 추출물을 투여하지 않은 당뇨병 질환 모델 쥐에 비해 근육의 무게가 증가된 것을 확인하였으며(도 1), 광학 현미경으로 근육 조직 절편을 관찰한 결과 근섬유의 크기가 회복됨을 확인하였는 바(도 2), 본 발명의 조성물이 인슐린 저항성 증가에 의한 근소실 및 근위축을 완화시키는 효과가 있음을 확인하였다. In the present invention, as a result of administering Ssari extract to
더욱 구체적으로 상기 싸리 추출물은 인슐린 신호전달 관련 인자들을 활성화시킴으로써 인슐린 저항성 증가에 의한 의한 근소실 및 근위축을 완화시킬 수 있다. More specifically, the Ssari extract may alleviate muscle loss and muscle atrophy caused by an increase in insulin resistance by activating factors related to insulin signaling.
본 발명의 일 실시예에서는 인슐린 저항성이 증가한 쥐에 싸리 추출물을 처리한 후 인슐린 신호 전달 관련 인자의 발현량을 측정하였으며, 싸리 추출물을 처리하지 않은 쥐에 비해 IRβ(insulin receptor β, IRS-1(insulin receptor substrate 1), pAkt/Akt(phosphorylation of Akt/Akt) 및 GLUT4(glucose transporter type 4)의 발현량이 현저히 증가했음을 확인하였다(도 3).In one embodiment of the present invention, the expression level of insulin signal transduction-related factors was measured after treatment of a mouse with increased insulin resistance in a mouse with increased insulin resistance, and IRβ (insulin receptor β, IRS-1 ( It was confirmed that the expression levels of insulin receptor substrate 1), pAkt/Akt (phosphorylation of Akt/Akt) and GLUT4 (glucose transporter type 4) were significantly increased (FIG. 3).
또한, 구체적으로 본 발명의 싸리 추출물은 근육에서 에너지 대사 관련 인자의 발현을 증가시킬 수 있다. 더욱 구체적으로, 상기 에너지 대사 관련 인자 중 pAPMK/AMPK(phosphorylation of APMK/APMK), SIRT1(sirtuin1), SIRT3(sirtuin3), SIRT4(sirtuin4) 및 PGC1-α(phosphatidylglycerol phospholipase1- α) 중 어느 하나 이상의 발현을 증가시킬 수 있다. In addition, specifically, the extract of the present invention can increase the expression of factors related to energy metabolism in muscles. More specifically, expression of any one or more of pAPMK/AMPK (phosphorylation of APMK/APMK), SIRT1 (sirtuin1), SIRT3 (sirtuin3), SIRT4 (sirtuin4), and PGC1-α (phosphatidylglycerol phospholipase1-α) among the energy metabolism-related factors Can increase.
제2형 당뇨의 경우 인슐린 신호 전달에 관여하는 IRS의 티로신 인산화가 증가하고 pAkt/Akt의 발현이 줄어들며, 그 결과 GLUT4의 발현이 감소하면서 조직 내 포도당 이용이 줄어들게 된다. 이는 SIRT1, SIRT3를 비롯한 미토콘드리아 합성관련인자를 감소시키고 각 조직의 에너지 대사를 원활하지 않게 하면서 조직 내 단백질 손상을 증가시켜 인슐린저항성 증가에 의한 근손상을 초래한다. 본 발명의 조성물은 상기 에너지 대사 관련 인자를 활성화시킴으로써 각 조직의 에너지 대사를 원활하게 하여 인슐린저항성 증가에 의한 근손상을 완화시킬 수 있다. In the case of
본 발명의 일 실시예에서 인슐린 저항성이 증가한 쥐에 싸리 추출물을 처리한 후 근육에서의 에너지 대사 관련 인자의 발현량을 측정하였으며 싸리 추출물을 처리하지 않은 쥐에 비해 pAPMK/AMPK, SIRT1, SIRT3, SIRT4 및 PGC1-α의 발현량이 증가함을 확인하였다(도 4).In one embodiment of the present invention, the expression level of energy metabolism-related factors in the muscles was measured after treatment with Ssari extract in mice with increased insulin resistance, and pAPMK/AMPK, SIRT1, SIRT3, SIRT4 compared to mice not treated with Ssari extract. And it was confirmed that the expression level of PGC1-α increased (Fig. 4).
또한 구체적으로 본 발명의 싸리 추출물은 근육의 염증반응 관련 인자의 발현을 감소시킬 수 있다. 더욱 구체적으로, 상기 염증반응 관련 인자인 MCP-1(monocyte chemoattractant protein 1)의 발현을 감소시킬 수 있다. In addition, specifically, the extract of the present invention can reduce the expression of factors related to the inflammatory response of the muscle. More specifically, it is possible to reduce the expression of the inflammatory response-related factor MCP-1 (monocyte chemoattractant protein 1).
본 발명의 일 실시예에서는 인슐린 저항성이 증가한 쥐에 싸리 추출물을 처리한 후 근육에서의 염증반응 관련 인자의 발현량을 측정한 결과, 싸리 추출물을 처리하지 않은 쥐에 비해 MCP-1의 발현량이 감소됨을 확인하였다(도 5).In one embodiment of the present invention, as a result of measuring the expression level of inflammatory response-related factors in the muscles after treatment with Ssari extract in mice with increased insulin resistance, the expression level of MCP-1 is reduced compared to mice not treated with Ssari extract. Was confirmed (Fig. 5).
상기 싸리 추출물은 단독 또는 혼합 용매로 추출하여 수득된 것일 수 있으며, 당해 공지된 임의의 용매를 사용할 수 있다.The Ssari extract may be obtained by extraction alone or with a mixed solvent, and any known solvent may be used.
구체적으로, 상기 싸리 추출물은 물, C1 내지 C4의 무수 또는 저급 알코올, 상기 물과 저급 알코올의 혼합 용매, 아세톤, 1,3-부틸렌글리콜, 에틸아세테이트, 클로로포름으로 이루어진 군에서 선택되는 1 종 이상의 단독 또는 혼합 용매로 추출하여 수득한 것일 수 있으나, 이에 제한되는 것은 아니다. Specifically, the Ssari extract is 1 selected from the group consisting of water,
또한, “추출물”은 상기 싸리 추출물 뿐 아니라 상기 추출액의 희석액이나 농축액, 상기 추출액을 건조하여 얻어지는 건조물, 상기 추출액의 조제물이나 정제물, 또는 이들의 혼합물 등, 추출액 자체 및 추출액을 이용하여 형성 가능한 모든 제형의 추출물을 포함한다.In addition, the "extract" can be formed using the extract itself and the extract, such as a dilution or concentrate of the extract, a dried product obtained by drying the extract, a preparation or purified product of the extract, or a mixture thereof, as well as the ssari extract. Contains extracts of all formulations.
본 발명의 상기 싸리 추출물은, 상기 싸리의 천연, 잡종 또는 변종 식물로부터 추출될 수 있고, 식물 조직의 배양물로부터도 추출이 가능하다.The Ssari extract of the present invention may be extracted from natural, hybrid or variant plants of Ssari, and may be extracted from cultures of plant tissues.
또한, 상기 싸리 추출물은 그 농도를 적절한 범위로 조절하여 사용할 수 있으며, 구체적으로 상기 싸리 추출물의 농도는 약 100mg/kg 내지 250mg/kg일 수 있다. 보다 더 구체적으로, 상기 싸리 추출물의 농도는 약 100mg/kg 또는 250mg/kg일 수 있으나, 이에 제한되는 것은 아니며, 필요에 따라 적절히 변경하여 사용할 수 있다. In addition, the sari extract may be used by adjusting its concentration to an appropriate range, and specifically, the concentration of the sari extract may be about 100 mg/kg to 250 mg/kg. More specifically, the concentration of the salicornia extract may be about 100mg/kg or 250mg/kg, but is not limited thereto, and may be appropriately changed and used as necessary.
본 발명의 추출물을 추출하는 방법은 특별히 제한되지 아니하며, 당해 기술 분야에서 통상적으로 사용하는 방법에 따라 추출할 수 있다. 상기 추출 방법의 비제한적인 예로는, 열수 추출법, 초음파 추출법, 여과법, 환류 추출법 등을 들 수 있으며, 이들은 단독으로 수행되거나 2 종 이상의 방법을 병용하여 수행될 수 있다. The method of extracting the extract of the present invention is not particularly limited, and may be extracted according to a method commonly used in the art. Non-limiting examples of the extraction method include a hot water extraction method, an ultrasonic extraction method, a filtration method, a reflux extraction method, and the like, and these may be performed alone or in combination of two or more methods.
본 발명에서 열수 추출 또는 냉침 추출한 추출물은 부유하는 고체 입자를 제거하기 위해 여과, 예를 들어 나일론 등을 이용해 입자를 걸러내거나 냉동여과법 등을 이용해 여과한 후, 그대로 사용하거나 이를 동결건조, 열풍건조, 분무건조 등을 이용해 건조시켜 사용할 수 있다.In the present invention, the extract extracted with hot water or cold needle is filtered to remove suspended solid particles, for example, nylon, etc. to filter the particles, or after filtration using a freeze filtration method, etc., or use as such, or freeze drying, hot air drying, It can be used after drying by spray drying or the like.
본 발명의 약학적 조성물은 투여를 위하여, 상기 본 발명의 싸리 추출물 외에 약학적으로 허용 가능한 담체, 부형제 또는 희석제를 포함할 수 있다. 상기 담체, 부형제 및 희석제로는 락토즈, 덱스트로즈, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아 고무, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로즈, 메틸 셀룰로즈, 미정질 셀룰로스, 폴리비닐 피롤리돈, 물, 메틸히드록시벤조에이트, 프로필히드록시벤조에이트, 탈크, 마그네슘 스테아레이트 및 광물유를 들 수 있다.For administration, the pharmaceutical composition of the present invention may contain a pharmaceutically acceptable carrier, excipient, or diluent in addition to the extract of the present invention. The carrier, excipient and diluent include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline Cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oils.
또한, 본 발명의 약학적 조성물은 어떠한 제형으로도 적용가능하며, 경구용 제형으로는 산제, 과립제, 정제, 캡슐제, 현탁액, 에멀젼, 시럽, 에어로졸 등일 수 있다. 비경구용 제형으로는 주사용, 도포용, 에어로졸 등의 스프레이 형일 수 있다. In addition, the pharmaceutical composition of the present invention can be applied in any dosage form, and oral dosage forms may be powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, and the like. The parenteral formulation may be a spray type such as injection, application, or aerosol.
상기 경구 투여를 위한 고형제제에는 정제, 환제, 산제, 과립제, 캡슐제 등이 포함되며, 이러한 고형제제는 상기 싸리 추출물에 적어도 하나 이상의 부형제, 예컨대, 전분, 칼슘, 카보네이트, 수크로오스, 락토오스, 또는 젤라틴 등을 혼합하여 조제할 수 있다. 상기 부형제 이외에 마그네슘 스티레이트, 탈크 같은 윤활제가 사용될 수도 있다.The solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and these solid preparations include at least one excipient, such as starch, calcium, carbonate, sucrose, lactose, or gelatin, in the salicornia extract. It can be prepared by mixing and the like. In addition to the above excipients, lubricants such as magnesium stearate and talc may be used.
상기 경구 투여를 위한 액상 제제로는 현탁제, 내용액제, 유제, 시럽제 등이 사용될 수 있으며, 단순희석제인 물, 리퀴드, 파라핀 외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다.Liquid preparations for oral administration may include suspensions, liquid solutions, emulsions, syrups, etc., and include simple diluents such as water, liquid, and paraffin, as well as various excipients such as wetting agents, sweetening agents, fragrances, and preservatives. I can.
비경구 투여를 위한 제제에는 멸균된 수용액, 비수성 용제, 현탁제, 유제, 동결건조 제제, 좌제가 포함된다. 비수성 용제, 현탁제로는 프로필렌글리콜(propylene glycol), 폴리에틸렌 글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르 등이 사용될 수 있다.Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized formulations, and suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used.
본 발명의 다른 측면은 싸리 추출물을 포함하는 약학적 조성물을 치료를 필요로 하는 개체에 약학적으로 유효한 양으로 투여하는 단계를 포함하는, 근위축증 치료방법에 관한 것이다. 구체적으로, 상기 근위축증은 인슐린저항성 증가에 의한 근위축증일 수 있다. Another aspect of the present invention relates to a method for treating muscular dystrophy, comprising administering a pharmaceutical composition comprising a salicornia extract to an individual in need thereof in a pharmaceutically effective amount. Specifically, the muscular atrophy may be a muscular atrophy caused by an increase in insulin resistance.
“싸리 추출물” 및 “근위축증” 등은 상기 설명한 바와 같다. "Ssari extract" and "muscular dystrophy" and the like are as described above.
본 발명에서 "약학적으로 유효한 양"은 의학적 치료에 적용 가능한 합리적인 수혜/위험 비율로 질환을 치료하기에 충분한 양을 의미하며, 유효 용량 수준은 환자의 성병, 연령, 질병의 종류, 중증도, 약물의 활성, 약물에 대한 민감도, 투여 시간, 투여 경로 및 배출 비율, 치료기간, 동시 사용되는 약물을 포함한 요소 및 기타 의학 분야에 잘 알려진 요소에 따라 결정될 수 있다. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is the patient's sexually transmitted disease, age, type of disease, severity, drug Activity, sensitivity to drugs, time of administration, route of administration and rate of excretion, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field.
본 발명의 약학적 조성물은 개별 치료제로 투여하거나 다른 치료제와 병용하여 투여될 수 있고, 종래의 치료제와 순차적으로 또는 동시에 투여될 수 있다. 또한 본 발명의 약학적 조성물은 단일 또는 다중 투여될 수 있다. 상기 요소를 모두 고려하여 부작용 없이 최소한의 양으로 최대 효과를 얻을 수 있는 양을 투여하는 것이 중요하며, 당업자에 의해 용이하게 결정될 수 있다. The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or administered in combination with other therapeutic agents, and may be administered sequentially or simultaneously with a conventional therapeutic agent. In addition, the pharmaceutical composition of the present invention may be administered single or multiple. It is important to administer an amount capable of obtaining the maximum effect in a minimum amount without side effects in consideration of all of the above factors, and can be easily determined by a person skilled in the art.
본 발명의 용어 "개체"는 본 발명에 따른 약학적 조성물의 투여에 의해 증상이 호전될 수 있는 인슐린 저항성 증가에 의한 근위축 질환을 가진 동물 또는 인간을 포함한다. 본 발명에 따른 치료용 조성물을 개체에게 투여함으로써, 근위축을 효과적으로 예방 및 치료할 수 있다. The term "subject" of the present invention includes an animal or human having a muscular dystrophy disease caused by an increase in insulin resistance whose symptoms can be improved by administration of the pharmaceutical composition according to the present invention. By administering the therapeutic composition according to the present invention to an individual, it is possible to effectively prevent and treat muscular atrophy.
본 발명의 용어 "투여"는 어떠한 적절한 방법으로 인간 또는 동물에게 소정의 물질을 도입하는 것을 의미하며, 본 발명에 따른 치료용 조성물의 투여 경로는 목적 조직에 도달할 수 있는 한 어떠한 일반적인 경로를 통하여 경구 또는 비경구 투여될 수 있다. 또한, 본 발명에 따른 치료용 조성물은 유효성분이 표적 세포로 이동할 수 있는 임의의 장치에 의해 투여될 수 있다. The term "administration" of the present invention means introducing a predetermined substance to humans or animals by any suitable method, and the route of administration of the therapeutic composition according to the present invention is through any general route as long as it can reach the target tissue. It can be administered orally or parenterally. In addition, the therapeutic composition according to the present invention may be administered by any device capable of moving the active ingredient to target cells.
본 발명에 따른 약학적 조성물의 바람직한 투여량은 환자의 상태 및 체중, 질병의 정도, 약물 형태, 투여 경로 및 기간에 따라 다르지만, 당업자에 의해 적절하게 선택될 수 있다. The preferred dosage of the pharmaceutical composition according to the present invention varies depending on the condition and weight of the patient, the degree of disease, the form of the drug, the route and duration of administration, but may be appropriately selected by those skilled in the art.
본 발명의 또 다른 측면은 싸리 추출물을 포함하는, 근위축 예방 및 개선용 식품 조성물에 관한 것이다. 구체적으로, 상기 싸리 추출물은 인슐린 저항성 증가에 의한 근위축을 개선시킬 수 있다. Another aspect of the present invention relates to a food composition for preventing and improving muscle atrophy, comprising a ssari extract. Specifically, the salicornia extract may improve muscle atrophy due to an increase in insulin resistance.
“싸리 추출물” 및 “인슐린 저항성 증가에 의한 근위축” 등은 상기 설명한 바와 동일하다."Ssari extract" and "muscular atrophy due to increased insulin resistance" are the same as described above.
본 발명의 싸리 추출물을 첨가할 수 있는 식품은 육류, 빵, 소시지, 초콜릿류, 스넥류, 캔디류, 과자류, 라면, 피자, 기타 면류, 껌류, 아이스크림류를 포함한 낙농제품, 각종 스프, 음료수, 차, 비타민 복합제, 건강기능성 보조식품 등이 있다. Foods to which the Ssari extract of the present invention can be added include meat, bread, sausage, chocolate, snacks, candy, confectionery, ramen, pizza, other noodles, gums, dairy products including ice cream, various soups, beverages, tea, There are vitamin complexes and health functional supplements.
상기 식품의 종류는 구체적으로 건강기능식품일 수 있다. 상기 건강기능 식품은 여러 가지 영양제, 비타민, 광물(전해질), 합성 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 증진제(치즈, 초콜릿 등), 펙트산 및 그의 염, 유기산, 보호성 콜로이드 점증제, pH 조절제, 안정화제, 보존제, 글리세린, 알코올, 탄산음료에 사용되는 탄산화제 등을 함유할 수 있다. 이러한 성분은 단독으로 또는 조합으로 사용될 수 있으며, 이러한 첨가제의 비율은 조성물 전체 중량당 0.001 내지 50 중량부의 범위에서 선택되는 것이 일반적이다.The type of food may specifically be a health functional food. The health functional foods include various nutrients, vitamins, minerals (electrolytes), synthetic flavors and flavoring agents such as natural flavors, coloring agents and enhancers (cheese, chocolate, etc.), pectic acid and its salts, organic acids, protective colloidal growth. It may contain an agent, a pH adjuster, a stabilizer, a preservative, glycerin, alcohol, a carbonating agent used in carbonated beverages, and the like. These components may be used alone or in combination, and the proportion of these additives is generally selected in the range of 0.001 to 50 parts by weight per total weight of the composition.
상기 건강기능식품은 식품의 생체 조절 기능을 강조한 식품으로 물리적, 생화학적, 생물공학적인 방법을 이용하여 특정 목적에 작용 및 발현하도록 부가가치를 부여한 식품이다. 이러한 건강기능 식품의 성분은 생체 방어와 신체 리듬의 조절, 질환의 방지 및 회복에 관계하는 신체 조절 기능을 생체에 대하여 충분히 발휘하도록 설계하여 가공하게 되며, 식품으로 허용 가능한 식품 보조 첨가제, 감미료 또는 기능성 원료를 함유할 수 있다. The health functional food is a food that emphasizes the biological regulation function of food, and is a food that has added value to act and express it for a specific purpose using physical, biochemical, and bioengineering methods. The ingredients of these health functional foods are designed and processed to sufficiently exert the body's control functions related to the body defense, regulation of body rhythm, prevention and recovery of diseases to the living body, and food supplementary additives, sweeteners, or functional foods that are acceptable as foods. It may contain raw materials.
상기 건강기능식품은 정제, 과립, 분말, 캅셀, 액상의 용액 및 환으로 이루어진 군에서 선택된 어느 하나의 제형일 수 있으나, 이에 제한되지 않는다. 구체적으로 상기 정제 형태의 건강기능식품은 싸리 추출물, 부형제, 결합제, 붕해제 및 다른 첨가제 혼합물을 통상의 방법으로 과립화한 다음, 활택제 등을 넣어 압축 성형하거나, 상기 혼합물을 직접 압축 성형하여 제조할 수 있다. 또한, 상기 정제 형태의 건강기능식품은 필요에 따라 고미제 등을 함유할 수 있으며, 필요에 따라 적당한 제피제로 제피할 수도 있다. The health functional food may be any one formulation selected from the group consisting of tablets, granules, powders, capsules, liquid solutions, and pills, but is not limited thereto. Specifically, the health functional food in the form of a tablet is prepared by granulating a mixture of extract, excipient, binder, disintegrant, and other additives in a conventional manner, and then compression molding with a lubricant, or compression molding the mixture directly. can do. In addition, the health functional food in the form of a tablet may contain a bitter agent or the like if necessary, and may be coated with a suitable skin-forming agent if necessary.
상기 캅셀 형태의 건강기능식품 중 경질캅셀제는 통상의 경질캅셀에 싸리 추출물 및 부형제 등의 첨가제와의 혼합물 또는 그의 입상물 또는 제피한 입상물을 충진하여 제조할 수 있다. 연질캅셀제는 싸리 추출물 및 부형제 등의 첨가제와의 혼합물을 젤라틴 등 캅셀기제에 충진하여 제조할 수 있다. 상기 연질캅셀제는 필요에 따라 글리세린 또는 솔비톨 등의 가소제, 착색제, 보존제 등을 함유할 수 있다. Among the health functional foods in the form of capsules, the hard capsules may be prepared by filling a conventional hard capsule with a mixture of additives such as ssari extract and excipients, or granular or skinned granules thereof. Soft capsules can be prepared by filling a mixture of additives such as salicornia extract and excipients into a capsule base such as gelatin. The soft capsule may contain a plasticizer such as glycerin or sorbitol, a colorant, a preservative, and the like, if necessary.
상기 환 형태의 건강기능식품은 싸리 추출물, 부형제, 결합제, 붕해제 등의 혼합물을 적당한 방법으로 성형하여 조제할 수 있으며, 필요에 따라 백당이나 다른 적당한 제피제로 제피를, 또는 전분, 탈크 또는 적당한 물질로 환의를 입힐 수도 있다. The ring-shaped health functional food may be prepared by molding a mixture of salicornia extract, excipients, binders, disintegrants, etc. by suitable methods, and if necessary, the skins may be coated with a white sugar or other suitable coating agents, or starch, talc, or a suitable substance. It can also be rejoicing.
상기 과립 형태의 건강기능식품은 싸리 추출물, 부형제, 결합제, 붕해제 등의 혼합물을 적당한 방법으로 입상으로 제조할 수 있으며, 필요에 따라 착향제, 고미제 등을 함유할 수 있다. The health functional food in the form of granules can be prepared in a granular form by a suitable method, such as a mixture of the extract, excipients, binders, disintegrants, etc., and may contain flavoring agents, bittering agents, and the like, if necessary.
상기 부형제, 결합제, 붕해제, 활택제, 고미제, 착향제 등에 대한 용어 정의는 당업계에 공지된 것으로 그 기능 등의 동일 내지 유사한 것들을 포함할 수 있다. The definition of terms for the excipients, binders, disintegrants, lubricants, bitters, flavoring agents, and the like are known in the art, and may include the same or similar functions such as the function thereof.
또한, 상기 식품의 종류는 식품 첨가제일 수 있으며, 상기 식품 첨가제는 식품의 제조, 가공, 또는 보존을 위해 식품에 첨가, 혼합, 침윤 기타의 방법에 의해 사용되는 물질을 의미한다. 상기 식품 첨가제는 천연물과 합성품이 있으며, 기능과 용도에 따라 분류할 수 있다. 현재 한국에 식품첨가물로 허가되어 있는 품목은 화학적 합성품 370여종, 천연첨가물 50여종이며, 주로 용도에 따라 보존료, 살균제, 산화방지제, 착색료, 발색제, 표백제, 조미료, 감미료, 착향료, 팽창제, 강화제, 개량제, 유화제, 증점제(호료) 및 안정제, 피막제, 껌 기초제, 소포제, 용제, 이형제, 방충제, 품질개량제와 기타 식품제조용 첨가제 등으로 분류되어 쓰이고 있다.In addition, the type of food may be a food additive, and the food additive refers to a substance used by a method such as adding, mixing, infiltrating or adding to food for the preparation, processing, or preservation of food. The food additives include natural products and synthetic products, and can be classified according to functions and uses. Currently, 370 kinds of chemical synthetic products and 50 kinds of natural additives are approved as food additives in Korea. Depending on the purpose, preservatives, fungicides, antioxidants, coloring agents, coloring agents, bleaching agents, seasonings, sweeteners, flavoring agents, expanding agents, reinforcing agents, and improving agents , Emulsifiers, thickeners and stabilizers, coating agents, gum base agents, defoaming agents, solvents, release agents, insect repellents, quality improvers and other additives for food manufacturing.
상기 식품첨가제의 형태는 분말, 과립, 정제, 캡슐 또는 액상 형태를 포함할 수 있으며 구체적으로는 캡슐의 형태일 수 있으나, 상기 형태에 제한되는 것은 아니다.The form of the food additive may include a powder, granule, tablet, capsule or liquid form, and specifically may be a capsule form, but is not limited to the form.
상기 싸리 추출물은 그 농도를 적절한 범위로 조절하여 사용할 수 있으며, 구체적으로 상기 싸리 추출물의 농도는 100mg/kg 내지 250mg/kg일 수 있다. 보다 구체적으로, 상기 싸리 추출물의 농도는 100mg/kg 또는 250mg/kg이나, 이에 제한되는 것은 아니며, 필요에 따라 적절히 변경하여 사용할 수 있다. The Ssari extract may be used by adjusting its concentration to an appropriate range, and specifically, the concentration of Ssari extract may be 100mg/kg to 250mg/kg. More specifically, the concentration of the salicornia extract is 100mg/kg or 250mg/kg, but is not limited thereto, and may be appropriately changed and used as necessary.
본 발명의 싸리 추출물을 식품용 조성물로 사용할 경우, 상기 싸리 추출물을 그대로 첨가하거나 다른 식품 또는 식품 성분과 함께 사용하고, 통상적인 방법에 따라 적절하게 사용할 수 있다. 상기 싸리 추출물의 혼합량은 그의 사용 목적(예방, 건강 또는 개선, 치료적 처치)에 따라 적합하게 결정될 수 있다. When using the extract of the present invention as a food composition, the extract may be added as it is or used with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixing amount of the salicornia extract may be appropriately determined according to the purpose of use (prevention, health or improvement, therapeutic treatment).
본 발명의 조성물은 인슐린 저항성에 의해 유발된 근위축을 효과적으로 완화시킬 수 있으며, 나아가 천연물 기반 물질로서 인체 안정성이 우수한 바, 인슐린 저항성 증가로 인해 유발된 근위축 및 근손상을 치료 및 예방시키는 용도로 널리 활용될 수 있다. The composition of the present invention can effectively alleviate muscle atrophy caused by insulin resistance, and further, as a natural product-based material, has excellent human stability, and is used to treat and prevent muscle atrophy and muscle damage caused by an increase in insulin resistance. It can be widely used.
본 발명의 효과는 상기한 효과로 한정되는 것은 아니며, 본 발명의 상세한 설명 또는 청구범위에 기재된 발명의 구성으로부터 추론 가능한 모든 효과를 포함하는 것으로 이해되어야한다. The effects of the present invention are not limited to the above effects, and should be understood to include all effects that can be inferred from the configuration of the invention described in the detailed description or claims of the present invention.
도1은 싸리(Lespedeza bicolor) 추출물의 혈당, 혈장 HbA1c 농도 및 근육 무게 변화를 측정한 결과를 나타낸 것이다(A: 공복 혈당치 B: HbA1c 혈장 농도 C: 근육 무게).
도2는 싸리 추출물의 근육조직의 조직학적 변화를 확인한 결과를 나타낸 것이다.
도3은 싸리 추출물이 인슐린 신호전달 기전 관련 인자를 활성화시키는지 여부를 확인한 결과를 나타낸 것이다.
도4는 싸리 추출물이 근육에서 에너지 대사 관련 인자를 활성화시키는지 여부를 확인한 결과를 나타낸 것이다(A: SIRT1, SIRT3, SIRT4 및 PGC1-α의 발현량 B: pERK/ERK 및 pAPMK/AMPK의 발현량).
도5는 싸리 추출물이 근육에서의 염증반응 관련 인자의 활성을 억제시키는지 여부를 확인한 결과를 나타낸 것이다.1 shows the results of measuring changes in blood sugar, plasma HbA1c concentration, and muscle weight of the extract of Lespedeza bicolor (A: fasting blood sugar level B: HbA1c plasma concentration C: muscle weight).
Figure 2 shows the results of confirming the histological changes of the muscle tissue of the extract.
Figure 3 shows the results of confirming whether or not the extract of salicornia ssari activates factors related to the insulin signaling mechanism.
Figure 4 shows the results of confirming whether or not the salicornia extract activates energy metabolism-related factors in muscle (A: expression levels of SIRT1, SIRT3, SIRT4 and PGC1-α B: expression levels of pERK/ERK and pAPMK/AMPK ).
Figure 5 shows the results of confirming whether or not the ssari extract inhibits the activity of factors related to the inflammatory response in the muscle.
이하, 첨부된 도면을 참고하여 본 발명의 실시예에 관하여 상세히 서술하나, 하기 실시예에 의해 본 발명이 제한되지 아니함은 자명하다.Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings, but it is obvious that the present invention is not limited by the following examples.
실시예 1. 싸리(Example 1. Sari ( Lespedeza bicolorLespedeza bicolor ) 추출물 제조) Extract preparation
싸리 추출물 제조를 위해 싸리와 증류수의 비율을 맞추어 100°C에서 열수추출과 121°C 오토클레이브(autoclave)에서 1시간동안 가압추출을 진행한 뒤 추출물을 상온에서 냉각시킨 후 폴리테트라플루오로에틸렌(Polytetrafluoroethylene, PTFE) 필터를 이용하여 여과시켜 싸리와 추출액을 분리 하였다. 여과 후 남은 싸리는 50°C 의 건조기에 건조시킨 후 분쇄해 저장하고 여과기를 통과한 추출액은 감압진공농축기를 이용하여 농축하였다. 농축이 완료된 싸리 추출액은 50°C의 건조기에서 24시간 동안 건조하여 80 mesh 표준체망을 통과시켜 냉동 보관한 후 싸리 추출물 시료로 사용하였다.For the preparation of the extract, after performing hot water extraction at 100 °C and pressure extraction in a 121 °C autoclave for 1 hour, cooling the extract at room temperature and then cooling the extract at room temperature, polytetrafluoroethylene ( Polytetrafluoroethylene (PTFE) filter was used to separate the extract from the salicornia. After filtration, the remaining sorghum was dried in a dryer at 50 °C, pulverized and stored, and the extract passed through the filter was concentrated using a vacuum concentrator. The concentrated Ssari extract was dried for 24 hours in a dryer at 50 °C, passed through an 80 mesh standard sieve, and stored frozen, and then used as Ssari extract sample.
실험예 1. 인슐린 저항성이 증가한 쥐에서의 혈당, HbA1c농도 및 근감소 변화 측정Experimental Example 1. Measurement of changes in blood glucose, HbA1c concentration and muscle reduction in mice with increased insulin resistance
4주령 C57BL6마우스를 35mg/kg BW으로 2회 스트렙토조토신 (Streptozotocin, 이하 STZ)을 투여하였으며, 고지방 식이를 통해 인슐린 저항성 증가 모델인 제2형 당뇨 모델을 유도하였다. 인슐린 저항성이 유발된 마우스 중 일부에 100 mg/kg의 적은 양의 싸리(low dose of Lespedeza bicolor, 이하 LL) 및 많은 양의 250 mg/kg 싸리(high dose of Lespedeza bicolor, 이하 HL) 농도의 싸리 추출물을 각각 살균된 3차 증류수에 녹여 12주 동안 경구 투여하였다. 매 주 공복 혈당을 측정하였으며, 혈장에서 헤모글로빈A1c(Hemoglobin A1c, 이하 HbA1c)를 측정하였다.Four-week-old C57BL6 mice were administered streptozotocin (STZ) twice at 35 mg/kg BW, and a
그 결과, 인슐린 저항성이 증가된 당뇨 대조군(DMC)에서 정상군(CON)에 비해 높은 혈당과 HbA1c 농도를 보이며, 근육의 무게가 감소됨을 확인하였다.As a result, it was confirmed that the diabetes control group (DMC) with increased insulin resistance showed higher blood sugar and HbA1c concentration than the normal group (CON), and the muscle weight was decreased.
또한, 인슐린 저항성이 증가된 쥐에 싸리 추출물을 처리한 결과 혈당 및 HbA1c가 유의적으로 감소되고 인슐린 저항성에 의해 감소한 근육의 무게가 회복되는 것을 확인하였다(도 1). 구체적으로, HL군에서 혈당 감소 효과가 특히 높게 나타났으며, LL군에서 HbA1c의 감소 효과가 특히 높게 나타나는 것을 확인하였다(도 1). In addition, it was confirmed that the blood glucose and HbA1c were significantly reduced as a result of treatment of the mice with increased insulin resistance, and the muscle weight decreased by the insulin resistance was recovered (FIG. 1). Specifically, it was confirmed that the blood sugar reduction effect was particularly high in the HL group, and the HbA1c reduction effect was particularly high in the LL group (FIG. 1).
상기 결과는 싸리 추출물이 높은 혈당, HbA1c 증상 및 근손실을 완화시킬 수 있는 효능을 가지고 있으며, 이를 통해 상기 싸리 추출물이 인슐린 저항성 증가로 인하여 높아진 혈당, HbA1c, 근감소를 효과적으로 개선할 수 있음을 시사한다. The above results suggest that Ssari extract has the effect of alleviating high blood sugar, HbA1c symptoms and muscle loss, and through this, the Ssari extract can effectively improve increased blood sugar, HbA1c, and muscle reduction due to increased insulin resistance. do.
실험예 2. 근육의 조직학적 변화 학인Experimental Example 2. Histological change of muscle
싸리 추출물의 근손상 완화 효과를 확인하기 위해, 인슐린 저항성이 증가된 쥐의 근섬유의 크기 변화를 측정하였다.In order to confirm the muscle damage alleviation effect of the extract of salicorniflora, the change in the size of muscle fibers of mice with increased insulin resistance was measured.
정상군(CON)과 인슐린 저항성이 증가된 당뇨 대조군(DMC)에는 살균된 3차 증류수를 처리하였으며, 인슐린 저항성이 유발된 쥐 중 일부는 각각 100 mg/kg의 적은 양의 싸리(low dose of Lespedeza bicolor, 이하 LL) 및 250 mg/kg 의 많은 양의 싸리(high dose of Lespedeza bicolor, 이하 HL) 농도의 싸리 추출물을 살균된 3차 증류수에 녹여 12주 동안 경구 투여한 후 쥐의 근육조직의 일부를 채취하였다. The normal group (CON) and the diabetic control group (DMC) with increased insulin resistance were treated with sterilized tertiary distilled water, and some of the mice with insulin resistance were treated with a low dose of Lespedeza (100 mg/kg, respectively). bicolor (hereinafter referred to as LL) and 250 mg/kg of high dose of Lespedeza bicolor (hereinafter referred to as HL) were dissolved in sterilized tertiary distilled water and administered orally for 12 weeks. Was collected.
채취한 근육조직을 4% 포르말린에 고정시키고 파라핀으로 포매(embedding)시킨 후, 조직을 7μm 두께로 동결 박절하여 조직 절편을 제작하였다. 이 후 H&E (hematoxylin & eosin) 염색을 실시하여 광학현미경을 사용하여 관찰하였다.The collected muscle tissue was fixed in 4% formalin and embedded with paraffin, and then the tissue was frozen and cut to a thickness of 7 μm to prepare a tissue section. Thereafter, H&E (hematoxylin & eosin) staining was performed and observed using an optical microscope.
그 결과 인슐린 저항성이 증가된 당뇨 대조군(DMC)에 비해 싸리 추출물을 투여한 LL 및 HL군에서 근섬유의 크기가 회복됨을 확인하였다(도 2).As a result, it was confirmed that the size of the muscle fibers was recovered in the LL and HL groups administered with Ssari extract compared to the diabetic control group (DMC) with increased insulin resistance (FIG. 2).
실험예 3. 쥐 근육의 인슐린 신호전달 기전 관련인자 측정Experimental Example 3. Measurement of factors related to insulin signaling mechanism in rat muscle
싸리 추출물의 근손상을 완화시키는 기전을 확인하고자 인슐린 신호전달 관련 인자의 발현량을 측정하였다. The expression level of insulin signaling related factors was measured to confirm the mechanism of alleviating the muscle damage of the salicornia extract.
정상군(CON)과 인슐린 저항성이 증가된 당뇨 대조군(DMC)에는 살균된 3차 증류수를 처리하였으며, 인슐린 저항성이 유발된 쥐 중 일부는 각각 100 mg/kg의 적은 양의 싸리(low dose of Lespedeza bicolor, 이하 LL) 및 250 mg/kg 의 많은 양의 싸리(high dose of Lespedeza bicolor, 이하 HL) 농도의 싸리 추출물을 살균된 3차 증류수에 녹여 12주 동안 경구 투여한 후 쥐의 근육조직의 일부를 채취하였다. The normal group (CON) and the diabetic control group (DMC) with increased insulin resistance were treated with sterilized tertiary distilled water, and some of the mice with insulin resistance were treated with a low dose of Lespedeza (100 mg/kg, respectively). bicolor (hereinafter referred to as LL) and 250 mg/kg of high dose of Lespedeza bicolor (hereinafter referred to as HL) were dissolved in sterilized tertiary distilled water and administered orally for 12 weeks. Was collected.
싸리 추출물을 투여한 쥐의 근육조직 일부를 단백질 분해효소 억제제(proteases inhibitor)가 포함된 용해 완충액을 이용하여 균질화한 후 18407 g, 4 ℃ 에서 30분간 원심분리 하여 상층액을 얻었다. 얻은 상층액을 이용하여 BCA(Bichinchominic acid assay)를 통해 샘플 내 단백질을 정량 한 뒤 562 nm에서 흡광도를 측정하여 표준곡선과 비교하여 시료 단백질 농도를 계산하였다. 30 ug의 단백질을 폴리아크릴아마이드 겔(polyacrylamide gel)에서 전기영동하여 분리한 후 니트로셀룰로오스 멤브레인(nitrocellulose membrane)으로 옮겼다. 멤브레인에 3% BSA(Bovine serum albumin) 로 다른 단백질이 붙지 않도록 blocking 한 후 원하는 단백질에 대한 항체 IRβ, IRS-1, pAkT, Akt, GLUT4(Santa Cruz Biotechnology, CA, USA, 1:200) 및 α-tubulin(Sigma-Aldrich, 1:500)를 차례대로 반응시켰다. ECL 용액을 반응시킨 후 ChemiDoc XRS+ imaging system 통하여 단백질 정도에 따른 밴드의 굵기로 그 발현 정도를 확인하였다.A portion of the muscle tissue of the rat to which the extract was administered was homogenized using a lysis buffer containing a protease inhibitor, and then centrifuged at 18407 g for 30 minutes at 4°C to obtain a supernatant. Using the obtained supernatant, the protein in the sample was quantified through BCA (Bichinchominic acid assay), and the absorbance was measured at 562 nm, and the sample protein concentration was calculated by comparing it with a standard curve. 30 ug of protein was separated by electrophoresis on a polyacrylamide gel, and then transferred to a nitrocellulose membrane. Antibodies against the desired protein IRβ, IRS-1, pAkT, Akt, GLUT4 (Santa Cruz Biotechnology, CA, USA, 1:200) and α after blocking the membrane with 3% Bovine serum albumin (BSA) to prevent adhesion of other proteins. -tubulin (Sigma-Aldrich, 1:500) was reacted in order. After reacting the ECL solution, the expression level was confirmed by the thickness of the band according to the protein level through the ChemiDoc XRS+ imaging system.
그 결과, 인슐린 저항성이 증가된 당뇨 대조군(DMC)에 비해 싸리 추출물을 투여한 LL 및 HL군에서 IRβIRS-1, pAkt/Akt, GLUT4의 발현이 현저히 증가함을 확인하였다(도 3).As a result, it was confirmed that the expression of IRβIRS-1, pAkt/Akt, and GLUT4 was significantly increased in the LL and HL groups administered with Ssari extract compared to the diabetic control group (DMC) with increased insulin resistance (FIG. 3).
상기 결과를 통해 싸리 추출물은 감소된 인슐린 신호전달체계에 관여하는 인자들을 활성화시킴으로써, 당뇨에 의한 혈당 감소 및 근 소실을 완화시키는 효과를 가짐을 시사한다. Through the above results, it is suggested that Ssari extract has an effect of reducing blood sugar and alleviating muscle loss due to diabetes by activating factors involved in the reduced insulin signaling system.
실험예 4. 쥐 근육의 에너지 대사 관련인자 측정Experimental Example 4. Measurement of factors related to energy metabolism in rat muscle
싸리 추출물의 근손상을 완화시키는 기전을 확인하고자 근육에서의 에너지 대사 관련 인자의 발현량을 측정하였다. The expression level of energy metabolism-related factors in the muscle was measured to confirm the mechanism of alleviating the muscle damage of the salicornia extract.
정상군(CON)과 인슐린 저항성이 증가된 당뇨 대조군(DMC)에는 살균된 3차 증류수를 처리하였으며, 인슐린 저항성이 유발된 쥐 중 일부는 각각 100 mg/kg의 적은 양의 싸리(low dose of Lespedeza bicolor, 이하 LL) 및 250 mg/kg 의 많은 양의 싸리(high dose of Lespedeza bicolor, 이하 HL) 농도의 싸리 추출물을 살균된 3차 증류수에 녹여 12주 동안 경구 투여한 후 쥐의 근육조직의 일부를 채취하였다. The normal group (CON) and the diabetic control group (DMC) with increased insulin resistance were treated with sterilized tertiary distilled water, and some of the mice with insulin resistance were treated with a low dose of Lespedeza (100 mg/kg, respectively). bicolor (hereinafter referred to as LL) and 250 mg/kg of high dose of Lespedeza bicolor (hereinafter referred to as HL) were dissolved in sterilized tertiary distilled water and administered orally for 12 weeks. Was collected.
싸리 추출물을 투여한 쥐의 근육조직 일부를 단백질분해효소 억제제(proteases inhibitor)가 포함된 용해 완충액을 이용하여 균질화한 후 18,407 g, 4 ℃ 에서 30분간 원심분리 하여 상층액을 얻었다. 얻은 상층액을 이용하여 BCA(Bichinchominic acid assay)를 통해 샘플 내 단백질을 정량 한 뒤 562 nm에서 흡광도를 측정하여 표준 곡선과 비교하여 시료 단백질 농도를 계산하였다. 30 ug의 단백질을 폴리아크릴아마이드 겔(polyacrylamide gel)에서 전기영동하여 분리한 후 니트로세룰로오스 멤브레인(nitrocellulose membrane)으로 옮겼다. 멤브레인에 3% BSA(Bovine serum albumin) 로 다른 단백질이 붙지 않도록 blocking한 후 원하는 단백질에 대한 항체 pERK ,ERK, pAMPK, AMPK, SIRT1, SIRT3, SIRT4, PGC1-α (Abcam, Cambridge, MA, USA, 1: 10000)와 α-tubulin(Sigma-Aldrich, 1:5000)를 차례대로 반응시켰다. ECL 용액을 반응시킨 후 ChemiDoc XRS+ imaging system 통하여 단백질 정도에 따른 밴드의 굵기로 그 발현 정도를 확인하였다. A portion of the muscle tissue of the rat to which the extract was administered was homogenized using a lysis buffer containing a protease inhibitor, and then centrifuged at 18,407 g for 30 minutes at 4°C to obtain a supernatant. Using the obtained supernatant, the protein in the sample was quantified through BCA (Bichinchominic acid assay), and the absorbance was measured at 562 nm, and the sample protein concentration was calculated by comparing it with a standard curve. 30 ug of protein was separated by electrophoresis on a polyacrylamide gel, and then transferred to a nitrocellulose membrane. After blocking other proteins with 3% BSA (Bovine serum albumin) on the membrane, antibodies against the desired protein pERK ,ERK, pAMPK, AMPK, SIRT1, SIRT3, SIRT4, PGC1-α (Abcam, Cambridge, MA, USA, 1: 10000) and α-tubulin (Sigma-Aldrich, 1:5000) were reacted in order. After reacting the ECL solution, the expression level was confirmed by the thickness of the band according to the protein level through the ChemiDoc XRS+ imaging system.
그 결과, 당뇨 대조군(DMC)에 비해 싸리 추출물을 투여한 LL 및 HL군에서 pERK/ERK의 발현은 감소되는 반면, pAPMK/AMPK, SIRT1, SIRT3, SIRT4, PGC1-α 의 발현이 증가되는 것을 확인하였다(도 4).As a result, compared to the diabetic control group (DMC), the expression of pERK/ERK was decreased in the LL and HL groups administered with Ssari extract, whereas the expression of pAPMK/AMPK, SIRT1, SIRT3, SIRT4, and PGC1-α was increased. Was done (Fig. 4).
상기 결과는 싸리 추출물이 근육의 에너지 대사에 관여하는 인자들의 활성을 회복시킴으로써, 인슐린 저항성 증가로 인한 근소실을 완화시키는 우수한 효과를 가짐을 시사한다.The above results suggest that Ssari extract has an excellent effect of alleviating muscle loss due to an increase in insulin resistance by restoring the activity of factors involved in muscle energy metabolism.
실험예 5. 쥐 근육의 염증반응 관련인자 측정Experimental Example 5. Measurement of Factors Related to Inflammatory Response in Rat Muscle
싸리 추출물의 근위축을 완화시키는 기전을 확인하고자 근육에서의 염증반응 관련 인자의 발현량을 측정하였다.In order to confirm the mechanism of relieving muscle atrophy of the salicornia extract, the expression levels of factors related to the inflammatory response in the muscles were measured.
정상군(CON)과 인슐린 저항성이 증가된 당뇨 대조군(DMC)에는 살균된 3차 증류수를 처리하였으며, 인슐린 저항성이 유발된 쥐 중 일부는 각각 100 mg/kg의 적은 양의 싸리(low dose of Lespedeza bicolor, 이하 LL) 및 250 mg/kg 의 많은 양의 싸리(high dose of Lespedeza bicolor, 이하 HL) 농도의 싸리 추출물을 살균된 3차 증류수에 녹여 12주 동안 경구 투여한 후 쥐의 근육조직의 일부를 채취하였다. The normal group (CON) and the diabetic control group (DMC) with increased insulin resistance were treated with sterilized tertiary distilled water, and some of the mice with insulin resistance were treated with a low dose of Lespedeza (100 mg/kg, respectively). bicolor (hereinafter referred to as LL) and 250 mg/kg of high dose of Lespedeza bicolor (hereinafter referred to as HL) were dissolved in sterilized tertiary distilled water and administered orally for 12 weeks. Was collected.
싸리 추출물을 투여한 쥐의 근육조직 일부를 단백질분해효소 억제제(proteases inhibitor)가 포함된 용해 완충액을 이용하여 균질화한 후 18407 g, 4 ℃ 에서 30분간 원심분리 하여 상층액을 얻었다. 얻은 상층액을 이용하여 BCA(Bichinchominic acid assay)를 통해 샘플 내 단백질을 정량 한 뒤 562 nm에서 흡광도를 측정하여 표준 곡선과 비교하여 시료 단백질 농도를 계산하였다. 30 ug의 단백질을 폴리아크릴아마이드 겔(polyacrylamide gel)에서 전기영동하여 분리한 후 니트로세룰로오스 멤브레인(nitrocellulose membrane)으로 옮겼다. 멤브레인에 3% BSA(Bovine serum albumin) 로 다른 단백질이 붙지 않도록 blocking한 후 원하는 단백질에 대한 항체 MCP-1 (Cell Signaling Technology, Inc., Danvers, MA, USA, 1:1000)과 α-tubulin (Sigma-Aldrich, 1:5000)를 차례대로 반응시켰다. ECL 용액을 반응시킨 후 ChemiDoc XRS+ imaging system 통하여 단백질 정도에 따른 밴드의 굵기로 그 발현 정도를 확인하였다. A portion of the muscle tissue of the rat to which the extract was administered was homogenized using a lysis buffer containing a protease inhibitor, and then centrifuged at 18407 g for 30 minutes at 4°C to obtain a supernatant. Using the obtained supernatant, the protein in the sample was quantified through BCA (Bichinchominic acid assay), and the absorbance was measured at 562 nm, and the sample protein concentration was calculated by comparing it with a standard curve. 30 ug of protein was separated by electrophoresis on a polyacrylamide gel, and then transferred to a nitrocellulose membrane. Block the membrane with 3% Bovine serum albumin (BSA) so that other proteins do not adhere, and then antibody MCP-1 (Cell Signaling Technology, Inc., Danvers, MA, USA, 1:1000) and α-tubulin ( Sigma-Aldrich, 1:5000) was reacted in order. After reacting the ECL solution, the expression level was confirmed by the thickness of the band according to the protein level through the ChemiDoc XRS+ imaging system.
그 결과, 당뇨 대조군(DMC)에 비해 싸리 추출물을 투여한 LL 및 HL군에서 MCP-1의 발현 수준이 현저히 감소한 것을 확인하였다(도 5).As a result, it was confirmed that the expression level of MCP-1 was significantly reduced in the LL and HL groups administered with Ssari extract compared to the diabetic control group (DMC) (FIG. 5).
상기 결과는 싸리 추출물이 근육의 염증반응에 관여하는 인자들의 활성을 저하시킴으로써 인슐린 저항성 증가로 인한 근위축 및 근소실을 완화시킬 수 있음을 시사한다.The above results suggest that the salicornia extract can alleviate muscle atrophy and muscle loss due to an increase in insulin resistance by reducing the activity of factors involved in the inflammatory response of muscles.
전술한 본 발명의 설명은 예시를 위한 것이며, 본 발명이 속하는 기술 분야의 통상의 지식을 가진 자는 본 발명의 기술적 사상이나 필수적인 특징을 변경하지 않고서 다른 구체적인 형태로 쉽게 변형이 가능하다는 것을 이해할 수 있을 것이다. 그러므로 이상에서 기술한 실시 예들은 모든 면에서 예시적인 것이며 한정적이 아닌 것으로 이해해야만 한다. 예를 들어, 단 일형으로 설명되어 있는 각 구성 요소는 분산되어 실시될 수도 있으며, 마찬가지로 분산된 것으로 설명되어 있는 구성 요소들도 결합된 형태로 실시될 수 있다.The above description of the present invention is for illustrative purposes only, and those of ordinary skill in the technical field to which the present invention pertains will be able to understand that other specific forms can be easily modified without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative and non-limiting in all respects. For example, each component described as a single type may be implemented in a distributed manner, and similarly, components described as being distributed may also be implemented in a combined form.
Claims (10)
상기 근위축은 인슐린 저항성 증가로 인한 근위축증인 것인, 약학적 조성물.The method of claim 1,
The muscular atrophy is the muscular atrophy caused by an increase in insulin resistance, the pharmaceutical composition.
상기 싸리 추출물은 근육의 에너지 대사 관련 인자의 발현을 증가시키는 것인, 약학적 조성물. The method of claim 1,
The ssari extract is to increase the expression of factors related to energy metabolism in muscle, pharmaceutical composition.
상기 에너지 대사 관련 인자는 pERK/ERK, pAPMK/AMPK(phosphorylation of APMK/APMK), SIRT1(sirtuin1), SIRT3(sirtuin3), SIRT4(sirtuin4) 및 PGC1-α(phosphatidylglycerol phospholipase1- α) 중 어느 하나 이상인 것을 특징으로 하는, 약학적 조성물.The method of claim 3,
The energy metabolism-related factor is at least one of pERK/ERK, pAPMK/AMPK (phosphorylation of APMK/APMK), SIRT1 (sirtuin1), SIRT3 (sirtuin3), SIRT4 (sirtuin4), and PGC1-α (phosphatidylglycerol phospholipase1-α). Characterized in, pharmaceutical composition.
상기 싸리 추출물은 근육의 염증반응 관련 인자의 발현을 감소시키는 것을 특징으로 하는, 약학적 조성물. The method of claim 1,
The psoriasis extract is characterized in that to reduce the expression of factors related to the inflammatory response of the muscle, pharmaceutical composition.
상기 염증반응 관련 인자는 MCP-1(monocyte chemoattractant protein 1)인 것인, 약학적 조성물.The method of claim 5,
The inflammatory response-related factor is MCP-1 (monocyte chemoattractant protein 1), the pharmaceutical composition.
상기 추출물은 물, C1 내지 C4의 무수 또는 저급 알코올, 상기 물과 저급 알코올의 혼합 용매, 아세톤, 1,3-부틸렌글리콜, 에틸아세테이트, 클로로포름으로 이루어진 군에서 선택되는 1 종 이상의 용매로 추출하여 수득한 것인, 약학적 조성물.The method of claim 1,
The extract is one or more solvents selected from the group consisting of water, C 1 to C 4 anhydrous or lower alcohol, a mixed solvent of water and lower alcohol, acetone, 1,3-butylene glycol, ethyl acetate, and chloroform. The pharmaceutical composition obtained by extraction.
상기 근위축은 인슐린 저항성 증가로 인한 근위축증인 것인, 식품 조성물.The method of claim 8,
The muscle atrophy is the muscle atrophy caused by the increase in insulin resistance, food composition.
상기 식품 조성물은 건강기능식품인 것인, 식품 조성물.
The method of claim 8,
The food composition is a health functional food, food composition.
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