KR102149079B1 - Method for predicting or determiing stroke using blood viscosity - Google Patents

Abstract

The present invention relates to a method of diagnosing a stroke using blood viscosity, identifying its subtype, and predicting the prognosis. The correlation between blood viscosity and acute or chronic signs of cerebrovascular disease, the cause of acute stroke and blood viscosity As a result of confirming the correlation, the correlation between blood viscosity and chronic radiologic signs of cerebrovascular disease, and the correlation between blood viscosity at or after acute stroke, the onset of stroke through diastolic and systolic blood viscosity levels, Since subtypes and prognosis can be predicted, it is related to providing information necessary for diagnosis of stroke, subtype discrimination, and prognosis monitoring using this.

Description

A method of predicting or diagnosing stroke using blood viscosity {METHOD FOR PREDICTING OR DETERMIING STROKE USING BLOOD VISCOSITY}

The present invention relates to a method for diagnosing stroke, identifying its subtype, and predicting prognosis using blood viscosity.

With the aging of the population, the incidence and prevalence of various senile diseases caused by aging are increasing, and in particular, central nervous system (CNS) diseases, including damage to the brain or spine, continue to increase. CNS disease, which has the highest proportion among them, is cerebrovascular disease, which ranks first in the single disease mortality rate, and there is a trend of 60,000 patients in Korea every year. The cerebrovascular disease is commonly referred to as a stroke, and is characterized by a sudden stop of circulation in a certain part of the brain due to an abnormal blood flow to the brain, resulting in loss of function of a nerve corresponding to that part. According to a recent study, it has been reported that the stroke causes more than 500,000 new strokes per year across all types of strokes. In addition, stroke is regarded as the first cause of permanent disorders such as muscle paralysis, consciousness disorder, visual impairment, and cognitive decline, unlike other diseases in which the defect is recovered during survival. Understanding the underlying etiology and developing treatment techniques Although the demand for is increasing, it is known that many recurrences even after treatment, causing enormous social and economic losses. According to a recent report, the direct costs (treatment and nursing) and indirect costs (loss of productivity, etc.) incurred for a stroke are estimated to be $43.3 billion per year in the United States alone.

The stroke is a hemorrhagic stroke caused by the bursting of blood vessels that supply blood to the brain due to shocks such as accidents, and the blood flow is temporarily blocked due to reduction of blood clots and perfusion, thereby supplying oxygen and nutrients to the brain. It can be classified as an ischemic stroke. In recent years, in Korea, the incidence of ischemic stroke continues to increase as compared to hemorrhagic stroke due to the westernization of diet and lifestyle. According to one study, ischemic stroke accounts for about 80% of all stroke types.

On the other hand, increased fibrinogen levels, whole blood viscosity, and plasma viscosity, which are the main factors of blood rheology, have been reported as risk factors or predictors of cardiovascular or cerebrovascular precautions (Coull BM, Beamer N, de Garmo P, Sexton G, Nordt F, Knox R, Seaman GV.Chronic blood hyperviscosity in subjects with acute stroke, transient ischemic attack, and risk factors for stroke.Stroke. 1991;22:162-8.). Whole blood viscosity can be determined through the complex interaction of blood components including red blood cells, white blood cells, platelets, fibrinogen and other plasma proteins (Grotta J, Ostrow P, Fraifeld E, Hartman D, Gary H. Fibrinogen, blood viscosity, and cerebral ischemia.Stroke.1985;16:192-8.). Whole blood viscosity measured at a high shear rate of 300 s ^-1 or more means systolic blood viscosity (SBV) (Kim H, Cho YI, Lee DH, Park CM, Moon HW, Hur M, Kim JQ). , Yun YM. Analytical performance evaluation of the scanning capillary tube viscometer for measurement of whole blood viscosity.Clin Biochem. 2013;46:139-42.). Blood viscosity at a high shear rate indicates the frictional properties of rapid blood flow, and is defined as having a velocity of 30 cm/s or more in a blood vessel with a large diameter (Noh HJ, Seo SW, Jeong Y, Park JE, Kim GH, Noh Y, Cho H, Kim HJ, Yoon CW, Ye BS, et al. Blood viscosity in subcortical vascular mild cognitive impairment with versus without cerebral amyloid burden.J Stroke Cerebrovasc Dis. 2014;23:958-66.). In addition, the whole blood viscosity measured at a low shear rate of 5 s ^-1 or less means diastolic blood viscosity (DBV). A similar brain microvascular disorder could be chronic cerebral ischemia due to cerebral small vessel disease (Pantoni L. Cerebral small vessel disease: from pathogenesis and clinical characteristics to therapeutic challenges. Lancet Neurol. 2010;9: 689-701.). Radiological signs such as leukemia, hiatus cerebral infarction and microbleeding are known as a result of small vessel damage in the brain and as chronic radiologic markers for cerebrovascular disease (Norrving B. Evolving concept of small vessel disease through advanced brain imaging. J Stroke. 2015;17:94-100.).

In the present invention, the correlation between blood viscosity and acute or chronic signs of cerebrovascular disease, correlation between the cause of acute stroke and blood viscosity, correlation between blood viscosity and chronic radiologic signs of cerebrovascular disease, and acute stroke The purpose of this study is to use blood viscosity for diagnosis or prognosis of stroke by confirming the correlation with blood viscosity at or after onset.

In order to achieve the above object, the present invention provides a method of providing information necessary for diagnosis or prognosis of a stroke.

In addition, the present invention provides a method for monitoring stroke treatment effect.

In addition, the present invention provides a method of providing information necessary for diagnosis of a stroke subtype.

In addition, the present invention provides a stroke subtype diagnostic kit including a quantification device for the viscosity of diastolic blood and systolic blood.

According to the present invention, the blood viscosity and subtype of stroke and cerebrovascular disease showed a significant correlation, and in particular, the onset, subtype and prognosis of stroke can be predicted through diastolic and systolic blood viscosity values. It can provide information necessary for diagnosis of stroke, subtype identification, and prognosis monitoring.

1 shows a patient registration flow chart.
Figure 2 is a diagram confirming the correlation between diastolic blood viscosity and red blood cell volume (a), fibrinogen level in blood (b), systolic blood viscosity and red blood cell volume (c) and fibrinogen level in blood (d):
*: p <0.05;
DBV: diastolic blood viscosity;
SBV: systolic blood viscosity; And
Hct: hematocrit.
Figure 3 is a diagram confirming the blood viscosity, red blood cell volume fibrinogen, and BUN/Cr ratio according to the stroke subtype:
DBV: diastolic blood viscosity;
SBV: systolic blood viscosity; And
Hct: volume of red blood cells.
Figure 4 is a laboratory findings including diastolic blood viscosity (a), systolic blood viscosity (b), red blood cell volume (c), and fibrinogen level in blood (d) and their relationship with chronic radiologic signs of cerebrovascular disease. This is a diagram confirming the correlation:
*: p <0.05;
DBV: diastolic blood viscosity;
SBV: systolic blood viscosity;
Hct: red blood cell volume;
PVH: periventricular hyperintensity; And
DWMH: Deep white matter hyperintensity.
Figure 5 is a diagram confirming the blood viscosity characteristics over time in SAO and non-SAO stroke:
*: p <0.05;
DBV: diastolic blood viscosity;
SBV: systolic blood viscosity; And
SAO: Arterioles occlusion.
6 is a diagram illustrating ROC analysis of diastolic blood viscosity over time.
7 is a diagram showing an ROC analysis of systolic blood viscosity over time.

Hereinafter, the present invention will be described in detail as an embodiment of the present invention. However, the following embodiments are presented as examples of the present invention, whereby the present invention is not limited, and the present invention can be variously modified and applied within the scope of equality interpreted from the description of the claims to be described later. .

In one aspect, the present invention provides the steps of obtaining blood isolated from a subject; And it relates to a method for providing information necessary for diagnosis or prognosis of a stroke, comprising measuring blood viscosity.

The term "detection" or "measurement" as used in the present invention means to confirm or quantify the presence of a detected or measured object.

The term "diagnosis" as used in the present invention is used to determine the susceptibility of a subject to a specific disease or disorder, or to determine whether a subject currently has a specific disease or disorder (e.g., Identification of a stroke or its subtype), determining the prognosis of a subject with a specific disease or condition, or therametrics (e.g., monitoring the condition of a subject to provide information about treatment efficacy) Thing). For the purposes of the present invention, the diagnosis means determining whether a stroke has occurred, distinguishing a stroke subtype, and determining whether a specific stroke subtype has occurred by measuring blood viscosity, and also includes determining the development and alleviation of a disease.

In one embodiment, the blood separated from the subject may be separated at the time of the onset of the stroke, 1 week after the onset, or 5 weeks after the onset, and may be separated during the systolic or diastolic phase.

In one embodiment, the blood viscosity may be the viscosity of whole blood.

In one embodiment, the blood viscosity may be systolic blood viscosity (SBV) or diastolic blood viscosity (DBV).

In one embodiment, the stroke is selected from the group consisting of small artery occlusion (SAO), large artery atherosclerosis (LAA), cardioembolism (CE), and cryptogenic stroke. It may be any one of the stroke subtypes, more preferably arteriole occlusion.

In one embodiment, the diastolic blood viscosity at the time of onset is greater than 296.1 mp or the systolic blood viscosity is greater than 37 mp; Diastolic blood viscosity greater than 245.9 mP or systolic blood viscosity greater than 39 mP 1 week after onset; And 5 weeks after onset, if the diastolic blood viscosity exceeds 225.2mp or the systolic blood viscosity exceeds 34.7mp, it can be determined that the risk of arteriolar obstruction is high.

As used herein, the term "at the time of onset" refers to the onset of a disease expected to be a stroke, and before medical intervention, including hydration therapy.

The method of checking the blood viscosity may be performed through various devices and methods known in the art, for example, but is not limited thereto, but is measured using a scanning capillary tube viscometer. Can be.

The term “subject” as used herein refers to human and non-human primates such as chimpanzees, and other ape and monkey species; Farm animals such as cows, sheep, pigs, goats and horses; Pet mammals such as dogs and cats; It refers to any member of the mammalian class, including, but not limited to, laboratory animals, including rodents such as mice, rats and guinea pigs. These terms do not denote a specific age or gender. Thus, adult and newborn subjects, as well as fetuses, whether male or female, are intended to be included within the scope of this term.

In one aspect, the present invention provides the steps of obtaining blood isolated from a subject; And to a method of monitoring the effect of treating stroke, comprising measuring blood viscosity.

In one embodiment, the treatment effect may be monitored by obtaining diastolic or systolic blood while performing treatment after the onset of a stroke and checking its viscosity.

In one aspect, the present invention relates to a stroke subtype diagnostic kit comprising a quantification device for the viscosity of diastolic blood and systolic blood.

The present invention will be described in more detail through the following examples. However, the following examples are only for embodiing the contents of the present invention and the present invention is not limited thereto.

Example 1. Patient registration

From July 2013 to February 2014, 127 consecutive acute ischemic stroke patients who were hospitalized within 3 days after the start of the stroke were enrolled, of which 70 samples for blood viscosity were obtained. . Acute ischemic stroke was confirmed in all patients by diffusionweighted magnetic resonance imaging (MRI). To homogenize the study population, 16 patients with strokes lasting 3 days or more, 4 patients with no confirmed stroke by MRI, 6 patients with strokes of other conclusive causes, and 7 with severe medical or neurological symptoms. Were excluded. In addition, 7 samples were deleted due to technical errors or poor quality. That is, the data of 63 patients were analyzed (FIG. 1), which is 7 patients with non-specific dizziness, 5 patients with temporary weakness without vascular cause, and vestibular neuronitis. 16 patients, including 3 patients and 1 patient with hypochondriasis, were classified as stroke-like groups. Patients suspected of having a transient ischemic attack (transient ischemic attack (TIA)) were not included in the stroke-like group because blood viscosity may be affected. Specifically, in order to minimize the dilution effect on blood viscosity by hydration therapy, the blood viscosity at the time of onset was measured from blood samples obtained in the emergency room before medical intervention including hydration therapy. At the time of onset, blood viscosity was measured twice in the first and fifth weeks after the measurement of blood viscosity at the time of onset, and in order to minimize the temporary dilution effect by oral water intake, it was measured from blood samples obtained in a fasting state. In addition, patients were instructed not to drink or eat anything, including water, after taking the evening medication until blood samples were taken early the next morning. As a control for comparison with blood samples from patients with acute ischemic stroke, hemoglobin samples from patients diagnosed as similar to stroke during the same study period were collected. Written consent was obtained from the patients, and was performed with the approval of the institutional review committee of Konkuk University Hospital.

Example 2. Statistical Analysis

In the following examples, the difference in vascular risk factors or test findings was evaluated using a chi-square test for dichotomized variables and an independent sample t test, and if appropriate, the Mann-Whitney U test for continuous variables or It was evaluated using ANOVA (analysis of variance) along with Tukey's post-hoc test. The correlation between SAO (small artery occlusion) stroke and test findings was evaluated using the calculation of crude ratios with 95% confidence intervals or adjusted odd ratios through univariate or multivariate logistic models. . The association between test findings including DBV, SBV, red blood cell volume and fibrinogen level was analyzed using Pearson correlation. The association between examination findings and radiographic evaluation results was also analyzed using Pearson correlation. There were considered significant differences at p<0.05 and all statistics were performed using SPSS version 17.0.

Example 3. Patient demographics and clinical characteristics

Vascular risk factors affecting blood viscosity and test findings were identified, and radiological evaluation was performed. Specifically, hemoglobin, hematocrit, leukocyte cells, platelets, blood proteins, fibrinogen, Ddimer, BUN/Cr (blood urea nitrogen-to-creatinine) ratio and lipid characteristics were confirmed as vascular risk factors. Patients were considered hypertensive if they used antihypertensive drugs or if their blood pressure was ≥140/90 mmHg in repeated measurements. Similarly, if you are taking diabetic medications, or your fasting blood sugar is ≥126 mg/dl, you were considered to have diabetes, and you are taking a lipid-lowering drug or fasting overnight if your cholesterol level is ≥220 mg/dl or your LDL cholesterol level is ≥140 mg/dl. In the case of dl, it was considered to be hyperlipidemia. Smoking was defined by the current smoking status. Stroke subtytes were classified according to the Trial of Org 10172 in the acute stroke treatment classification.

In addition, radiological evaluation was performed. Specifically, using a 3 T MR system (Signa HDx; GE Medical Systems, Milwaukee, WI, USA), T1-weighted images, T2-weighted images, FLAIR (fluid attenuation inversion recovery) images and T2* Gradient-echo images were obtained. T1-weighted images and T2-weighted images were obtained during the same imaging session in the same orientation and slice position: Matrix, 128; Field of view, 280 x 280 mm; Section thickness, 5 mm; Spacing between sections, 2 mm. Radiological signs associated with small blood vessel diseases of the brain have been defined, for example, white matter hyperintensities, lacrimal lacunes and microbleeds. The severity of white matter hyperintension on FLAIR images was evaluated on the Periventricular Hyperintensity (PVH) and Deep White Matter Hyperintensity (DWMH) scales according to the Fazekas scale. Lacrimal cerebral infarction is a small, distinct lesion (diameter <15 mm) with a low signal on the T1-weighted image, a strong signal on the T2-weighted image, and perilesional gliotic changes on the FLAIR image.White matter ischemic. changes). Unlike Virchow Robin space or ischemic white matter changes, hiatus cerebral infarction was distinguished by a high-intensity rim around the cavity on FLAIR images. Acute lacunar infarctions, expressed as stroke index at patient registration, were not included in the analysis. Only chronic hiatal cerebral infarction was included, regardless of whether symptoms were present or not. Microbleeding was defined as a small, uniform and rounded loss of signal lesion (<10 mm in diameter) on the T2* gradient-echo image. Statistical analysis was performed as described in Example 2 above.

The clinical demographic, radiological evaluation and examination findings of 63 enrolled patients are shown in Table 1 above. The most common stroke subtypes are SAO (n = 25, 39.7%), LAA (n = 23, 36.5%), CE (n = 12, 19.0%) and cryptogenic stroke (Cryp; n = 3, 4.8). %). The prevalence of vascular risk factors excluding atrial fibrillation did not differ between stroke subtypes.

Example 4. Analysis of correlation between blood viscosity, red blood cell volume, and fibrinogen level in blood

To measure the whole blood viscosity, a scanning capillary tube viscometer (BVD-PR01, Bio-Visco Inc., Korea) was used. The scanning capillary viscometer can simultaneously measure systolic blood viscosity (SBV) and diastolic blood viscosity (DBV) corresponding to whole blood viscosity at high and low shear rates, respectively. Samples containing 3 ml of whole blood were collected in a vacutainer containing ethylenediaminetetraacetic acid (EDTA), an anticoagulant, and stored refrigerated at 4° C. until the test. All measurements were performed within 24 hours of sample collection. Whole blood viscosity was measured over a wide range of shear rates from 1 to 1000 s ^-1 . In the low shear rate range of 5 s ^-1 or less, whole blood viscosity 1 s ^-1 was chosen as the DBV. In the high shear rate range of 300 s ^-1 or higher, a whole blood viscosity of 300 s ^-1 was chosen as SBV. The correlation between SBV and DBV, red blood cell volume and fibrinogen in whole blood viscosity was statistically analyzed as described in Example 2 above.

As a result, DBV and SBV had a significant correlation with each other (r = 0.456, p <0.001), DBV had a significant correlation with red blood cell volume (r = 0.421, p = 0.001) (Fig. 2a), SBV Was significantly correlated with the blood fibrinogen level (r = 0.340, p = 0.01) (Fig. 2d).

Example 5. Relationship between blood viscosity and stroke subtype

As a result of analyzing the relationship between the blood viscosity analyzed in Example 3 and the stroke subtype as described in Example 2, the DBV at the time of onset in SAO was 274.7 ± 70.9 mP, and the DBV at the time of onset in the stroke of unknown cause was 258.9 ± In 46.5 mP, CE, DBV at onset was 252.0 ± 57.5 mP, and in LAA, DBV at onset was 214.5 ± 75.9 mP. Specifically, DBV at the onset of SAO was significantly higher than that of other stroke subtypes (p = 0.037). In addition, in the post-hoc test, DBV at the onset of SAO was significantly higher than that of DBV at the onset of LAA (p = 0.021; Table 1 and Fig. 3a). Among laboratory variables significantly associated with SAO stroke, such as DBV, hemoglobin, and red blood cell volume in univariate analysis, only DBV is a number of vascular risk factors, including age, sex, hypertension, diabetes, hyperlipidemia, atrial fibrillation and smoking. The multivariate analysis applied to was still significant (p = 0.033) (Table 2). The DBV in the stroke-like group was 240.8 ± 38.1 mP. Test findings affecting blood viscosity did not differ among stroke subtypes except for increased D-dimer levels in CE (Table 1). SBV, red blood cell volume, blood fibrinogen levels and BUN/Cr ratio at the time of onset did not differ between stroke subtypes (FIGS. 3b to e).

Example 6. Confirmation of blood viscosity and chronic radiographic signs of cerebral small vessel disease

Correlation between major determinants of blood viscosity, such as DBV, SBV, erythrocyte volume and fibrinogen level, and chronic radiological signs such as white matter hyperplasia, hiatus cerebral infarction and microbleeding in all patients, regardless of stroke subtype. Was investigated.

As a result, a statistically significant correlation was observed between DBV and the number of chronic hiatus brain attack (r = 0.274, p = 0.030) (Fig. 4a). DBV appeared to have a positive correlation with the severity of PVH (r = 0.135, p = 0.291) or the number of microbleeds (r = 0.145, p = 0.256), but not statistically significant (Fig. 4a). SBV, red blood cell volume, and fibrinogen levels in blood were not shown to be significantly correlated with radiologic signs (FIGS. 4b to d ).

Example 7. Confirmation of blood viscosity characteristics over time

Follow-up blood viscosity was measured in 41 of 63 patients whose stroke subtype was determined (65.1%). The demographic characteristics of patients who were measured and who did not differ significantly (Table 3).

To confirm the role of blood viscosity in SAO stroke, patients were dichotomized into SAO and non-SAO groups (other subtypes). Blood viscosity was measured at the time of onset, 1 week, and 5 weeks (subsequent). At the time of onset, DBV was significantly higher in the SAO group than in the non-SAO group (274.7 ± 70.9 mP and 229.8 ± 70.0 mP, p = 0.016, respectively) (Fig. 5A). The difference between the DBV of the two groups decreased after 1 week (SAO, 232.7 ± 54.9 mP; non-SAO, 216.2 ± 48.0 mP; p = 0.394), and increased again after 5 weeks (SAO, 273.2). ± 48.9 mP; non-SAO, 218.6 ± 65.4 mP; p=0.009) (FIG. 5A ). In the SAO group, DBV increased again and returned to baseline at the time of stroke onset. On the other hand, DBV in the non-SA0 group was found to remain within the same range as the value measured after 1 week. SBV measured after 5 weeks was also significantly higher in the SAO group than in the non-SAO group (42.0 ± 4.3 mP and 36.1 ± 6.9 mP, respectively; p = 0.005) (Fig. 5b).

Example 8. Arterial artery occlusion (SAO) stroke subtype-specific cut-off value determination

After dividing into SAO and non-SAO (nSAO) using the blood viscosity values of the patients whose stroke subtype was determined in Example 7, diastolic blood viscosity (DBV) and systolic blood (SBV) were analyzed through ROC analysis. viscosity, systolic blood viscosity) The cut-off values with optimal sensitivity and specificity for each measurement period were derived, measured at the time of onset, 1 week, and 5 weeks after each onset. Specifically, the derived DBV at the time of onset was Cut-off>296.1mP, p=0.0155, sensitivity=48%, specificity=84%, DBV after 1 week cut-off>245.9mP, p=0.3393, sensitivity=63 %, specificity=79% and after 5 weeks, DBV was cut-off>225.2mP, p=0.0049, sensitivity=87%, specificity=55% (Figs. 6a to c), and SBV at the time of onset was cut -off>37mP, p=0.0170, sensitivity=88%, specificity=45%, SBV after 1 week Cut-off>39mP, p=0.5912, sensitivity=55%, specificity=74% and SBV after 5 weeks Was found to be Cut-off>34.7mP, p=0.0019, sensitivity=100%, specificity=41% (Figs. 7a to c).

That is, the cut-off value of the diastolic/systolic phase and the blood viscosity for each period that determines the arterial obstruction (SAO) based on the capillary blood viscosity measurement was >296.1mp (p<0.05); Systolic blood viscosity at onset >37mp (p<0.05); Diastolic blood viscosity >225.2mp (p<0.01) after 5 weeks; And systolic blood viscosity >34.7mp (p<0.01) after 5 weeks.

Claims (12)

In the method of providing information for diagnosis or prognosis of a stroke using a medical diagnosis device,
(a) Diastolic blood viscosity (DBV) and systolic blood viscosity using a diastolic blood, systolic blood and a scanning capillary tube viscometer isolated from the subject at the time of the invention or 5 weeks after the onset of the stroke. measuring (systolic blood viscosity, SBV); And
(b) the diastolic blood viscosity at the time of onset is greater than 296.1 mp and the systolic blood viscosity is greater than 37 mp; Or if the diastolic blood viscosity exceeds 225.2mp and the systolic blood viscosity exceeds 34.7mp 5 weeks after the onset, determining that there is a risk of small artery occlusion (SAO); small artery occlusion comprising , SAO), a method of providing information for the diagnosis or prognosis of stroke.
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