JP2012181070A - Health degree detection method - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a detection method capable of detecting a change in a process from a health state to a disease state concerning a subject individual.SOLUTION: A method for detecting a health degree in a subject individual includes: measuring 8-OHdG and HEL in urine collected from the subject individual; acquiring change tendencies in the blood HDL cholesterol and BMI of the subject individual within a normal range with the measured 8-OHdG density in the urine as an index; and acquiring the intake tendency of an antioxidant compound in the subject individual within the normal range with the measured HEL density in the urine as an index.

Description

  The present invention relates to a health level detection method, and more particularly to a method for detecting a health level (health level) in a healthy person.

  Although national medical expenses are increasing steadily, Japan has already reached a super-aging society, medical expenses are increasing rapidly, and restraining medical expenses is a national proposition. In particular, since lifestyle-related diseases such as dialysis and cardiovascular diseases and their complications occupy a large share of medical expenses, there is an urgent need to prevent lifestyle-related diseases early. In recent years, there has been a demand for early detection of disease and realization of early treatment in a period from a healthy state to a disease state. Therefore, an inspection method that can detect such changes in the course is also demanded.

  Various attempts have been disclosed for such inspection methods. For example, a technique for evaluating oxidative stress characteristics using two indexes of oxidative damage degree and oxidative defense ability in two-dimensional coordinates is disclosed (Patent Document 1). In this method, the production rate per weight of 8-OHdG (8-hydroxy 2'-deoxyguanosine) and other compounds in urine is evaluated.

  In addition, urinary 8-OHdG and HEL (hexanoyl lysine) are measured by the subject, a group with relatively high values is selected, broccoli sprout extract is administered, and these urine concentrations are measured. It has been disclosed that the antioxidant action of the extract was evaluated (Patent Document 2).

  Furthermore, in the evaluation of antioxidant capacity by measuring the redox balance of a living body, it is disclosed that 8-OHdG in urine is quantitatively measured as an evaluation factor (Patent Document 3).

  Furthermore, it has been reported that urinary 8-OHdG in healthy subjects decreased from the subject group by ingesting vegetables, juice, and the like (Non-patent Document 1). Moreover, the change of 8-OHdG in the accumulation | storage urine for 12 hours is measured by ingesting vegetable juice for a healthy soccer player (nonpatent literature 2).

JP 2003-310261 A JP 2008-74816 A JP 2003-315327 A

Asia Pac J Clin Nutr. 2007; 16 (3): 411-21. J. Physiol. Anthropol. 19 (6), p287-289 (2000)

  Indeed, 8-OHdG and HEL are DNA and lipid oxides, respectively, and are known as in vivo oxidative stress markers. And about 8-OHdG, it is also known that the concentration in urine falls by ingestion of an antioxidant compound. However, it was unclear whether it could be a marker for detecting changes in the process from health to disease, that is, changes in the process from health to disease in healthy individuals.

  In addition, these oxidative stress markers have little change in the measurement target in healthy individuals due to the homeostasis maintenance function of the living body, and it is difficult to easily detect small changes due to problems with detection sensitivity and accuracy. .

  Then, this invention aims at providing the detection method which can detect the change in the process from a healthy state to a disease state in a test individual.

  The present inventors pay attention to 8-OHdG and HEL which are known oxidative stress markers, and it is possible to detect HEL and 8-OHdG in urine simultaneously using an antigen-antibody reaction. We have found that it is useful to detect changes in the process from disease to disease. According to the present invention, the following means are provided.

  According to the present invention, there is provided a method for detecting the health level of a test individual, the step of measuring urinary 8-OHdG and HEL collected from the test individual, the measured urinary 8-OHdG concentration, and And a step of acquiring a tendency of the degree of health of the test individual in a healthy range using the HEL concentration as an index.

The acquisition step uses the measured urinary 8-OHdG concentration as an index to acquire the change tendency of the blood HDL cholesterol and BMI of the test individual in the healthy range, and uses the measured urine HEL concentration as an index to determine the healthy range. The above-mentioned health is obtained by obtaining the tendency of intake of the antioxidant compound of the test individual in the above and obtaining the change tendency of the systolic blood pressure of the test individual in the healthy range using the measured urinary 8-OHdG concentration and / or HEL concentration as an index. It may be a step of acquiring the degree tendency. The health degree may be a tendency for metabolic syndrome.
Using the measured urinary 8-OHdG concentration as an index, the change tendency of blood HDL cholesterol and BMI (Body Mass Index, hereinafter the same) of the test individual in the healthy range is obtained, and the measured urine HEL concentration is used as an index. As a method, a detection method is provided, comprising: obtaining a tendency of intake of an antioxidant compound of a test individual in a healthy range. The measurement step may be a step of detecting 8-OHdG and HEL simultaneously by an antigen-antibody reaction.

  According to the present invention, there is provided a method for evaluating food, the step of administering the food to a test individual, the step of measuring 8-OHdG and HEL in urine collected from the test individual, and the measured urine The change tendency of the blood HDL cholesterol and BMI of the test individual in the healthy range with the 8-OHdG concentration as an index was obtained, and the test in the healthy range was performed using the measured urinary HEL concentration as the index. An evaluation method is provided, comprising: acquiring an individual's tendency to take an antioxidant compound and evaluating the administration effect of the food.

  According to the present invention, there is provided a healthy human health test kit including a first antibody that specifically detects 8-OHdG and a second antibody that specifically detects HEL. The kit may contain an 8-OHdG mock antigen and a HEL mock antigen. It may include a solid phase body having a plurality of sets of the first antibody and the second antibody or a set of 8-OHdG mock antigen and HEL mock antigen on the surface.

It is a figure which shows the figure in the 2nd item simultaneous measurement antibody chip | tip whole layout, spot layout, the supply layout of a biotinylated antibody, and the signal obtained from the said chip | tip. It is a table | surface which shows the comparison result of 8-OHdG and HEL before and after vegetable intake intervention. It is a table | surface which shows the correlation with 8-OHdG, HEL, and other clinical data.

  The present invention measures 8-OHdG and HEL in urine collected from a test individual, and uses the measured urinary 8-OHdG concentration and HEL concentration as an index to indicate the tendency of the test subject's health in a healthy range. Is something to get. Furthermore, using the measured urinary 8-OHdG concentration as an index, the change tendency of blood HDL cholesterol and BMI of the test individual in the healthy range is obtained, and using the measured urine HEL concentration as an index, the test individual in the healthy range The anti-oxidant compound intake tendency is obtained, and the change tendency of the systolic blood pressure of the test individual in the healthy range is obtained using the measured 8-OHdG concentration and / or HEL concentration in urine as an index. The present inventors measure the urinary 8-OHdG and urinary HEL of a test individual, and in healthy individuals, these values can be associated with specific health indicators and also associated with intake of antioxidant compounds. I found that I could do it for the first time. To date, 8-OHdG has been known to be elevated in individuals already diseased. However, it has not been associated with a specific health index, and it has been unclear whether it corresponds to changes in the course from health to disease. It was also unclear whether these minute changes in the health status of healthy individuals could be found, regardless of the test individual or patient in the disease boundary region where 8-OHdG and HEL were high. All of these indicators are strongly related to metabolic syndrome, and these urinary concentrations were found to be useful for predicting metabolic syndrome. According to the present invention, it is possible to capture a subtle change (health level) in the health state of a subject. Hereinafter, embodiments of the present invention will be described in detail.

(Detection method)
The detection method of the present invention is a method for detecting the health level of a test individual, the step of measuring 8-OHdG and HEL in urine collected from the test individual, and the measured 8-OHdG concentration in urine And the process of acquiring the tendency of the test subject's health degree in a healthy range can be provided using HEL density | concentration as a parameter | index. Alternatively, using the measured urinary 8-OHdG concentration as an index, the change tendency of blood HDL cholesterol and BMI of the test individual in the healthy range is obtained, and using the measured urine HEL concentration as an index, the test individual in the healthy range The tendency of intake of antioxidant compounds in the urine was obtained and the change tendency of the systolic blood pressure of the test individual in the healthy range was obtained using the measured 8-OHdG concentration and / or HEL concentration in the urine as an index. Obtaining a health degree tendency.

  The test individual is an animal including a human. In addition to humans, pets such as dogs and cats and various animals are also included. Preferably, it is a human. Especially, it is preferable that a test individual exists in a healthy range, ie, is a healthy person. A healthy person is, for example, an individual who has not yet been diagnosed with a specific disease or syndrome or a border region thereof (known as a disease state) by a known index. For example, an individual who has not been diagnosed as metabolic syndrome can be mentioned.

The following criteria can be used for diagnosis of metabolic syndrome. That is, the waist circumference is 85 cm or more for men and 90 cm or more for women, and the following (1) to (3):
(1) Neutral fat 150 mg / dL or more, HDL cholesterol 40 mg / dL or less, or both;
(2) one or both of the highest (systolic) blood pressure 130 mmHg or higher, the lowest (diastolic) blood pressure 85 mmHg or higher; and (3) fasting blood glucose 110 mg / dL or higher,
An individual to which two or more items correspond is said to be an individual diagnosed with metabolic syndrome.

  In addition, healthy individuals include individuals who have not been diagnosed at the present time but are highly likely to develop so-called metabolic syndrome such as hypertension, diabetes and arteriosclerosis by genetic diagnosis.

  The health degree means a tendency in the course (healthy range) from a state of being a healthy person to a disease state. For example, the direction of change (trend), whether it is toward a health state or a disease state.

  In this method, urine collected from a test individual is used as a test sample. The collection form of urine is not particularly limited and may be a known form. The test sample is preferably 24-hour urine. This is because 8-OHdG and HEL are susceptible to daily fluctuations in urine concentration, meals, and the like.

(Measurement process of 8-OHdG and HEL)
This method includes the step of measuring urinary 8-OHdG and HEL. It is known that 8-OHdG is similarly decomposed by DNA-derived peroxide and excreted in urine, and HEL is a lipid peroxide-derived decomposed product and is concentrated as excrement in urine. know. The measurement step is preferably performed so that urine is periodically collected and the concentrations of 8-OHdG and HEL in the urine are measured.

  These measurement methods may be any known measurement method and are not particularly limited. For example, antigen-antibody reaction such as HPLC, ELISA, LC-MS and the like can be mentioned. An antigen-antibody reaction using an antibody chip is preferable. More preferably, it is an antibody chip which is a solid phase carrier provided with both mock antigens (such as 8-OHdG-BSA and HEL-BSA) on the surface so that 8-OHdG and HEL can be detected simultaneously, and more preferably, The antibody chip is provided with many sets of such mock antigens on the same solid phase carrier, and can simultaneously detect 8-OHdG and HEL for many samples. Such an antibody chip can be applied to the indirect competition method. The antibody value chip is an antibody chip that is a solid phase carrier having both antibodies on the surface so that 8-OHdG and HEL can be detected simultaneously, and more preferably, such an antibody set is used as the same solid phase carrier. It is an antibody chip that is provided in large numbers and that can simultaneously detect 8-OHdG and HEL for a large number of specimens. Such an antibody chip can be directly applied to the competition method. The 8-OHdG antibody, the HEL antibody, and these mock antigens can be obtained commercially or by a well-known technique of those skilled in the art, and those skilled in the art can also immobilize these solid-phase bodies using a known technique. It can be realized by appropriately using it.

(Various trends acquisition process)
This method can obtain the change tendency of blood HDL cholesterol and BMI of a test individual using the measured urinary 8-OHdG concentration as an index. Moreover, the change tendency of systolic blood pressure can also be acquired. According to the present inventors, it has been found that the 8-OHdG concentration in the urine of a healthy person is significantly inversely correlated with the blood HDL cholesterol level and significantly correlated with BMI and systolic blood pressure. From this, by measuring the urinary 8-OHdG concentration and monitoring it, it is possible to obtain a change tendency of these useful health indexes without performing a separate test. These known indicators are useful in that they are related to metabolic syndrome.

  Moreover, this method can acquire the tendency of intake of antioxidant compounds and the change tendency of systolic blood pressure of a test individual using the measured HEL concentration in urine as an index. According to the present inventors, it is known that the HEL concentration in the urine of a healthy person is significantly correlated with the intake tendency of antioxidant compounds such as vegetables and the systolic blood pressure. From this, by measuring the urinary HEL concentration and monitoring it, the intake tendency of the antioxidant compound and the change tendency of the systolic blood pressure can be grasped.

  According to the combination of these two kinds of oxidative stress markers, as described above, it can be associated with a plurality of useful indicators related to metabolic syndrome in test individuals in the healthy range. That is, since a sign of metabolic syndrome can be detected at an early stage, it is a test method useful for applying to healthy individuals for the onset and prevention of metabolic syndrome.

  In addition, by measuring 8-OHdG and HEL simultaneously on the same chip using an antigen-antibody reaction, these data can be acquired quickly and easily, and evaluation can also be performed quickly.

(Evaluation methods)
The food evaluation method of the present invention includes a step of administering the food to a test individual, a step of measuring urinary 8-OHdG and HEL collected from the test individual, and a measured urinary 8-OHdG concentration. A step of evaluating the administration effect of the food in a test individual in a healthy range can be provided using as an index. According to this evaluation method, the effect of the food intake on the health of the test individual can be evaluated using the urinary 8-OHdG concentration and the HEL concentration. According to this method, since the urine concentration of two markers closely related to each other is used as an index, highly accurate evaluation is possible.

  The food is not particularly limited. The food includes, for example, nutritional supplements such as so-called supplements. Moreover, since all the markers in this method are oxidative stress markers, they are useful for antioxidants or antioxidant foods related to oxidative stress, but the present method is applied without being limited to this type of food. be able to.

  In the evaluation step, using the measured urinary 8-OHdG concentration as an index, the blood HDL cholesterol and BMI blood HDL cholesterol and BMI change tendency of the test individual in a healthy range are obtained, and the measured urine HEL concentration Using the urine as an index, the intake tendency of the antioxidant compound in the test individual in the healthy range is obtained, and the change tendency of the systolic blood pressure of the test individual in the healthy range is measured using the measured 8-OHdG concentration and / or HEL concentration in the urine as an index. It is good also as a process of acquiring and analyzing these and evaluating the administration effect of the food.

(Detection kit)
The health test kit for healthy persons of the present invention can include a first antibody that specifically detects 8-OHdG and a second antibody that specifically detects HEL. According to such a kit, since the antibody specific to the two oxidative stress markers serving as an index of the health level of a healthy person is contained, the concentration of these markers can be easily measured by antigen-antibody reaction. The kit may further contain a 8-OHdG mock antigen and a HEL mock antigen. This is because the competing method can be easily implemented. Further, the kit preferably includes a solid phase body having a set of a first antibody and a second antibody / or a set of 8-OHdG mock antigens and HEL mock antigens on the surface. With such a form, two markers can be measured simultaneously and rapidly for a large number of specimens.

  EXAMPLES Hereinafter, although an Example is given and this invention is demonstrated concretely, this invention is not limited to these Examples.

  In this example, a multi-item chip targeted for detection of health level in a healthy person was produced. That is, a new layout that can measure a maximum of four items with one specimen of 8-OHdG and HEL was devised and examined for trial production. The layout on the chip and the layout of the spots are shown in FIG. As shown in FIG. 1, four spots were spotted at 1 mm intervals at the specimen dropping site. In addition, the sample was dropped onto the chip at 40 locations of 4 × 10.

  The antibody chip of this example was prepared by using a commercially available mock antigen as a methacrylic monomer containing an azo dye disclosed in Example 5 of JP-A-2003-329682 (an azo dye containing an amino group and a cyano group). Obtained) and copolymerized with a methacrylic monomer to obtain an azo polymer. A substrate on which this azopolymer was spin-coated was immobilized by a light immobilization method. In the antigen-antibody reaction, a sample urine, a biotinylated antigen, and a mixed solution of antibodies were dropped and reacted with each dropping site of the antibody chip, and then washed with PBS containing 0.01% Tween 20 (TPBS). . Further, the reaction with alkaline phosphatase labeled avidin was followed by washing with TPBS. Thereafter, a solution of CDP-STAR (chlorine-substituted 1,2-dioxetane chemiluminescence substrate of alkaline phosphatase, Roche Applied Science) was dropped onto the antibody value chip, and chemiluminescence was detected with a CCD camera.

  The light emission state is also shown in FIG. As shown in FIG. 1, 8-OHdG and HEL are detected as independent spots, respectively, and even when a sample in which two items are mixed is applied, each signal is quantified without interfering with each other. I understood that I could do it.

  In this example, a measurement method using healthy subjects as subjects was examined. That is, in order to confirm the correlation between the HEL / 8-OHdG two-item chip practicality and the measurement value of the oxidative stress marker on the antibody chip with human clinical data, human urine samples were obtained and implemented Measurement with a two-item chip similar to that obtained in Example 1 was performed.

  Regarding samples, we collected samples from healthy university students in a vegetable intervention test project. That is, a urine sample was provided in an intervention test to examine the effects of different vegetable intakes on the prevention of lifestyle-related diseases, and oxidative stress markers were measured for this urine sample. The subjects of the experiment were healthy male and female college students and graduate students who were divided into a group that continuously took 350g of vegetables per day for two weeks and a group that did not perform such intervention.

  General examinations such as blood and urine were performed twice before and after the examination. In principle, urine samples were collected for 24 hours. The urine sample was dispensed as an oxidative stress marker measurement sample using an antibody chip and measured by Healthcare Systems. The number of specimens was approximately 200 specimens before and after the intervention.

(Sample pretreatment)
Urine stored at −80 ° C. was dissolved on ice and then centrifuged at 3000 rpm, 4 ° C. for 10 min with a centrifuge, and the supernatant was taken as a measurement sample. Pre-treated samples, graded HEL and 8-OHdG standards, and appropriately diluted primary antibody solution were placed in a 384 plate, placed in a dispenser, and sampled and sampled by the program. . Then, it detected by the CCD camera by chemiluminescence through reaction of alkaline phosphatase labeled avidin. The signal of the obtained image was digitized, and data analysis was performed with analysis software.

(Results of dietary intervention test samples)
General test items and blood test items were measured for BMI, blood pressure, HDL-C, and the like. The measurement value by the antibody chip was a value corrected by 24H urine volume for those who successfully collected urine for 24 hours (HEL / day, 8-OHdG / day). About these, we analyzed before and after the comparison, correlation of the amount of change, correlation coefficient before the intervention (previous value). The results are shown in FIG. As shown in FIG. 2, in the result of comparison before and after the intervention, regarding HEL, a significant difference (both sides) was found with a significance value (both sides) of 0.04 before and after the intervention in a 24H urine collection successful person.

  FIG. 3 shows the relationship between the pre-intervention data and the general clinical examination items. As shown in FIG. 3, the 8-OHdG value corrected with the 24H urine volume had a correlation coefficient of −0.26 with the HDL-C value, and an inverse correlation was observed with a significance probability (two-sided) of 0.02. In addition, the 8-OHdG value and HEL value corrected with 24H urine volume have a correlation coefficient with the systolic blood pressure of 0.28 and 0.32, and positive (both sides) is positive with 0.01 and 0.00 Correlation was seen. Furthermore, the 8-OHdG value corrected with the 24H urine volume had a correlation coefficient of 0.23 with BMI, and a positive correlation with a significance probability (two-sided) of 0.03 was observed.

  It is said that HEL and 8OHdG in urine reflect the degree of oxidative stress in the body, and as a new finding, 8OHdG value in urine (in urine accumulated for 24 hours) correlates with BMI in HDL- It was found that there was an inverse correlation with C (good cholesterol), and the HEL value in the urine (24-hour accumulated urine) in the dietary intervention test was significantly higher than the non-intake group due to the intake of antioxidant compounds such as vegetables. The results show that both the 8-OHdG value in urine (24-hour accumulated urine) and the HEL value correlate with systolic blood pressure. I was able to find a measurement method that seems to be demonstrated. In particular, since the urinary 8-OHdG value and the HEL value are both strongly associated with clinical values that serve as indicators of the diagnostic criteria for metabolic syndrome, these measurements lead to metabolic syndrome in the healthy range. Can be predicted very early.

  As mentioned above, although the Example of this invention was described in detail, these are only illustrations and do not limit a claim. The technology described in the claims includes various modifications and changes of the specific examples illustrated above.

The technical elements described in this specification or the drawings exhibit technical usefulness alone or in various combinations, and are not limited to the combinations described in the claims at the time of filing. In addition, the technology exemplified in this specification or the drawings can achieve a plurality of objects at the same time, and has technical usefulness by achieving one of the objects.

Claims (9)

A method for detecting the health level of a test individual,
Measuring 8-OHdG and HEL in urine collected from a test individual;
Using the measured urinary 8-OHdG concentration and HEL concentration as indices, obtaining a trend of the health degree of the test individual in the healthy range;
A detection method comprising:   The acquisition step uses the measured urinary 8-OHdG concentration as an index to acquire the change tendency of the blood HDL cholesterol and BMI of the test individual in the healthy range, and uses the measured urine HEL concentration as an index to determine the healthy range. The above-mentioned health is obtained by obtaining the tendency of intake of the antioxidant compound of the test individual in the above and obtaining the change tendency of the systolic blood pressure of the test individual in the healthy range using the measured urinary 8-OHdG concentration and / or HEL concentration as an index. The detection method according to claim 1, which is a step of acquiring a tendency of degree.   The detection method according to claim 1, wherein the health degree is a tendency for metabolic syndrome.   The detection method according to any one of claims 1 to 3, wherein the measurement step is a step of simultaneously detecting 8-OHdG and HEL by an antigen-antibody reaction. A method for evaluating food,
Administering the food to a test individual;
Measuring 8-OHdG and HEL in urine collected from a test individual;
Using the measured 8-OHdG concentration in urine as an index, and evaluating the administration effect of the food in a test individual in a healthy range;
An evaluation method comprising:   The evaluation step obtains the change tendency of the blood HDL cholesterol and BMI of the test individual in the healthy range, and obtains the tendency of intake of the antioxidant compound of the test individual in the healthy range by using the measured urine HEL concentration as an index. The evaluation method according to claim 5, which is a step of evaluating the administration effect of the food. A first antibody that specifically detects 8-OHdG;
A second antibody that specifically detects HEL;
Including a health test kit for healthy people.   Furthermore, the kit of Claim 7 containing the mock antigen of 8-OHdG and the mock antigen of HEL.   9. The test kit according to claim 8, comprising a solid phase body having a plurality of sets of the first antibody and the second antibody / or 8-OHdG mock antigen and HEL mock antigen on the surface.