KR101019506B1 - Indicator Using Cortisol and Dehydroepiandrosterone in Saliva for Evaluating the Function of Adrenal Gland and the Functional Evaluation Method of Adrenal Gland Using Thereof - Google Patents

Abstract

The present invention relates to an indicator capable of quickly and objectively determining an adrenal state of an individual and a method for measuring adrenal state using the indicator. In detail, the indicator of the present invention includes the following two-dimensional map; A cortisol distribution function that is a sum of a first Gaussian function and a second Gaussian function; And a dehydroisoandrosterone distribution function which is a sum of a third Gaussian function and a fourth Gaussian function, wherein the x-axis of the two-dimensional map is a concentration of cortisol and the y-axis of the two-dimensional map is dehydro. It is the concentration of isoandrosterone (DHEA; dehydroepiandrosterone) and the ratio of cortisol concentration and dehydroisoandrosterone concentration is the value, and the cortisol distribution function is a concentration distribution of cortisol measured from saliva of the normal group without disease, respectively. The sum of a first Gaussian function and a second Gaussian function obtained by fitting to two Gaussian distributions having a weight of 0, wherein the dehydroisoandrosterone distribution function is a concentration distribution of dehydroisoandrosterone measured from saliva of the normal group. Is a third Gaussian function and a fourth Gaussian obtained by fitting the two Gaussian distributions with respective weights. A summation of the functions, wherein the region composed of the x-axis and the y-axis of the two-dimensional map is the intersection of the first Gaussian function and the second Gaussian function, a specific percentile of the first or second Gaussian function, The third Gaussian function and the fourth Gaussian function may be divided into regions by a specific percentile of the third or fourth Gaussian function, or a combination thereof.

The indicators of the present invention can provide a quick and objective criterion in the assessment of the health status of patients with a variety of clinical and non-clinical studies and adrenal related diseases, and are useful indicators for determining adrenal status associated with stress. Can be used as

Saliva, cortisol, dehydroisoandrosterone, adrenal gland, indicator

Description

Indicator using Cortisol and Dehydroepiandrosterone in Saliva for Evaluating the Function of Adrenal Gland and the Functional Evaluation Method of Adrenal Gland Using Thereof of Saliva Cortisol and Dehydroisoandrosterone Concentration }

The present invention relates to an indicator capable of quickly and objectively determining an adrenal state of an individual and a method for measuring adrenal state using the indicator.

The adrenal gland is located in the shape of a cone above the kidneys. The adrenal glands are anatomically divided into the medulla and the cortex, and the adrenal medulla mainly produces catecholamines such as epinephrine and norepinephrine, and the cortex is aldosterone, cortisol and dehydro. It is known to produce mainly steroids such as isoandrosterone (DHEA; dehydroepiandrosterone, hereinafter DHEA).

The adrenal cortex is histologically divided into glomerulosa, fasciculata and redicularis. Cortisol is mainly produced from the fasciculata, and DHEA is mainly produced from the redicularis. All tissues use 17α-hydroxypregenolone as a precursor to produce cortisol and DHEA.

The hormone production of the adrenal glands has a very elaborate circadian rhythm and is the only endocrine organ in the body that secretes hormones in response to external stimuli (stresses). A patent has been applied for measuring and evaluating an individual's stress index by using a concentration of corticosteroids using this feature (WO 2002/62198).

Steroid production in the adrenal cortex is regulated by corticosteroids (ACTH) secreted by the pituitary gland. The secretion of the pituitary gland's corticosteroids has a constant circadian rhythm (the highest after climatic phase and gradually decreases to the lowest before bedtime). Of the many steroids produced and secreted by the adrenal cortex, cortisol best reflects the circadian rhythm of the pituitary gland. Cortisol is often referred to as the "stress hormone" because of the rapid increase in cortisol production in the adrenal glands.

Temporary stress is known to rapidly increase the level of catecholamines in the autonomic nervous system and cortisol levels along with the levels of corticosteroids in the pituitary gland. However, when the stressors disappear, elevated levels of catecholamines and cortisol will return to normal levels within 30 minutes.However, when stress is chronically acting, the daily rhythm, which has remained constant, will slow down or the cortisol levels will rise or fall abnormally. Loss forms.

Hormone levels have been measured mainly in the blood. From an endocrinology point of view, blood is not a hormone's target organ, but a simple hormone carrier. Most blood protein hormones (growth hormones, insulin, etc.) are well soluble in water and are present in activated form in the blood, but because steroids are insoluble in water, they are present in the blood in combination with other media (carrier proteins). When steroids are combined with the carrier, they do not act on the hormone receptors in the target organs, so they are called "inactive states" and cannot induce physiological mechanisms. Most of the steroids in the blood (95-99%) are present in this inactive state and only a fraction (1-4%) are known to be in an unbound state (active state). Therefore, the concentration of steroids in the blood can be said to be the concentration of most inactive rather than the concentration of the hormone in the active state.

Unlike blood, all of the steroids in saliva are not active with the carrier medium, and active steroids contained in the extracelluar fluid are transferred to the salivary glands by passive diffusion. will be. Therefore, the steroid concentration contained in saliva is recognized as an active steroid contained in the tissue.

In addition, saliva, unlike blood, can be easily collected by the general public in accordance with the individual's sleep cycle without the help of medical personnel, so it has the advantage of easily conducting research on the hormonal circadian rhythm of the individual. In other words, using saliva has the advantage of easy access to the concentration of active steroids and the hormonal circadian rhythms, which were difficult to access to blood. However, because the concentration of steroids in saliva is as low as 1 / 100-1 / 1000 of the blood, it cannot be analyzed by commercially available hormone analysis kits.

Among the various types of steroids in saliva, corticosteroids (cortisol and DHEA) have been widely used as tools for identifying stress-related symptoms (fatigue, depression, post-traumatic syndrome, migraine, immune disorders, etc.). In particular, the cortisol concentration, cortisol diurnal rhythm, and the corticol and molar C / D ratio (hereinafter referred to as molar C / D ratios), which indicate the balance of corticosteroids, are important indicators for identifying these symptoms. .

In patients with chronic fatigue symptoms, cortisol concentrations are lower than normal after waking up, and the cortisol daytime rhythm is also less clear than normal. Cortisol levels, especially those with severe stress, are not significantly different from normal subjects, but the high molar C / D ratio has been reported to result in excessive secretion of cortisol in functional adrenal glands.

Although cortisol is known only as a stress hormone to the general public, cortisol is at the center of the body's endocrine system and is responsible for regulating sugar, fat and protein metabolism, mineral balance, and immune balance. Cortisol's role, as can be guessed, is intimately linked to good health.

The symptoms that occur when the concentration of cortisol in the body is abnormally high and low are summarized in Tables 1 and 2, respectively.

Table 1 Symptoms of excessively high concentrations of cortisol

Table 2 Symptoms when the concentration of cortisol is low

As described above, although corticosteroids have been used in various fields of study as useful biomarkers of adrenal status associated with stress, the relative comparison / analysis by case is mostly quantified to assess adrenal status to date. There is no objective standard.

Accordingly, the present inventors objectively and quantitatively determine the state of the adrenal gland, in particular, the balance of the adrenal cortex hormones by using the concentrations of cortisol and DHEA in the saliva and the ratio of cortisol and DHEA in saliva that are active as means for evaluating the adrenal state. The aim of this study is to provide indicators that can be evaluated and to provide a method for evaluating adrenal function using these indicators.

It is an object of the present invention to provide an indicator capable of quickly and objectively determining an adrenal state of an individual and an adrenal state estimating method using the indicator.

Specifically, the cortisol concentration, the dehydroisoandrosterone concentration, the cortisol and dehydroisoandrosterone concentration ratios were measured from the saliva of the normal group without disease, and the measured cortisol concentration distribution and the dehydroisoandrosterone concentration were measured. Multidimensional indicators are divided into functions that are each formulated with a Gaussian mixture model, and areas based on which cortisol concentrations and dehydroisoandrosterone concentrations can be assessed. It is to provide, and to provide a method for evaluating the adrenal state of the individual by using the multi-dimensional indicator.

Indicators for evaluating adrenal function of the present invention include a two-dimensional map; A cortisol distribution function that is a sum of a first Gaussian function and a second Gaussian function; And a dehydroisoandrosterone distribution function which is a sum of a third Gaussian function and a fourth Gaussian function, wherein the x-axis of the two-dimensional map is a concentration of cortisol and the y-axis of the two-dimensional map is dehydro. It is the concentration of isoandrosterone (DHEA; dehydroepiandrosterone) and the ratio of cortisol concentration and dehydroisoandrosterone concentration is the value, and the cortisol distribution function is a concentration distribution of cortisol measured from saliva of the normal group without disease, respectively. Is a sum of a first Gaussian function and a second Gaussian function obtained by fitting two Gaussian distributions having a weight of. The dehydroisoandrosterone distribution function is a dehydroisoandrosterone measured from saliva of the normal group. The third Gaussian function and the fourth value obtained by fitting the concentration distribution of to two Gaussian distributions having respective weights The sum of the rightian functions and the region consisting of the x and y axes of the two-dimensional map is the intersection of the first Gaussian function and the second Gaussian function and the percentile of the first or second Gaussian function. ), Divided into regions by the intersection of the third Gaussian function and the fourth Gaussian function, the percentile of the third or fourth Gaussian function, or a combination thereof. .

In this case, as described above, the first to fourth Gaussian functions are functions having respective weights. That is, the cortisol distribution function is a sum of two normal distribution functions (first Gaussian function and second Gaussian function) multiplied by their respective weights, and the dehydroisoandrosterone distribution function is two normal distributions by which their respective weights are multiplied. The sum of the functions (third Gaussian function and fourth Gaussian function).

The indicator further includes a C / D map, wherein the x-axis of the C / D map is a concentration of cortisol, and the y-axis of the C / D map is a dehydroepiandrosterone (DHEA). Concentration, wherein the region consisting of the x- and y-axes of the C / D map is determined by the percentile of the concentration ratio distribution of cortisol and dehydroisoandrosterone as measured from the saliva of the normal group. The concentration of cortisol belonging to the percentile of the concentration ratio distribution is divided into / D region, wherein the one C / D region is a range (determined by the percentiles that divide the C / D region). Range and concentration range of dehydroisoandrosterone.

Preferably, the reference percentiles that divide the C / D zones are 10, 25, 75, and 90 percentiles, and specific ranges of one C / D zone are 0-10 percentiles, 10-25 percentiles, 25, respectively. To 75th percentile, 75 to 90th percentile, and 90 to 100th percentile.

Specifically, the region of the C / D map has a percentile of the concentration ratio distribution of 0 to 10 percentile, 10 to 25 percentile, 25 to 75 percentile, 75 to 90 percentile, and 90 to 100 percentile. It is divided into the concentration range of cortisol belonging to and the concentration range of D isoandrosterone, and it is characterized by 5 zones.

The boundaries of the zones belonging to the percentiles of the specific range of concentration ratio distributions are the angles of the concentration ratios of cortisol and dehydroisoandrosterone measured from saliva with the percentiles (10, 25, 75, and 90) that are the zone boundaries. It consists of the concentration of cortisol and the concentration of dehydroisoandrosterone, each boundary is characterized by having the following linear formula on the C / D map.

(Four equations bounded by five regions of a C / D map)

10th quartile boundary: y (concentration of dehydroisoandrosterone) = a * x (concentration of cortisol)

25th quartile boundary: y (concentration of dehydroisoandrosterone) = b * x (concentration of cortisol)

75th quartile boundary: y (concentration of dehydroisoandrosterone) = c * x (concentration of cortisol)

90th quartile boundary: y (concentration of dehydroisoandrosterone) = d * x (concentration of cortisol)

The concentration of the cortisol and dehydroisoandrosterone of the present invention is preferably a molar concentration, and the concentration ratio of the cortisol and the dehydroisoandrosterone is preferably a molar concentration ratio.

In this case, a is 0.13 to 0.6, b is 0.10 to 0.12, c is 0.055 to 0.075, and d is 0.04 to 0.05.

Upon fitting the cortisol concentration distribution, based on the work ratio of cortisol concentration and dehydroisoandrosterone concentration, the distribution of cortisol concentration having more than the daily concentration ratio is fitted to the first Gaussian function and having less than the work concentration ratio. The distribution of cortisol concentrations is fitted to the second Gaussian function.

Upon fitting the dehydroisoandrosterone concentration distribution, based on the ratio of cortisol concentration and dehydroisoandrosterone concentration, the distribution of cortisol concentration having less than the daily concentration ratio is fitted to the third Gaussian function, The distribution of cortisol concentrations with concentration ratios above is characterized by fitting to the fourth Gaussian function.

The ratio (the daily concentration ratio which is a reference when fitting the cortisol concentration distribution and the standard concentration ratio that is the reference when fitting the dehydroisoandrosterone concentration distribution) is a C / D ratio which is a ratio obtained by dividing the cortisol concentration by the dehydroisoandrosterone concentration, The specific concentration ratio is a specific C / D ratio within a region consisting of the maximum value and the minimum value of the C / D ratios measured from the saliva of the normal group without the disease, and the one C / D which is a reference for fitting the cortisol concentration distribution. The ratio is characterized by greater than the one C / D ratio that is the basis for fitting the dehydroisoandrosterone concentration distribution.

The cortisol concentration and the dehydroisoandrosterone concentration are characterized in that the molar concentration.

One C / D ratio, which is a reference when fitting the cortisol concentration distribution, is 13 to 16, and one C / D ratio, which is a reference when fitting the dehydroisoandrosterone concentration distribution, is 8 to 10.

An area of the 2D map may be an intersection point of the first Gaussian function and the second Gaussian function, 85 to 95 percentiles of the second Gaussian function, an intersection point of the third Gaussian function and the fourth Gaussian function, and The fourth Gaussian function is divided into nine regions by 85 to 95 percentiles.

The sum of weights of each Gaussian function of the first Gaussian function and the second Gaussian function is 1, the average of the first Gaussian function is 10 to 12, and the standard deviation is 18.5 to 20, and The average is 20 to 24, the standard deviation is 60 to 80, the sum of the weights of each Gaussian function of the third and fourth Gaussian functions is 1, and the average of the third Gaussian function is 0.7 to 1.1, The standard deviation is 0.1 to 0.15, the average of the fourth Gaussian function is 1.6 to 2, the standard deviation is 0.3 to 0.45.

The weights of the first Gaussian function and the second Gaussian function are 0.6 to 0.8 and 0.2 to 0.4, respectively, and the weights of the third and fourth Gaussian functions are 0.6 to 0.8 and 0.2 to 0.4, respectively. .

The concentrations of dehydroisoandrosterone and cortisol in saliva are characterized by radioimmunoassay, and the saliva is characterized by saliva of the awakening cortisol response (ACR) phase.

Adrenal function evaluation method according to the present invention is a) by measuring the concentration of cortisol and dehydroisoandrosterone in saliva by radioimmunoassay, dehydroisoandrosterone concentration, cortisol concentration, and the ratio of the concentration ratio of cortisol and dehydroisoandrosterone Obtaining; And b) obtaining a region of the indicator corresponding to the ratio of the concentrations of cortisol and dehydroisoandrosterone obtained in step a) using the above-described indicators.

The method of evaluating adrenal function includes a) using a cortisol distribution function including the first Gaussian function and the second Gaussian function, and the dehydroisoandrosterone distribution function by adding the third and fourth Gaussian functions. Obtaining the cortisol distribution percentile at the cortisol concentration value of the step and the dehydroisoandrosterone distribution function percentile at the dehydroisoandrosterone concentration value of step a) is preferably performed. Do.

In the adrenal function evaluation method, the step of obtaining the percentile region on the C / D map to which the concentration ratio of cortisol and dehydroisoandrosterone obtained in step a) is further performed using the above-described C / D map.

The index of the present invention provides a stochastic distribution function of the levels of active corticosteroids, particularly cortisol and dehydroisoandrosterone, and provides a two-dimensional region with two axes of cortisol and dehydroisoandrosterone. By providing a two-dimensional map divided into sections to assess the function of the adrenal gland, it is possible to accurately, objectively and easily assess the function of the adrenal gland as assessed by relative comparison, clinical care experience, or any variable criteria. There is a possible advantage, and it is possible to determine whether the disease associated with adrenal function or the disease causing stress, or the quantitative assessment of the stress itself is possible.

Evaluation method of the present invention can determine the adrenal function objectively, quickly and easily, more accurate and reliable evaluation can be performed by using the active cortical hormone, cortisol modeling the distribution of cortisol concentration of the population The advantage of comprehensively and accurately evaluating the adrenal function of individuals based on various objectives such as distribution function, dehydroisoandrosterone distribution function, C / D map, and two-dimensional map modeling the concentration distribution of dehydroisoandrosterone in the population There is this.

Hereinafter, the indicators and the evaluation method of the present invention will be described in detail with reference to the accompanying drawings. The drawings introduced below are provided by way of example so that the spirit of the invention to those skilled in the art can fully convey. Accordingly, the present invention is not limited to the drawings presented below and may be embodied in other forms. Also, throughout the specification, like reference numerals designate like elements.

Hereinafter, the technical and scientific terms used herein will be understood by those skilled in the art without departing from the scope of the present invention. Descriptions of known functions and configurations that may be unnecessarily blurred are omitted.

The present invention is characterized by using the cortisol concentration in the saliva, dehydroisoandrosterone (DHEA) concentration, cortisol and DHEA concentration ratio in the saliva that is active as an index for evaluating the adrenal function of the individual.

In addition, the saliva is characterized in that the saliva of the cortisol reactor by the gas phase. The saliva cortisol concentration gradually increases after the weather and reaches a maximum in about 30 minutes. This phenomenon is called the awakening cortisol response (ACR). Cortisol concentration after waking means the maximum degree of activation of the adrenal cortex that can be expressed in response to changes in the weather, and cortisol, DHEA, and the ratio of cortisol and DHEA during cortisol Hormonal balance, including stress disorders).

Hereinafter, the following examples were performed with the population living in the Republic of Korea, but the present invention is not limited by race and residential area, of course.

Measurement target

The normal subjects were 1009 patients (male: 506, female: 503) who had not been hospitalized for 2 years after the previous medical examination. The average age was 41.6 years. All of them were office workers, people with normal sleep cycles, except those with diabetes or heart disease, or those taking birth control pills.

Saliva collection

The saliva collection tube was sent to each mail or direct health checkup subject before the checkup. Subjects were asked to collect saliva samples 25-35 minutes after waking up on the day of test (average sampling time: 27 ± 2.3 minutes), and no food, tea, coffee smoking, and carbonated drinks were taken 30 minutes before saliva collection. Salivary cortisol concentrations were independent of flow rate, and sugar-free gums did not affect hormonal measurements, allowing chewing sugar-free gums when the subject needed saliva stimulation. They were asked to rinse their mouths with water before collecting saliva samples and to record how many minutes after salvation they were taken on the label provided.

Determination of Saliva Cortisol and DHEA Levels

Since steroid concentrations in saliva are dozens of times lower (20-30 times for cortisol) than blood, commercially available hormone assay kits are not suitable for measuring cortisol and DHEA in saliva. Therefore, precise measurement technology was required for salivary steroid analysis, and thus the applicants measured saliva cortisol concentration and DHEA concentration using radioimmunoassay, in particular, liquid phased-double antibody method.

In detail, pure cortisol and DHEA used as standard samples were purchased from Sigma-Aldrich. This standard cortisol or DHEA was dissolved in an organic solvent in which ethanol and propylene glycol were mixed in a 1: 1 ratio, and phosphorylation buffer solution (GPBS, pH = 7.2) were further diluted to prepare standard samples.

Cortisol antibodies were purchased from Biogenesis. Cortisol antibodies are 0.001% for aldosterone, 4.1% for 11-deoxycorticosterone, 5.7% for deoxycortisol, 0.5% for 21-deoxycortisol, and cortisol It cross-reacted to 1.2% for corticosterone and less than 0.001% for other steroid hormones.

The antibody of DHEA was purchased from AbCam, used 0.01% in Testosterone, 0.05% in Dihydrotestosterone, 5 alpha-Androstane-diol-3 beta. ) 0.01% for 5 alpha-Androstane-diol-3 alpha, 0.01% for Androstenediol, 0.08% for Androstenediol, 0.01% for Androsterone, 0.01% for Epiandrosterone, 0.01% for 5 beta-Androstane-3 alpha, 0.01% for Estradiol, 0.01% for Progesterone Cross-reacted with 0.1% in Pregnenolone, 0.02% in Sulfate DHEA, 0.01% in Cholesterol and 0.01% in Estrone.

The procedure is briefly described, and for a more detailed measurement, see the applicant's paper on hormonal determination using liquid phased-double antibody methods (Salivary cortisol and DHEA levels in the Korean population: age- related differences, diurnal rhythm, and correlations with serum levels.Yonsei Med J. 2007; 48: 379-88).

Into the test tube, each of the standard samples and 0.1 ml of the sample was placed, and GPBS was substituted for the antibody in the test tube without cortisol or DHEA to measure the nonspecific reaction, and the standard sample cortisol or DHEA Charcoal-stripped saliva was added to the included test tubes.

The total radiation dose of I125-labeled cortisol and DHEA was approximately 20,000 cpm / tube and antibodies were diluted to bind 30-35%. The total volume of the first reaction was 0.4 ml per tube, and the standard and saliva samples were reacted at 4 ° C. for 24 hours. Then, the reaction was further reacted at room temperature for 30 minutes for the second reaction, and then 0.1 ml of the secondary antibody was added to each test tube and reacted for another 30 minutes. Then, 0.5 ml of 10% PEG was added and reacted for 15 minutes and centrifuged at 1500 g for 20 minutes at 4 ° C. The supernatant was removed using a vacuum pump, and the radiation dose of the precipitate remaining in each test tube was measured using a gamma ray meter (Cobra 5005, PerkinElmer Life and Analytical Sciences). Each measured cortisol or DHEA level was calculated from a standard curve using RIASMART software (PerkinElmer Life and Analytical Sciences).

Cortisol had an intra-assay coefficient of variation of 8.1% and an inter-assay coefficient of variation of 7.5%. Analytical sensitivity of cortisol was 11 pg / tube. The intra-assay coefficient of variation of DHEA was 9.34%, and the inter-assay coefficient of variation was 6.51%. Analytical sensitivity of DHEA was 1 pg / tube.

Extraction of indicators

Molecular concentrations of cortisol and DHEA contained in saliva collected after meteorology from 1009 normal (men; 506; female; 503; mean age; 41.16 ± 6.57 years) without organic diseases using radioimmunoassay Based on the measurement results, as shown in FIG. 1 (c), the distribution of the concentration ratio of cortisol and DHEA (cortisol molar concentration / DHEA molar concentration, FIG. 1 (c), hereinafter molar C / D ratio) was obtained.

The concentration of cortisol measured was in the range of 3.2-48.8 nmol / L (FIG. 1A), and the concentration of DHEA was 0.14-3.74 nmol / L (FIG. 1B). The mean concentration of cortisol (mean ± SD) was 15.0 ± 7.8 nmol / L and the average concentration of DHEA was 1.3 ± 0.6 nmol / L.

Molar C / D ratio ranged from 3.1-39.8 (FIG. 1 (c)) and mean (mean ± SD) was 12.8 ± 5.5.

Shapiro-Wilk test confirmed that the distribution of cortisol, DHEA, and molar C / D ratios did not follow the normal distribution (p <0.001).

Cortisol concentrations were higher in females (15.5 nmol / L) than in males (14.5 nmol / L), but there was no significant difference between the sexes (p> 0.05). DHEA was also slightly higher in females (1.27 nmol / L) than in males (1.27 nmol / L), but there was no significant difference as in cortisol.Molar C / D ratio was also slightly higher in females (13.0) than in males (12.5). No significant difference was observed (p> 0.05).

In addition, although the age of the subjects is in their 20s and 50s, the DHEA concentration (20-50s; 1.2-1.3 nmol / L) and the molar C / D ratio (20-50s; 12.2-13.2) of each age group show a big change. There was no.

Molar C / D ratio is determined by two variables: the concentration of cortisol and DHEA. The change in the distribution of the molar C / D ratio according to the concentrations of cortisol and DHEA can confirm the direction of steroid production in the adrenal cortex.

Figure 2 (a) and 2 (b) shows the distribution of the molar C / D ratio according to the concentration of cortisol and DHEA, as shown in Figure 2 was not uniformly distributed according to the concentration of the hormone.

In order to examine the distribution of the Molar C / D ratio values according to the concentrations of cortisol and DHEA in more detail, as shown in FIG. 3, the values of the molar C / D ratio are divided by the interquartile range (IQR). , Molar C / D ratio values below the first quartile (25% percentile; below 9.0) and between the middle quartiles (25-75% percentile; 9.0-15.2) above the third quarter (75% percentile; above 15.2) The distribution is shown at which cortisol and DHEA concentrations they occur mainly.

Figure 3 (a) shows the cortisol concentrations corresponding to each of the quartile range of the value of the Molar C / D ratio, Figure 3 (b) shows each of the quartile range of the value of the Molar C / D ratio The corresponding DHEA concentration is shown.

As shown in FIG. 3 (a), the low Molar C / D ratio (1/4 quartile value (9.0) or less) was mainly distributed in the cortisol 6-10 nmol / L range, and the cortisol concentration was Almost none appeared above 24 nmol / L.

The populations in the middle quartile (1/4 ~ 3/4; 9.0-15.2) with Molar C / D ratios were mainly distributed in the range of 10-14 nmol / L of cortisol. However, the populations with molar C / D ratios above 3/4 quartiles (above 15.2) were distributed in almost all regions regardless of cortisol concentration.

These results indicate that steroid production and secretion in the adrenal cortex are not always balanced, especially in those with high molar C / D ratio (relatively high cortisol).

On the other hand, as shown in FIG. 3 (b), the low molar C / D ratio value was found in the entire range of the measured DHEA concentrations, and the molar C / D ratio was higher than the middle or 3/4 quartiles. Was mainly in the range of 0.6-1.2 nmol / L of DHEA, and the distribution frequency of these populations decreased drastically when the DHEA concentration was above 2.4 nmol / L.

These results indicate that the molar C / D ratio value depends on the concentrations of cortisol and DHEA. Based on the result of FIG. 3, the molar C / D ratio value is based on the specific molar C / D ratio value. The concentration distribution of cortisol belonging to below and the concentration of cortisol belonging to above a certain molar C / D ratio value were formulated into different Gaussian distribution functions, and the total cortisol concentration distribution was modeled by the sum of these two Gaussian functions. Is based on different specific molar C / D ratio values and formulates the concentration distribution of DHEA below a certain molar C / D ratio value and the concentration distribution of DHEA above a certain molar C / D ratio value with different Gaussian distribution functions. The total DHEA concentration distribution was modeled as the sum of these two Gaussian functions.

Therefore, the cortisol distribution function or the DHEA distribution function estimated by EM algorism based on a specific molar C / D and presented in GMM is shown in Equation (1) below.

-----(수식 1)

----- (Equation 1)

(Where x is the concentration of cortisol or DHEA, f (x) is the cortisol distribution function or DHEA distribution function, and f _i (x / u _i , σ _i ² ) is the mean u _i and the standard deviation (σ _i ) i th normal distribution function with w), where w _i is the weight of the i th normal distribution function.)

In detail, based on the results of FIG. 3 (a), a population having a Molar C / D ratio of less than quartile (9.0 or less) and a population having a molar C / D ratio of between quartiles (9.0-15.2) Cortisol distribution was estimated as the 1st component (Gaussian mixture model) of GMM (Gaussian mixture model, GMM), and the cortisol distribution of the population whose molar C / D ratio was above 3/4 quartile (15.2 or higher) It is estimated as the 2nd component of the second Gaussian function.

The mean and standard deviation of the 1st component group were 11.5 and 4.36 nmol / L, respectively, and the mean and standard deviation of the 2nd component group were 22.4 and 8.32 nmol / L, respectively. The proportion of each component (weight of each Gaussian function) in the total population was 68% and 32%.

Specifically, based on the results of FIG. 3 (b), the population in which the Molar C / D ratio is included in the interquartile (9.0-15.2) and the molar C / D ratio are 3/4 quartiles or more (15.2 or more) The DHEA distribution of the population was estimated as the 1st component of the GMM (third Gaussian function), and the DHEA distribution of the population with molar C / D ratio below the quartile (9.0 or less) was calculated as the 2nd component of the GMM (fourth Gaussian function). Estimated as According to this estimated model, the 1st component of the DHEA distribution function accounts for the distribution of DHEA in a population with low molar C / D ratios (below 1/4) and the 2nd component represents the intermolecular and high molar C / D. Explain the distribution of DHEA in the population with ratio. The mean and standard deviation of the groups corresponding to the 1st component of the DHEA distribution were 0.99 and 0.12 nmol / L, respectively, and the mean and standard deviation of the groups corresponding to the 2nd component were 1.87 and 0.38 nmol / L, respectively. The proportion of each component in the total group was 69.4% and 30.5%.

4 (a) shows the concentration distribution of cortisol and the cortisol distribution function obtained based on the measurement result of the normal group, and FIG. 4 (b) shows the concentration distribution of DHEA and the DHEA distribution function together. It is.

The molar C / D ratio shows the ratio of the concentration of cortisol and DHEA because 17α-hydroxypregnenolone is commonly used as a precursor in the biosynthesis of cortisol and DHEA in the adrenal cortex. It is used to assess the condition of the neocortex. In order to evaluate the function of the adrenal cortex two-dimensionally, as shown in FIG. 5, a C / D map and a multidimensional map, each of which was divided into quartiles of molar C / D ratio, were constructed.

In FIG. 5, the distribution of the molar C / D ratio together with the concentrations of cortisol and DHEA is shown in a scatter plot, and each symbol is different according to the percentile of the molar C / D ratio (90th percentile or more; Δ, 75th-90th percentile; ●, 25th-75th percentile; ○, 25th-10th percentile; ●, 10th percentile or less; △).

The C / D map of FIG. 5 shown with the distribution of the molar C / D ratio is divided into regions according to the percentiles of the molar C / D ratio, and is divided by four percentiles of 10, 25, 75, and 90. It is divided into four areas.

10th quartile boundary: y (concentration of dehydroisoandrosterone) = 0.1498 * x (concentration of cortisol)

25th quartile boundary: y (concentration of dehydroisoandrosterone) = 0.1109 * x (concentration of cortisol)

75th quartile boundary: y (concentration of dehydroisoandrosterone) = 0.06526 * x (concentration of cortisol)

90th quartile boundary: y (concentration of dehydroisoandrosterone) = 0.04952 * x (concentration of cortisol)

The 10th-quartile boundary is a region with a higher DHEA concentration than the cortisol concentration, indicating an inadequate response of adrenal function to internal / external stress. The 10-25 quartile boundary is an area in which the adrenal function may be deteriorated due to chronic stress. The 25-75 quartile boundary indicates the normal functioning of the adrenal function, and the 75-90 quartile boundary indicates the state of adrenal function enhancement due to acute stress. It is an area that needs to be relieved, and the area by the 90th quartile boundary indicates that the adrenal function is hyperactive due to acute stress, and the adrenal function is suspected along with acute stress, such as weight gain, depression, sexual dysfunction, etc. If accompanied by symptoms, this is an area that requires further examination.

The CD map described above with reference to FIG. 5 is preferably used together with the multi-dimensional map of FIG. 6 to be used as an index for evaluating the function of the adrenal gland.

The multidimensional map of FIG. 6, shown with the distribution of the molar C / D ratio, simultaneously shows the cortisol distribution function and the distribution function of DHEA on the x-axis (cortisol) and y-axis (DHEA). Distribution function and DHEA distribution function Intersection of 1st and 2nd components (cortisol; 19.04 nmol / L, DHEA; 1.61 nmol / L) and 90th percentile of each 2nd component (cortisol 30.8 nmol / L, DHEA The area is divided by 2.48 nmol / L).

According to the intersection and the 90th percentile, the multi-dimensional map region is divided into nine regions, and according to the reference cortisol concentration as shown in FIG. 6, a, b and c, 1, according to the reference DHEA concentration, Divided into 2 and 3, the nine clusters are collectively referred to as a1, a2, a3, b1, b2, b3, c1, c2, c3.

FIG. 7 illustrates the statistical characteristics of nine clusters divided by the criteria of the cortisol distribution function and the DHEA distribution function, and the graph of each zone shows the molar C / D ratios of the populations included in the zone. (box; 25-75 quartile, bar; median, whisker; 5-95 quartile) n is the number of populations included in the zone, and P0.9 is the molar C / D for each zone population. The D ratio value means the probability that it can be included in the 90th quartile or more, and P0.1 means the probability that each molar C / D value can be included below the 10th quartile in the molar C / D ratio value of the entire population. do.

Hereinafter, the features of each zone will be described in detail with reference to FIGS. 6 to 7.

Zone a1 is a population in which the concentrations of cortisol and DHEA fall below the intersection point (0.20-1.61 nmol / L) of the cortisol distribution function and the DHEA distribution function, respectively. Approximately 65% of the total population is included in zone a1 and the mean molar C / D ratio is 12.3 ± 4.7. Since the molar C / D ratio values are different in the population included in the zone a1, it is preferable to use the percentiles of the major molar C / D ratios as summarized in Table 3 below.

(Table 3)

Zone a2 is a population in which the cortisol concentration is in the 1st component (1.58-19.04 nmol / L) of the cortisol distribution function and the concentration of DHEA corresponds to the 2nd component (1.61-2.48 nmol / L) of the DHEA distribution function. This is a relatively high group. The mean molar C / D ratio for zone a2 is 7.6 ± 1.9 and approximately 9% of the total population belongs to zone a2.

Zone a3 is a population in which the cortisol concentration is in the 1st component (1.58-19.04 nmol / L) range of the cortisol distribution function and the concentration of DHEA is greater than or equal to 90th percentile (2.48 nmol / L or more) of the DHEA distribution function. Extremely high concentrations of DHEA. The mean molar C / D ratio for zone a3 is 5.0 ± 1.3 and about 1.3% of the population is in zone a3.

Zone b1 has a cortisol concentration ranging from the intersection of the 1st and 2nd components of the cortisol distribution function to the 2nd component 90th percentile (19.04-30.84 nmol / L) and the concentration of DHEA from the start of the 1st component of the DHEA distribution function. (0.20-1.61 nmol / L), a relatively high molar C / D ratio value (mean 19.7 ± 4.8), with approximately 8.8% of the population included in zone b1.

In zone b2, the cortisol and DHEA concentrations range from the intersection of the cortisol distribution function 1st component and the 2nd component to the 2nd component 90th percentile (19.04-30.84 nmol / L), and the concentration of DHEA is also the 1st component and 2nd component of the DHEA distribution function. Molar C / D ratio (12.0 ± 2.0), which represents a relatively high concentration of cortisol and DHEA, but corresponds to the average of the entire population, from the point of intersection to the range of 2nd component 90th percentile (1.61-2.48 nmol / L) 9% of the total population is in Zone b2.

Zone b3 has a concentration of cortisol and DHEA in the range of the 2nd component 90th percentile (19.04-30.84 nmol / L) from the intersection of the cortisol distribution function 1st component and the 2nd component, and the concentration of DHEA is 90th percentile of the 2nd component of the DHEA distribution function. This group is relatively high (more than 2.48 nmol / L) but has a relatively high cortisol concentration, but molar C / D ratio is rather low (8.4 ± 16) due to the non-ideally high concentration of DHEA. About 2% are in this group.

Zone c1 has a cortisol concentration of at least 90th percentile (30.84 nmol / L) of the 2nd component of the cortisol distribution function and a concentration of DHEA at the intersection of the 1st component and the 2nd component of the DHEA distribution function (0.20-1.61 nmol / L). Cortisol concentration was higher than DHEA concentration and molar C / D ratio was high (26.5 ± 4.8). About 2% are in this group.

Zone c2 shows that the concentration of cortisol is greater than 90th percentile (more than 30.84 nmol / L) of the 2nd component of the cortisol distribution function and the concentration of DHEA is greater than 90th percentile of the 2nd component from the intersection of the 1st and 2nd components of the DHEA distribution function (1.61-2.48). nmol / L), which is characterized by high cortisol and DHEA concentrations and slightly higher molar C / D ratio (18.2 ± 2.5). About 2.4% is in zone c2.

Zone c3 is the cortisol concentration of 90% percentile or more of the 2nd component of the cortisol distribution function (30.84 nmol / L) and DHEA concentration of 90% percentile or more (2.48 nmol / L) of the 2nd component of the DHEA distribution function. The molar C / D ratio is relatively close to the overall mean (12.1 ± 1.9). Approximately 0.6% fall into this group.

Zone a1 is a zone where the cortisol and DHEA concentrations are included in the first ginsian function distribution of each Gaussian distribution, indicating normal adrenal function, and according to Mcween's theory of stress response, This is an area where you can respond and adapt. However, even in this region, if the cortisol concentration is very low (cortisol concentration: 4.4 nmol / L or less), the adrenal insufficiency (adrenal insufficient) should be considered with reference to the symptoms in Table 2.

Zones a2, a3 and b3 have higher levels of DHEA rather than cortisol levels, indicating an improper response of adrenal function to internal / external stress, or taking anabolic steroids such as DHEA or polycystic follicles in women.

Zone b1 is a region where the concentration of cortisol is rather high and the concentration of DHEA is low, and the ratio of CD ratio is also relatively high. This area is Selye. According to H's General Adaption Syndrom, this is an area that indicates the state of adrenal function (end-resistance) due to acute stress. The risk of adrenal insufficiency is at risk if the subject falls within this area.

Zone b2 is a region where the cortisol concentration and the DHEA concentration are included in the overlapping portions of the first and second Gaussians of each Gaussian distribution, indicating that the adrenal function remains stable.

Zone C1 is a region with very high cortisol and low DHEA and relatively high CD ratios. This situation is Selye. According to H's Genetic Adaption Syndrom, this is an area showing the state of adrenal gland functioning (mid-resistance period) due to acute stress. However, hyperactivity of the adrenal gland is an area of risk due to factors of organic factors (adrenal hypertrophy, tumors, etc.), and therefore is an area to be accompanied by other overhaul of the subject. Symptoms corresponding to pseudo-Cushing's syndrome (Table 1) may appear, and if the subject is female, the symptoms of menstrual irregularity may occur.

Zones C2 and C3 have a high concentration of cortisol and high concentrations of DHEA, so that the ratio of CD ratios belongs to a normal region. Selye. According to H's Genetic Adaption Syndrom, this is an area that shows the state of adrenal function being advanced due to acute stress (initial resistance). This is the area where stressors act temporarily, or if there are no stressors but continue to be in this area, there is a risk of organic factors, including adrenal tumors. do.

Adrenal Function Assessment Using Extracted Indicators

Chronic musculoskeletal pain due to spinal structural abnormalities is considered inevitable perceived stress. In the present invention, saliva was collected for 30 minutes after waking up in patients with musculoskeletal pain (neck, shoulder or back pain) for at least 6 months, and the concentrations of cortisol and DHEA contained in saliva were measured (normal group for extraction of indicators). We used a method similar to the method of measuring saliva cortisol and DHEA concentrations.

The levels of cortisol and DHEA were not significantly different from those of normal subjects, but the molar C / D ratio was significantly higher than that of normal subjects. 8 is an example in which the concentrations of cortisol and DHEA of individual patients with different shapes by disease type are projected onto a multi-dimensional map. From an overall perspective, the cortisol concentrations of patients were distributed over 1st and 2nd components, and DHEA is mainly 1st. It was mainly distributed in the component. These results indicate that the molar C / D ratio of most patients is mainly distributed in b1, c1, and c2, where the molar C / D ratio is high, resulting in an imbalance in the production of corticosteroids.

Specifically, cortisol concentrations in healthy, painless subjects were predominantly distributed within the first and third gown cyanide distribution functions (ie, belonged to zone a1) and C / D ratios were also predominantly in the 25-75 percentile range.

12 of 38 patients with musculoskeletal pain that may be stressed due to pain are distributed in the second Gaussian distribution function, and 38 C / D ratios are included in the 75th-90th percentile range. It can be confirmed that is a functional hyperactivity state.

Furthermore, the distribution of cortisol and DHEA in patients with intermittent sleep disorders and fatigue symptoms and those with chronic symptoms does not appear to be significantly different, but patients with chronic sleep disorders have relatively low cortisol concentrations in the a1 region. Patients with chronic fatigue were mainly distributed in c1 and c2 areas, and patients with other chronic symptoms were mainly distributed in a1, b1 and c1.

As described above, the present invention has been described by specific matters such as the specific number of specimens and limited embodiments and drawings. However, the present invention is provided only for better understanding of the present invention, and the present invention is limited to the above embodiments. Various modifications and variations can be made by those skilled in the art to which the present invention pertains.

Therefore, the spirit of the present invention should not be limited to the described embodiments, and all of the equivalents or equivalents of the claims as well as the claims to be described later will belong to the scope of the present invention. .

FIG. 1 shows the distribution of cortisol concentration (FIG. 1 (a)), DHEA concentration (FIG. 1 (b)), and cortisol concentration and DHEA concentration (molar C / D ratio) measured from the saliva of the normal group (FIG. 1 (c). )),

2 is a scatter plot of the molar C / D ratio according to the measured concentrations of cortisol (FIG. 2 (a)) and DHEA (FIG. 2 (b)),

Figure 3 shows the distribution of the concentration of cortisol divided into quarter quarter or less, between 2 / 4-3 / 4 quartiles and above 3/4 quartiles based on the molar C / D ratio obtained by the measurement (FIG. 3 (a)) and DHEA concentration distribution (Fig. 3 (b)) is shown as a scatter plot,

4 shows the cortisol distribution function with the cortisol concentration distribution (FIG. 4 (a)), and shows the DHEA distribution function with the DHEA concentration distribution (FIG. 4 (b)),

5 shows a C / D map with a molar C / D ratio scatter plot,

6 illustrates a multi-dimensional map with a molar C / D ratio scatter plot,

7 shows statistical characteristics of each zone of the multi-dimensional map,

8 illustrates an example of adrenal function evaluation using a multi-dimensional map.

Claims (12)

As an indicator of adrenal function, The indicator is a two-dimensional map; A cortisol distribution function that is a sum of a first Gaussian function and a second Gaussian function; And a dehydroisoandrosterone distribution function which is a sum of the following third Gaussian function and the fourth Gaussian function. The x-axis of the two-dimensional map is the concentration of cortisol, the y-axis of the two-dimensional map is the concentration of dehydroisoandrosterone (DHEA) and the ratio of the cortisol concentration and the dehydroisoandrosterone concentration Has, The cortisol distribution function is a sum of the first Gaussian function and the second Gaussian function obtained by fitting the concentration distribution of cortisol measured from the saliva of the normal group without disease to two Gaussian distributions having respective weights. The dehydroisoandrosterone distribution function is a sum of the third and fourth Gaussian functions obtained by fitting the concentration distribution of dehydroisoandrosterone measured from the saliva of the normal group to two Gaussian distributions having respective weights. Is, An area composed of the x-axis and y-axis of the two-dimensional map includes an intersection point of the first Gaussian function and the second Gaussian function, a percentile of the first or second Gaussian function, and the third Gaussian function. And an adrenal function evaluation index divided into regions by a crossing point of the fourth Gaussian function, a percentile of the third or fourth Gaussian function, or a combination thereof. The method of claim 1, The indicator further includes a C / D map, The x-axis of the C / D map is the concentration of cortisol, the y-axis of the C / D map is the concentration of dehydroisoandrosterone (DHEA), Two or more C / D zones based on the percentiles of the concentration ratio distribution of cortisol and dehydroisoandrosterone measured from the saliva of the normal group divided into regions, Said one C / D zone is characterized in that the adrenal function evaluation index, characterized in that consisting of the concentration range of the cortisol and the concentration range of the dehydroisoandrosterone belonging to the percentile of the concentration ratio distribution of the specific range. The method of claim 1, Upon fitting of the cortisol concentration distribution, based on the work ratio of cortisol concentration and dehydroisoandrosterone concentration, the distribution of cortisol concentration having more than the work concentration ratio is fitted to the first Gaussian function, and less than the work concentration ratio. The distribution of cortisol concentrations with is fitted as the second Gaussian function, Upon fitting the dehydroisoandrosterone concentration distribution, based on the ratio of cortisol concentration and dehydroisoandrosterone concentration, the distribution of cortisol concentrations having less than the work concentration ratio is fitted as the third Gaussian function. A distribution of cortisol concentration having a daily concentration ratio or higher is fitted to the fourth Gaussian function. The method of claim 3, wherein The ratio is the C / D ratio, which is the ratio of the cortisol concentration divided by the dehydroisoandrosterone concentration, and the concentration ratio is the one in the region consisting of the maximum to minimum values of the C / D ratio measured from the saliva of the normal group without the disease. Adrenal function evaluation, characterized in that the C / D ratio, the one C / D ratio that is a reference when fitting the cortisol concentration distribution is greater than the one C / D ratio that is a reference when fitting the dehydroisoandrosterone concentration distribution Indicators. The method of claim 4, wherein Adrenal function evaluation index, characterized in that one C / D ratio which is a reference when fitting the cortisol concentration distribution is 13 to 16. The method of claim 4, wherein Adrenal function evaluation index, characterized in that the one C / D ratio as a reference when fitting the dehydroisoandrosterone concentration distribution is 8 to 10. The method of claim 3, wherein An area of the 2D map may be an intersection point of the first Gaussian function and the second Gaussian function, 85 to 95 percentiles of the second Gaussian function, an intersection point of the third Gaussian function and the fourth Gaussian function, and Adrenal function evaluation index, characterized in that divided into nine regions by the 85 to 95 percentile of the fourth Gaussian function. The method of claim 3, wherein The sum of weights of each Gaussian function of the first Gaussian function and the second Gaussian function is 1, the average of the first Gaussian function is 10 to 12, and the standard deviation is 18.5 to 20, and Average is 20 to 24, standard deviation is 60 to 80, The sum of the weights of each Gaussian function of the third and fourth Gaussian functions is 1, the average of the third Gaussian function is 0.7 to 1.1, the standard deviation is 0.1 to 0.15, and Adrenal function evaluation index, characterized in that the average is 1.6 to 2, the standard deviation is 0.3 to 0.45. The method of claim 8, The weights of the first Gaussian function and the second Gaussian function are 0.6 to 0.8 and 0.2 to 0.4, respectively, and the weights of the third and fourth Gaussian functions are 0.6 to 0.8 and 0.2 to 0.4, respectively. Adrenal function evaluation indicators. The method of claim 1, The concentration of dehydroisoandrosterone and cortisol in saliva is measured by adrenal function evaluation index, characterized in that. The method of claim 1, The saliva is an adrenal function evaluation index, characterized in that the saliva of the awakening cortisol response (ACR) phase. a) measuring the concentration of cortisol in saliva and dehydroisoandrosterone by radioimmunoassay to obtain dehydroisoandrosterone concentration, cortisol concentration, and the ratio of cortisol and dehydroisoandrosterone; And b) using the indicator of any one of claims 1 to 11 to obtain a region of said indicator to which the concentration ratio of cortisol and dehydroisoandrosterone obtained in step a) corresponds; Adrenal function evaluation method, characterized in that it is performed including.

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