JP2016044146A - Muscle atrophy inhibitor and muscle growth promoter - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a nutrient effective in skeletal muscle atrophy inhibition.SOLUTION: The inventor of this application focused on a soybean peptide, examined the effect of the soybean peptide to muscle atrophy caused by a no-load model in terms of elderly people's muscle atrophy, and as a result of eager studies, found out that the soybean peptide had a skeletal muscle atrophy-inhibiting action and a skeletal muscle growth promoting action. That is, the invention relates to a muscle atrophy inhibitor and muscle growth promoter containing the soybean peptide as an active ingredient. The kind of soybean peptide is not particularly limited, but the peptide with a molecular weight of 5000 or less is preferable.SELECTED DRAWING: None

Description

  The present invention relates to a muscle atrophy inhibitor having an action of suppressing skeletal muscle atrophy and a muscle growth promoter having an action of promoting skeletal muscle growth.

  According to the Ministry of Health, Labor and Welfare, there are currently nearly 5 million people in need of assistance and nursing care in Japan. Among them, there are patients who are bedridden or have difficulty in walking due to lower limb disorders. In such patients, skeletal muscle atrophy progresses and muscle mass decreases, thus leading to functional decline. In addition, the aging of society has caused sarcopenia and skeletal muscle mass has decreased due to aging. Such a decline and transformation of behavioral force associated with skeletal muscle atrophy is a serious social problem, and elucidation of the mechanism and development of a treatment method are required.

  In addition, although there are harmful changes in the body such as atrophy of skeletal muscles, especially slow muscle fibers, which are anti-gravity muscles, and a decrease in muscular strength associated with them in the zero-gravity space, an effective countermeasure has not been developed yet. However, the suppression of muscular atrophy of astronauts who stay in space for a long period of time is an important issue that human beings must solve in the future in space development.

  In space flight under zero gravity, anti-gravity muscles (trunk muscle, quadriceps, triceps, etc.) are more likely to atrophy than voluntary muscles (biceps), and slow muscles become faster. Muscle atrophy under microgravity is most severe in the first one or two weeks and then slows down. After 6 months of flight, muscle mass, muscle, and maximum oxygen uptake are reduced by an average of 10-20%, and recovery after returning takes 6 weeks.

  Body composition varies greatly with age. In humans, after 30 years of age, muscle mass decreases at a rate of about 5% every 10 years. In addition, the rate of decline is said to accelerate further after age 60. This skeletal muscle atrophy is a muscle atrophy caused by an imbalance between protein synthesis and degradation in skeletal muscle and a decrease in the amount of muscle protein.

  The causes of muscular atrophy in the elderly include aging, malnutrition, and a decrease in physical activity. In addition, in the case of elderly people, a decrease in muscular strength is induced, which increases the risk of falls and fractures and often results in a need for nursing care such as being bedridden. Although it has been reported that exercise is effective as a method to improve this muscle atrophy, exercise is necessary to prevent or improve muscle atrophy in those who are unable to exercise or who are forced to bet. Other methods are essential.

  Therefore, an object of the present invention is to find a nutrient that is effective in suppressing skeletal muscle atrophy. It is a further object of the present invention to find nutrients that are effective in promoting skeletal muscle growth.

  The inventor of the present application focused on soybean peptide, examined the effect of soybean peptide on muscle atrophy caused by the no-load model from the viewpoint of muscle atrophy in elderly people, and as a result of earnest research, Have been found to have an atrophy-suppressing action and a skeletal muscle growth-promoting action.

  That is, the present invention is a muscle atrophy inhibitor and a muscle growth promoter containing soybean peptide as an active ingredient. Although the kind of soybean peptide is not particularly limited, preferably a soybean peptide having a molecular weight of 5000 or less is desirable.

  In the present invention, in order to prepare a preparation of a desired form, various components and additives are appropriately selected according to the use and form within a range not impairing the effects of the present invention, and one or more of them are selected. You may make it mix | blend together.

  By continuously ingesting the muscle atrophy inhibitor of the present invention even in an environment where muscle weight can be reduced by fixation or non-use with casts, aging, bedridden, exposure to weightless space, etc. It is possible to effectively suppress skeletal muscle atrophy. Moreover, by continuously ingesting the muscle growth promoter of the present invention, it is possible to effectively suppress skeletal muscle atrophy and promote skeletal muscle growth.

  Therefore, the muscle atrophy inhibitor and the muscle growth promoter of the present invention can prevent, for example, prevention of muscular weakness accompanying aging, prevention of physical strength of children, recovery of physical strength after illness (recovery of muscular atrophy due to fractures, etc.), postmenopausal It is useful for preventing muscle weakness, preventing muscular atrophy due to cachex, and preventing astronaut muscle atrophy. Furthermore, the muscle growth promoting agent of the present invention is also useful for increasing the strength of athletes, increasing the strength of the growing season, increasing the strength of workers, and the like.

It is a figure which shows the tail suspension method which is a muscle atrophy induction method with respect to a mouse | mouth. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result. It is a graph which shows an experimental result.

[Experiment 1] Effect of AM intake on muscle wet weight In this experiment, 20 male ICR retired mice (body weight; average: 42.3 ± 2.7 g, Nippon SLC Co., Ltd.) were used.

  In an environment of room temperature 24 ± 1 ° C., 12 hours day / night reversal light / dark cycle, food and drinking water were allowed to be freely ingested and reared on an individual gauge for 7 days. MF (Oriental Yeast Industry Co., Ltd.) was used as the diet, and pure water (Seiki Pharmaceutical Co., Ltd.) and Soy Peptide High Newt AM (Fuji Oil Co., Ltd.) prepared to 175 mg / mL were used as the drinking water. The molecular weight of soybean peptides used in this experiment was 5000 or less.

  As shown in the table below, these mice were treated with a pure water intake group (hereinafter “W group”), a pure water intake + tail suspension group (hereinafter “WTS group”), a soybean peptide high newt AM intake group (hereinafter “AM group”). “) Soy peptide high-newt AM intake + tail suspension group (hereinafter referred to as“ AMTS group ”) was divided into 5 groups of 5 animals. The tail suspension method devised by Emily R. Morey in 1949 was used as the inactivity method (see Fig. 1).

  Seven days after the tail suspension, mice in the W group, WTS group, AM group, and AMTS group were sacrificed under anesthesia, and the soleus muscle, which is the soleus muscle and the fast muscle, which were the body weight and the slow muscle, were collected. The weight of soleus and plantar muscles was measured to the second decimal place with an electronic balance (METTLER TOLEDO Co., Ltd.).

  In this experiment, the intake of soy peptide AM suppressed the muscle wet weight, skeletal muscle protein, and muscle cell atrophy. However, in this experiment, the tail suspension model affected the soleus muscle, but no significant difference was observed with only a slight decrease in the soleus muscle wet weight and muscle cell area. .

[Experiment 2] Effect of AM intake on skeletal muscle protein amount The skeletal muscle collected in Experiment 1 was stored and used at -80 ° C until analysis. The myofibrillar protein content was measured for absorbance with Odyssey (LI-OCR, Inc) at a wavelength of 595 nm based on the Bradford method. A calibration curve was drawn using BSA (BIO RAD, Bovine Serum Alubumin Standard Set) to determine the amount of protein. As shown in FIG. 2, skeletal muscle atrophy was clearly suppressed by administration of soybean peptide high-newt AM (hereinafter abbreviated as “AM” as necessary).

[Experiment 3] Effect of AM ingestion on the cross-sectional area of muscle cells In order to measure the cross-sectional area of muscle cells, the skeletal muscles were reared and collected with isopentane cooled in liquid nitrogen in the same manner as in Experiment 1 after the mice were raised and collected. What was transferred to liquid nitrogen and stored was used. A section of the middle abdomen of skeletal muscle was prepared with cryostat (LEICA, CM1510 S). HE staining was performed, and the muscle cell area was measured with 100 digital microscopes (KH-7700, HYROX) in each group.

<Statistical processing>
Data were expressed as mean ± standard deviation. The data were checked for significance using a one-way analysis of variance, and the significance of each group was examined by multiple comparison (Tukey). P <0.05 was considered significant.

  As shown in FIG. 3, atrophy caused by tail traction of the soleus was obviously suppressed by AM administration. Thus, AM seems to suppress skeletal muscle atrophy by some mechanism.

[Experiment 4] Effect of AM intake on TNFα expression that promotes muscle atrophy TNFα has been reported as a powerful factor inducing skeletal muscle atrophy by tail traction. Therefore, since there is a possibility that AM works as an inhibitor of TNFα, the activity of TNFα and gene expression in atrophy muscle were examined.

<TNF-α mRNA gene expression by RT-PCR method>
a. Preparation of samples The instruments used before collecting skeletal muscle were sterilized by autoclaving. For those that cannot be heated, RNase AWAY (Molevular Bio Productsa) was used. The collected skeletal muscle was stabilized with RNAlater (QUIAGEN).

b. RNA purification
RNA was purified using QUIAGEN's miRNeasy Mini kit.
The sample prepared in the above a was transferred to a tube (1.5 mL sterilized biomasher: Assist Co., Ltd.), and 700 μL of QlAzol Lysis Regent was added and homogenized. For homogenization, a power smasher from Nippi Corporation was used. The operation method followed the kit protocol. The centrifuge operation used Himac CR 15D of Hitachi Koki Co., Ltd.

c. Confirmation of RNA purity A portion of the RNA purified in b above was transferred to another tube and diluted 12-fold with 10 mM Tris-HCl pH 7.5. This was measured with A260 and A280 to confirm the degree of purification. Bio Photometer Plus (eppendorf) was used for the measurement. A suitable degree of purification is 1.8 to 2.1 for A260 / A280. The RNA concentration was calculated from the formula of RNA concentration = 40 μg / mL × A260 × dilution ratio, assuming that A260 = 1 was 40 μg / mL.

d. Synthesis of cDNA cDNA was synthesized using the RNA purified in b above. For synthesis of cDNA, Applied Biosystems High Capacity RNA-to-cDNA kit was used. The operation method followed the kit protocol. The enzyme reaction apparatus used was Gene Thermo Unit GTU-1615 manufactured by Taitec Corporation.

e. Analysis by RT-PCR ΔΔCt PCR was performed using the synthesized cDNA as a template. RT-PCR used ABI PRISM 7000 from Applied Biosystems. Analysis was performed by ΔΔCt method using β-actin as the endogenous control in a 96-well plate.

<Statistical processing>
Analysis was performed, Ct values of the target gene and the endogenous control gene (β-actin) were calculated on the software, and the average value and standard deviation of the Ct value of the target gene and the Ct value of the endogenous control were determined. The initial RNA amount was corrected using the Ct value of the endogenous control, and the standard deviation of the ΔCt value was calculated from the standard deviation value. The value obtained by subtracting the ΔCt value of the reference sample from the ΔCt values of the other samples was defined as the ΔΔCt value.

  The results are as shown in FIGS.

  It is considered that the concentration of TNFα in the blood was clearly decreased by AM administration and the activity of TNFα was suppressed. However, the gene expression in skeletal muscle is not considered to suppress the activity of AM TNFα.

[Experiment 5] Effect of ingestion of AM on IGF-1 expression that promotes muscle growth The above-mentioned TNFα-induced proteolytic inhibitory effect was sought for AM administration, but no clear change was observed. Therefore, considering the possibility that AM administration more positively suppresses skeletal muscle atrophy, the experiment was advanced focusing on the factor IGF-1, which induces skeletal muscle hypertrophy. It is known that IGF-1 significantly increases during skeletal muscle hypertrophy or during development of skeletal muscle.

  The experimental method for gene expression is basically the same as the method for measuring TNFα.

  The results are as shown in FIGS.

  As a result of the experiment, it was found that the concentration of IGF-1 was clearly increased by AM administration as shown in the figure. Furthermore, it was revealed that the gene expression of IGF-1 increases with AM administration. An increase in IGF-1R, which is a receptor for IGF-1, was also observed. From the above results, it is considered that the IGF-1 gene is expressed by AM administration, and the synthesized IGF-1 promotes protein synthesis by the autocrine mechanism and suppresses muscle atrophy.

  The following experiment was conducted to examine the effects of human consumption of soy peptide on skeletal muscle.

[Target of experiment]
There are 30 female college students aged 20-22 years old who belong to the lacrosse club of Jissen Women's University.

[Experiment grouping]
It was divided into 3 groups: 4 soy peptide high-newt AM (Fuji Oil Co., Ltd.) intake group, 3 placebo (lactose) intake group, and 10 control group not taking anything. The soy peptide high newt AM intake group and the placebo (lactose) intake group were ingested by students in the form of supplements. In addition, the molecular weight of the soybean peptite used in this experiment was 5000 or less.

[Supplement intake period]
99 days.

[Supplement intake]
In the morning, immediately after exercise, each day before bedtime 3 tablets (total 150mg) were taken by themselves.

[Measurement method of experiment]
The body composition was measured using a body composition analyzer Inbody manufactured by Biospace Corporation.

[Experimental result]
FIG. 9 shows the results of measuring the amount of body protein constituting the body. The group that took soybean peptide high-newt AM clearly increased after ingestion.

  Also, as shown in FIG. 10, the amount of skeletal muscle among the body proteins increased statistically significantly.

  Moreover, as shown in FIG. 11, the amount of fat decreased conversely with skeletal muscle.

[Summary of experimental results]
From the experimental results of each of the above-described examples, the skeletal muscle atrophy can be effectively suppressed and the skeletal muscle atrophy can be effectively suppressed by continuously providing soy peptide even under conditions that cause disuse muscle atrophy. It was confirmed that muscle growth was promoted.

  Therefore, the muscle atrophy inhibitor of the present invention is continuously ingested even in an environment where muscle weight can be reduced by fixation or non-use with casts, aging, bedridden, exposure to weightless space, etc. Thus, skeletal muscle atrophy can be effectively suppressed. Moreover, by continuously ingesting the muscle growth promoter of the present invention, it is possible to effectively suppress skeletal muscle atrophy and promote skeletal muscle growth.

Claims (2)

  A muscle atrophy inhibitor comprising soybean peptide as an active ingredient.   A muscle growth promoter containing soy peptide as an active ingredient.