CN109061137A - Purposes of 13 (the S)-HODE levels as cardiovascular disease Testing index - Google Patents

Purposes of 13 (the S)-HODE levels as cardiovascular disease Testing index Download PDF

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Publication number
CN109061137A
CN109061137A CN201811029161.8A CN201811029161A CN109061137A CN 109061137 A CN109061137 A CN 109061137A CN 201811029161 A CN201811029161 A CN 201811029161A CN 109061137 A CN109061137 A CN 109061137A
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hode
myocardial infarction
acute myocardial
purposes
index
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夏勇
罗素新
皮强中
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Chongqing Medical University
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Chongqing Medical University
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/531—Production of immunochemical test materials
    • G01N33/532—Production of labelled immunochemicals
    • G01N33/535—Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6854—Immunoglobulins
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/32—Cardiovascular disorders
    • G01N2800/324—Coronary artery diseases, e.g. angina pectoris, myocardial infarction

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Abstract

A kind of purposes the invention discloses 13 (S)-HODE levels as cardiovascular disease Testing index, belong to field of biomedicine technology, specifically, the reagent for providing detection people's isolated blood 13 (S)-HODE level is preparing the purposes in cardiovascular disease diagnosis reagent, and index of the measurement from 13 (S)-HODE levels in vitro human plasma is used for the purposes in prognosis evaluation cardiovascular disease kit in preparation.The present invention is experimentally confirmed, and (the S)-HODE level of peripheral blood 13 is chosen in close relations with prognosis after acute myocardial infarction and therapeutic scheme.

Description

Purposes of 13 (the S)-HODE levels as cardiovascular disease Testing index
Technical field
The invention belongs to field of biomedicine technology, are related to 13 (S)-HODE levels of one kind and refer to as cardiovascular disease detection Target purposes.
Background technique
Acute myocardial infarction AMI is one kind of acute coronary syndrome, is the emergency and severe disease with high lethality rate.The hair of screening as early as possible Existing acute myocardial infarction AMI, for rescue, patient vitals are most important.At present in acute myocardial infarction AMI diagnosis and treatment, coronarography It is the goldstandard of its diagnosis, in addition there are electrocardiogram, myocardial enzymes detection, optical coherence tomography and intravascular ultrasound inspections The diagnostic means such as look into.But coronarography is a kind of invasive inspection, and has one to the technology of Medical Devices and healthcare givers Provisioning request.Myocardial enzymes usually obviously rise after acute myocardial infarction AMI 3-4h, for Early acute myocardial infarction, especially Acute myocardial infarction AMI within morbidity 3-4h, can not efficient diagnosis and antidiastole.At present these means be all unable to satisfy it is objective, Easy identification myocardial infarction, the especially demand of screening early myocardial infarction.How effectively to identify that acute myocardial infarction AMI is outstanding It is that early myocardial infarction is current urgent problem to be solved.Wherein coronary plaque does not rupture the acute of (fibrous cap is complete) The treatment of myocardial infarction may not necessarily be implanted into bracket, and clinical prognosis is more preferable, and the probability of Cardia cevent is also lower (Prati F, Uemura S, Souteyrand G et al. OCT-based diagnosis and management of STEMI associated with intact fibrous cap. JACC: Cardiovascular Imaging 2013; 6:283-287.).Although optical coherence tomography and intravascular ultrasound can show the shape of patient's coronary artery and its patch State, but relevant device expensive equipment, technical requirements are high.Therefore effectively easily identify the method for coronary plaque rupture very It is important.
In the prior art, there are no effective method come it is objective, easily identify Early acute myocardial infarction (morbidity 3-4h Within), myocardial infarction coronary plaque the technical issues of whether rupturing.
Summary of the invention
The present invention provides purposes of 13 (the S)-HODE levels of one kind as cardiovascular disease Testing index.This purposes Solve in the prior art without effective method come it is objective, easily identify Early acute myocardial infarction (morbidity 3-4h with It is interior), myocardial infarction coronary plaque the technical issues of whether rupturing.
Its technical solution is as follows:
The present invention provides one kind to derive from purposes of (the S)-HODE level of people's isolated blood 13 as Testing index.
The present invention also provides the reagents of detection people's isolated blood 13 (S)-HODE level to prepare cardiovascular disease diagnosis Purposes in reagent.
Further, the cardiovascular disease includes that Early acute myocardial infarction, acute myocardial infarction AMI, coronary plaque are broken It splits.
Further, 13 (S)-HODE levels of measurement are compared with scheduled index 540ng/ml, if being higher than 540ng/ml, It is diagnosed as Early acute myocardial infarction (within morbidity 3-4h).
The present invention also provides 13 (S)-HODE, and as Testing index, in preparation, for Early acute myocardial infarction, (fall ill 3- Within 4h) purposes in screening agent.
The present invention also provides be used for the prognosis of acute myocardial infarction AMI using (the S)-HODE level of human peripheral 13 as index Assessment.
The present invention also provides using (the S)-HODE level of human peripheral 13 as index in controlling for acute myocardial infarction AMI Treat the assessment of scheme selection.
The present invention also provides a kind of indexs of measurement (S)-HODE level 13 in vitro human plasma to use in preparation Purposes in prognosis evaluation cardiovascular disease kit.
Further, the cardiovascular disease includes that Early acute myocardial infarction, acute myocardial infarction AMI, coronary plaque are broken It splits.
Further, 13 (S)-HODE levels of measurement are compared with scheduled index 540ng/ml, if being higher than 540ng/ml, It is diagnosed as Early acute myocardial infarction (within morbidity 3-4h).
Beneficial effects of the present invention:
13 (S)-HODE levels of the invention can effectively identify acute myocardial infarction AMI as the purposes of cardiovascular disease Testing index, Especially early myocardial infarction (within morbidity 3-4h), and 13 (S)-HODE and plaque rupture are closely related, are suitable for identifying Coronary plaque rupture determines the prognosis of myocardial infarction patient.The present invention measures people by competitiveness enzyme-linked immunoabsorption 13 (S)-HODE in Blood plasma in vitro are horizontal, it is found that 13 (S)-HODE levels are compared with just in patients of acute myocardial infarction (AMI) blood plasma 13 (S)-HODE raising in the blood plasma of normal coronary artery group (NCA) and unstability blood patient with angina pectoris group (UA) (AMI: 554.18 ± 52.91ng/ml, NCA:325.07 ± 45.89ng/ml, UA:332.30 ± 28.013ng/ml, single factor test variance point Analysis, P < 0.01).And Early acute myocardial infarction (within morbidity 3-4h, AMI (onset within 3-4h)) patients blood plasma In 13 (S)-HODE horizontal (761.67 ± 125.41ng/ml) more compared with 13 (S)-HODE levels in NCA and UA patients blood plasma High (one-way analysis of variance, P < 0.01).And the ROC curve by the patient for calculating AMI patient and exclusion AMI, finds it Youden index is that 0.265, corresponding 13 (S)-HODE levels are 540ng/ml, can be used as diagnosis acute myocardial infarction AMI Index is 540ng/ml(p < 0.01).Early acute myocardial infarction (AMI (the onset being further discovered that within morbidity 3-4 h Within 3-4h)) patient and exclude acute myocardial infarction AMI patient ROC curve, Youden index be 0.265, it is corresponding 13 (S)-HODE levels be also 540ng/ml, can be used as diagnosis Early acute myocardial infarction screening index be 540ng/ml (p < 0.01).
Detailed description of the invention
Figure 1A is by the detection discovery of competitiveness enzyme-linked immuno analytical method in acute myocardial infarction AMI (AMI) peripheral blood in patients In the blood plasma of acquirement, 13 (S)-HODE levels less Stable Angina Pectoris and normal person increase (one-way analysis of variance, P < 0.01), wherein NCA: normal group of coronary artery, UA: unstable angina pectoris group, AMI: acute myocardial infarction AMI group, * * P < 0.01；
Figure 1B is the ROC curve of the patient of AMI patient and exclusion AMI, and Youden index is 0.265, corresponding 13 (S)- HODE level is 540ng/ml, and the index that can be used as diagnosis acute myocardial infarction AMI is 540ng/ml(p < 0.01).
Fig. 2A is the early stage Acute myocardial by the detection discovery of competitiveness enzyme-linked immuno analytical method within the 3-4h that falls ill In the blood plasma that infarct (AMI (onset within 3-4h)) patient's human peripheral obtains, 13 (S)-HODE levels are more unstable Property patient with angina pectoris and normal person increase (one-way analysis of variance, P < 0.01), wherein: NCA: normal group of coronary artery, UA: Unstable angina pectoris group, AMI (onset within 3-4h): Early acute myocardial infarction group (within morbidity 3-4h), * * P <0.01；
Fig. 2 B is the ROC of the patient of the acute myocardial infarction AMI (AMI) patient and exclusion acute myocardial infarction AMI within the 3-4h that falls ill Curve, Youden index are that 0.265, corresponding 13 (S)-HODE levels are 540ng/ml, and it is anxious to can be used as screening early stage Property myocardial infarction index be 540ng/ml(p < 0.01)
Fig. 3 is to carry out immunohistochemical staining for human coronary artery to find that the expression of 13 (S)-HODE in its tissue is mainly enriched in Patch coronarius.Show that (S)-HODE is that the coronary plaque of rupture discharges 13 in patients of acute myocardial infarction blood plasma.
Specific embodiment
Technical solution of the present invention is described in more detail with reference to the accompanying drawings and detailed description.
Experimental material used in following embodiment and reagent are as follows:
13 (S)-HODE competitiveness enzyme linked immune assay kits are purchased from U.S. Enzo Life Sciences company；
13 (S)-HODE IgG primary antibody goat derived antibodies are purchased from U.S. USBiological company；
HRP marks two antiantibody of rabbit-anti goat to be purchased from Wuhan Seville company；
3% hydrogen peroxide is purchased from Wuhan Seville company；
Normal rabbit serum is purchased from Wuhan Seville company；
Groupization kit DAB color developing agent is purchased from Wuhan Seville company.
Embodiment 1
13 (S)-HODE levels are in normal coronary group (NCA) peripheral blood, unstable angina group (UA) and Acute myocardial Detection in the peripheral blood of Infarction Patients (AMI).
The present embodiment selects normal coronary group (NCA) peripheral blood of the same symptom of cardiac muscle, unstable angina group (UA) and the human peripheral of patients of acute myocardial infarction (AMI).It is detected in blood plasma by competitiveness enzyme linked immune assay kit 13 (S)-HODE are horizontal.
Specific implementation method are as follows: competitiveness enzyme linked immune analytic method
The peripheral blood 3000rpm centrifugation 10min of acquirement is obtained into blood plasma；Prepare 2000 ng/ml, 1000 ng/ml, 250 ng/ The concentration gradient dilution of 13 (S)-HODE standard items of ml, 62.5 ng/ml, 15.6 ng/ml, 3.9 ng/ml；50 ul 13 (S) the standard detection liquid dilution of 450 ul is added in-HODE Conjugate, is used for gross activity degree detection hole；In each detection hole It is interior that blood plasma, 13 (S)-HODE gradient solutions, 13 (the S)-HODE detected accordingly are added by step according to kit specification Conjugate stoste, standard detection liquid, antibody stoste；Incubation at room temperature, 3-4h；It is rinsed 3 times with the rinsing liquid of preparation；It lives always Property degree detection hole in 13 (S)-HODE Conjugate blue liquids of 1:10 are added；It is added in kit and provides in each detection hole PNpp substrate solution, 200 ul are incubated for 3-4h under the conditions of 37 DEG C；Offer in 50 ul kits is provided in each detection hole React stop solution；The absorbance of each detection hole is detected under 405 nm luminosity in microplate reader immediately；With four parameter Logistic The method of regression curve fitting, establishes standard concentration curve；It is horizontal to calculate 13 (S)-HODE.
As shown in Figure 1A, (the S)-HODE level of AMI group 13 increases (P < 0.01) compared with NCA group or UA group.
As shown in Figure 1B, 13 (S)-HODE level in AMI and non-AMI group (UA group and NCA group) blood plasma is assessed by SPSS ROC curve discovery, Youden index (sensitivity+specificity -1) be 0.265, corresponding 13 (S)-HODE levels be 540ng/ml, the index that can be used as diagnosis acute myocardial infarction AMI is 540ng/ml(p < 0.05).
As shown in Figure 2 A, early stage AMI(AMI (onset within 3-4h)) (the S)-HODE level of patient 13 is compared with NCA group Or UA group increases (P < 0.01).
As shown in Figure 2 B, early stage AMI group (within 2 h of morbidity) and non-AMI group (UA group and NCA group) blood are assessed by SPSS The ROC curve discovery of (S)-HODE level 13 in slurry, Youden index (sensitivity+specificity -1) is 0.350, corresponding 13 (S)-HODE levels are 540ng/ml, and the index that can be used as screening Early acute myocardial infarction (within 2 h of morbidity) is 540ng/ml(p < 0.05).
Implementation column 2
The present embodiment selects human coronary artery's wax histotomy, and the expression of 13 (S)-HODE, HE are dyed by immunohistochemistry antibody Dyeing observation tissue morphology.
Specific implementation method are as follows: after obtaining paraffin tissue sections
1) immunohistochemical staining: 60 DEG C of paraffin tissue sections are used roasting piece 2 hours；Successively dimethylbenzene I, dimethylbenzene II, dehydrated alcohol I, dehydrated alcohol II, 85% alcohol, 75% alcohol, distilled water dewaxing treatment；Repairing for buffer (PH6.0) is repaired with citric acid antigen Multiple box carries out antigen retrieval in heating 10min in micro-wave oven；PBS(PH7.4 it) elutes 3 times, each 5min；3% hydrogen peroxide solution, Room temperature, which is protected from light, is incubated for 20 min；It is eluted 3 times with PBS(PH7.4), each 5min；Rabbit anteserum confining liquid after dilution, room temperature closing 30min；Primary antibody after 1:400 dilution, 37 DEG C of 1 h of incubation；PBS wash 5min, 3 times；Secondary antibody is incubated at room temperature 30min；It is added dropwise new The fresh DAB developing solution matched, microscopically observation colour developing；After haematoxylin dyeing, indigo plant is returned in differentiation；It is dehydrated mounting；It takes pictures.
2) HE dye: 60 DEG C of paraffin tissue sections roasting piece 2 hours；Successively dimethylbenzene I, dimethylbenzene II, dehydrated alcohol I, nothing Water-ethanol II, 85% alcohol, 75% alcohol, distilled water dewaxing treatment；Few drops of bush sperm covering tissues are added dropwise, dye 3 min- 5min；Break up 10 s-30 s with the acidic alcohol of 1 %；Blue liquid is returned in dropwise addition, returns indigo plant；5min is rinsed with tap water again；She is added dropwise Red colouring liquid dyes 2-5min on histotomy；Histotomy be dehydrated transparent；Mounting；It takes pictures.
As shown in figure 3,13 (S)-HODE more other vascular site expression in coronary plaque are significantly increased.Display 13 (S) the raising of-HODE in patients of acute myocardial infarction blood plasma is compared is caused by coronary artery rupture release；Show peripheral blood 13 (S)-HODE is increased in close relations with coronary artery rupture.Due to the uncracked acute myocardial infarction AMI of coronary plaque, prognosis More preferably, treatment can take thrombus to draw, anticoagulant, antiplatelet is without being implanted into bracket, therefore peripheral blood 13 (S)-HODE is horizontal It is chosen with prognosis after acute myocardial infarction and therapeutic scheme in close relations.
The foregoing is only a preferred embodiment of the present invention, the scope of the present invention is without being limited thereto, it is any to be familiar with Those skilled in the art within the technical scope of the present disclosure, can be readily available the simple change of technical solution, Or equivalence replacement, these change or replacement are fallen within the protection scope of the present invention.

Claims (10)

1. one kind derives from purposes of (the S)-HODE level of people's isolated blood 13 as Testing index.
2. the reagent of detection people's isolated blood 13 (S)-HODE level is preparing the purposes in cardiovascular disease diagnosis reagent.
3. purposes according to claim 2, which is characterized in that the cardiovascular disease include Early acute myocardial infarction, Acute myocardial infarction AMI, coronary plaque rupture.
4. purposes according to claim 2, which is characterized in that 13 (S)-HODE levels of measurement and scheduled index 540ng/ Ml compares, if being higher than 540ng/ml, is diagnosed as Early acute myocardial infarction.
5.13 (S)-HODE are as Testing index in preparation for the purposes in Early acute myocardial infarction screening agent.
6. assessment of (the S)-HODE level of human peripheral 13 as index for the prognosis of acute myocardial infarction AMI.
7. the assessment that (the S)-HODE level of human peripheral 13 is chosen as index in the therapeutic scheme for acute myocardial infarction AMI.
8. the index for measuring (S)-HODE level 13 in vitro human plasma is used for prognosis evaluation cardiovascular disease in preparation Purposes in kit.
9. purposes according to claim 8, which is characterized in that the cardiovascular disease include Early acute myocardial infarction, Acute myocardial infarction AMI, coronary plaque rupture.
10. purposes according to claim 8, which is characterized in that 13 (S)-HODE levels of measurement and scheduled index 540ng/ml compares, if being higher than 540ng/ml, is diagnosed as Early acute myocardial infarction.
CN201811029161.8A 2018-09-05 2018-09-05 Purposes of 13 (the S)-HODE levels as cardiovascular disease Testing index Pending CN109061137A (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7202276B2 (en) * 2000-04-07 2007-04-10 1411198 Ontario Limited Use of 13-HODE as a regulator of vascular biocompatibility and an inhibitor of cell hyperplasia
CN107271681A (en) * 2017-06-05 2017-10-20 中国人民解放军沈阳军区总医院 Applications of the blood plasma S100A12 in ST sections of elevation myocardial infarction early diagnosis

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7202276B2 (en) * 2000-04-07 2007-04-10 1411198 Ontario Limited Use of 13-HODE as a regulator of vascular biocompatibility and an inhibitor of cell hyperplasia
CN107271681A (en) * 2017-06-05 2017-10-20 中国人民解放军沈阳军区总医院 Applications of the blood plasma S100A12 in ST sections of elevation myocardial infarction early diagnosis

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
袁梦: "运用代谢组学的手段研究ω-3多不饱和脂肪酸对急性心肌梗死患者的预后的影响及相关分子机制", 《中国博士学位论文全文数据库医药卫生科技辑》 *

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Application publication date: 20181221