CN103901214A - Reagent for predicting micro-bleeding of acute cerebral infarction patient - Google Patents
Reagent for predicting micro-bleeding of acute cerebral infarction patient Download PDFInfo
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Abstract
The invention belongs to the field of biological detection, and relates to a reagent for predicting micro-bleeding of an acute cerebral infarction patient. The reagent consists of reagents for detecting any one or more of adiponectin, E-selectin, S100B proteins and soluble advanced glycation end product receptors sRAGE. The content of the adiponectin, the E-selectin, the S100B proteins and the soluble advanced glycation end product receptors in the venous blood of the acute cerebral infarction patient is detected, so that the occurrence possibility of micro-bleeding of the patient is judged in an early stage, and a basis is provided for early intervention.
Description
Technical field
The invention belongs to field of biological detection, relate to a kind of micro-hemorrhage reagent of acute cerebral infarction patients generation of predicting.
Background technology
Brain micro-hemorrhage be in brain due to Microangiopathy take the hemorrhage subclinical infringement of brain essence as principal feature of small quantity.On gtadient echo T2 weighted imaging, micro-hemorrhage one or more mottled signal deletion kitchen ranges that show as, diameter 2-5mm, around without oedema.Histopathology confirms, signal deletion is mainly because the arteriole Blood Trace of fiber hyaloid degeneration exosmoses, and causes due to the deposition of iron content protoheme.After research shows brain micro-hemorrhage and age, Analysis on Cardiovascular Risk Factors, myelinopathy, cerebral apoplexy, cerebral amyloid angiopathy change, cognition dysfunction, palsy, the disturbance of emotion has certain relation, and after brain micro-hemorrhage and Ischemic Stroke, cerebral hemorrhage and thrombolysis or the treatment of anti-bolt, symptomatic cerebral hemorrhage is in close relations.
The micro-hemorrhage risk that likely increases hemorrhage conversion after acute cerebral infarction thrombolysis of early stage perspective study prompting brain.After some researchs recently show brain micro-hemorrhage and thrombolysis there is not correlativity in hemorrhage conversion.Kim etc. in row vein tPA thrombolysis or the 6 hours of onset expert artery urokinase thrombolytic therapy in 3 hours of falling ill, studies show that the micro-hemorrhage number of brain is not the independent hazard factor of hemorrhage conversion after Hyperacute cerebral infarction thrombolysis to acute cerebral infarction patients.But it is only 5 examples that the micro-hemorrhage number of this test midbrain is greater than 10 patient, may have impact to result.Fiehler etc. are to giving studies show that of patient of intravenous thrombolysis therapy in 570 routine 6 hours of onsets, it is micro-hemorrhage that 86 routine patients baseline period before thrombolysis finds that there is brain, and wherein the micro-hemorrhage number of 6 routine patients' brain is greater than 5.In this 86 routine patient, there are 5 examples (5.8%) that hemorrhage conversion occurs, in the micro-hemorrhage patient of anencephaly, have 13 examples (2.7%) to occur hemorrhage conversion, difference not statistically significant.This research points out, compared with the yield (≈ 10%) of the thromboembolism treatment of the past bibliographical information, even if the risk that hemorrhage conversion occurs the micro-hemorrhage patient of brain slightly raises, also can not exceed the benefit that thrombolysis brings; Simultaneously fewer in number owing to having the micro-hemorrhage patient of multiple brains, the relation of the micro-hemorrhage number of brain and hemorrhage conversion still needs further to be confirmed.This research is the retrospective study of the multicenter cooperation of maximum at present, but this research is at still Shortcomings of aspect such as the selection of the micro-hemorrhage assessment of brain, statistical method and statistics effect, evaluations to the micro-hemorrhage latent effect of the brain of different parts, and its conclusion still awaits further checking.For the anti-bolt treatment of post-stroke, the hemorrhage conversion after micro-hemorrhage may the treatment with anti-bolt of brain is relevant in current studies show that, but its risk-benefit ratio is still not clear at present.On this basis, micro-hemorrhage existence can provide guidance to dispel poly-, anti-freezing and thrombolytic drug of Ischemic Stroke choice for use.
Adiponectin (adiponectin) is a kind of endogenous biologically active polypeptide of being secreted by adipocyte.Be present in a large number in blood circulation.Adiponectin is known as again Acrp30, apM1, AdipoQ, GBP28.Adiponectin can affect fat and sugar class metabolism, regulate blood vessel inner skin cell function, anti-inflammatory, in the generation of cerebrovascular disease and development, play important nerve and vascular protection effect.
E-Selectin (E-selectin) is a member of selecting plain family, is the cell adhesion molecule that a class Ca2+ relies on, and can identify and combination with special glycosyl.Main identification and the adhesion participating between leucocyte and vascular endothelial cell.E-Selectin is present in the Surface of Vascular Endothelial Cells of activation, claims again Adhesion Molecules on Endothelial Cells-1.Micro-hemorrhage generation may cause the permeability of blood-brain barrier (BBB) relevant with vascular endothelial dysfunction, once endothelial injuries will cause BBB permeability to increase, blood constituent exosmoses, and occurs petechial hemorrhage, and along with the endothelial cell quantity increase activating, expression increases E-Selectin.
S100B is a member of S100 protein family, is the specific albumen of Deiter's cells, mainly expresses at astroglia, especially the astroglia of those parcel vascular endothelial cells.When damage occurs nervous system, S100B is released into blood vessel external series gap from damaging cells, enters blood by BBB, causes the S100B level in blood to increase.
Solubility Advanced Glycation End Product Receptors (sRAGE) is the soluble component of Advanced Glycation End Product Receptors (RAGE).RAGE is the signal conduction acceptor of advanced glycation end products, is a kind of multiple ligand acceptor in cell surface immunoglobulin superfamily, at many cells.The endogenic ligand of its sugar-free comprises proinflammatory molecule S100B, HMGB-1, amyloid polypeptide and b2 integral protein macrophage antigenic complex-1 etc.SRAGE has the similar structure of part to RAGE, can with its ligand binding, the competitive activity that suppresses RAGE.Therefore, can avoid the damaging reaction that RAGE-part axle mediates by Cell protection.In addition, sRAGE also can be used as the scavenger receptor of AGEs and other RAGE parts in circulation.
How to predict whether Patients with Cerebral Infarction occurs micro-hemorrhage, there is no at present definite method, only have some clinical score scales, but accuracy and specificity are all not high, and not yet in population of China, verify its validity.There is no at present can be for the biochemistry detection means of the hemorrhage generation of pre-micrometer.
Summary of the invention
The object of the invention is the above-mentioned deficiency for prior art, provide a kind of for predicting that micro-hemorrhage reagent occurs acute cerebral infarction patients.
Object of the present invention can be achieved through the following technical solutions:
A kind of for predicting that micro-hemorrhage reagent occurs acute cerebral infarction patients, by detecting any one in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE, or multiple reagent composition, preferably by detecting any one in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE, or multiple ELISA reagent composition.
The ELISA reagent that detects adiponectin comprises HRP compound, the benzidine reaction substrate etc. of the human adiponectin specific antibody of biotin coupling, human adiponectin protein standard substance, Streptavidin coupling.
The ELISA reagent that detects E-Selectin comprises HRP compound, the benzidine reaction substrate etc. of people's E-Selectin specific antibody of biotin coupling, people's E-Selectin protein standard substance, Streptavidin coupling.
The ELISA reagent that detects S100B comprises HRP compound, the benzidine reaction substrate etc. of the people S100B specific antibody of biotin coupling, people's S100B standard items, Streptavidin coupling.
The ELISA reagent that detects sRAGE comprises HRP compound, the benzidine reaction substrate etc. of the people sRAGE specific antibody of biotin coupling, people sRAGE protein standard substance, Streptavidin coupling.
Above-mentioned ELISA reagent can be made by oneself, also can buy commercial corresponding antibodies and ELISA kit.
Any one in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE or multiple application in the micro-hemorrhage reagent of preparation prediction acute cerebral infarction patients generation.
There is micro-hemorrhage combinations of biomolecules in prediction acute cerebral infarction patients, by any two kinds or two or more composition in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE.
Beneficial effect:
The micro-hemorrhage significant correlativity that has of the content of adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors in Patients with Cerebral Infarction venous blood and brain, detection reagent provided by the invention, to the micro-hemorrhage good predictive value that has of brain, is the micro-hemorrhage foundation that provides of early intervention brain.
Embodiment
Embodiment
1 data and method
1.1 research object
Research object is starting Ischemic Stroke of being in hospital in Nanjing General Hospital, Nanjing Military Area Command, PLA during year August in January, 2012 to 2012, and the same period because of headache, dizzy health examination volunteer is as control group, all completes MRI(3.0T) magnetic susceptibility sequence (SWI) scanning.Neurological functional deficit adopts NIH stroke scale (NIHSS) scoring by two Neurology doctors that do not know imaging examination result.Enter group standard as follows :≤8 points of (1) NIHSS scorings; (2) be admitted to hospital in 1 week in morbidity; (3) check contraindication (for example having magnetic metal material in body) without MRI.Exclusion standard is as follows: (1) kidney or liver diseases; (2) there is acute or chronic inflammation disease, as pneumonia, meningitis etc., use recently any antiphlogistic; (3) malignant tumour or cerebral operations.Collect all experimenters' Baseline Data, as sex, age, smoking history, history of drinking history, past medical history and medication history.
1.2MRI inspection method
MRI checks the 3.0T MRI of the Siemens company machine that adopts, the conventional sequence of row spin echo, three-dimensional time leap blood vessel imaging and GER-T2*WI and SWI sequence scanning.SWI sweep parameter is: matrix 298x448; Inspection area 230mm × 184mm, bed thickness 1.5mm, repetition time, echo time are respectively 28ms and 20ms, encourage 1 time.Micro-hemorrhage low signal disappearance that is defined as diameter 2-10mm, clear border, around without oedema.Get rid of the empty shadow of globus pallidus calcification, cavum subarachnoidale hypointense signal and vessel flow etc.Ischemic Stroke is micro-ly hemorrhagely divided into two groups according to having or not.Micro-be hemorrhagely divided under deep, cerebral lobe and curtain according to position.And according to micro-hemorrhage quantity classification: without micro-hemorrhage, slight (micro-hemorrhage total 1 to 5), moderate (6 to 15) and severe (being greater than 15).Above results of imaging is analyzed the unwitting neuroimaging doctor of section of patient clinical data by 2.
1.3ELISA method detects adiponectin, E-Selectin, S100B and sRAGE
In-patient morning next day gathers patient's median basilic vein blood 2ml on an empty stomach, and control group be outpatient service on an empty stomach blood sampling be all placed in EDTA anticoagulant tube, 1500 revs/min after centrifugal 10 minutes, separated plasma, is placed in-80 ℃ of refrigerator storages for subsequent use.Adiponectin, E-Selectin, S100B and sRAGE detection method are basic identical, adiponectin detection kit is purchased from Millipore company of the U.S., E-Selectin kit is purchased from eBioscience company, S100B detection kit is purchased from BioVendor company, sRAGE detection kit, purchased from R & D company, all operates by kit instructions flow process.Illustrate as an example of S100B example below.
1. application of sample: by the standard items of variable concentrations (50pg/ml, 100pg/ml, 200pg/ml, 500pg/ml, 1000pg/ml, 2000pg/ml), dilution buffer liquid (blank), serum sample to be checked adds in ELISA Plate with the amount in 100 microlitres/hole, and 2 multiple holes are set.Cover shrouding film, room temperature (25 ℃) is hatched 2 hours.
2. wash plate: button removes liquid in hole, and 300 microlitres/hole adds eluent; Stop and discard liquid in hole after 1 minute.Repeat 3 times, on filter paper, buckle for the last time dry.
3. add detection antibody: 100 microlitres/hole adds the primary antibodie of the biotin coupling after dilution.Cover shrouding film, room temperature (25 ℃) is hatched 1-1.5 hour.
4. wash plate: button removes liquid in hole, and 300 microlitres/hole adds eluent; Stop and discard liquid in hole after 1 minute.Repeat 3 times, on filter paper, buckle for the last time dry.
5. enzyme-added: 100 microlitres/hole adds Streptavidin-HRP.Cover shrouding film, incubated at room 30 minutes.
6. wash plate: button removes liquid in hole, and 300 microlitres/hole adds eluent; Stop and discard liquid in hole after 1 minute.Repeat 3 times, on filter paper, buckle for the last time dry.
7. colour developing: 100 microlitres/hole adds TMB, and room temperature lucifuge is hatched 5-30 minute.Judge cessation reaction according to the depth of color in hole (mazarine).Conventionally colour developing can reach good effect in 10-20 minute.
8. cessation reaction: rapid 100 microlitres/hole adds stop buffer cessation reaction.
9. read plate: stop in latter 10 minutes, with detecting the wavelength 450nm value of reading, draw the OD value in every hole with microplate reader.
2 statistical analysis
Application SPSS16.0 software carries out statistical study, normal distribution measurement data is with mean ± standard deviation (x ± s) represent, adopt independent sample t check or one-way analysis of variance, skewed distribution measurement data is relatively with Kruskal-Wallis or Mann – Whitney U check, and enumeration data is relatively used x
2check or Fisher check.The factor relevant with micro-hemorrhage order of severity carried out to P < 0.2 variable row grade Logistic regretional analysis again after single factor analysis.In addition, adopt partial Correlation Analysis in micro-bleeding group, and proofread and correct possible Confounding Factor, as the age, sex, the use of statins etc.P < 0.05 has statistical significance for difference.
3 results
3.1 patients' general demography data
Have 170 routine patients and be included into this research, 63.9 years old mean age, wherein the male sex's 109 examples, account for 64.1%.81 people's Finding cases occur micro-hemorrhage, account for 47.6%.Wherein micro-hemorrhage 27 examples of single-shot, account for 33.3%, and multiple micro-hemorrhage 54 examples, account for 66.7%.Patient's medical history situation and main laboratory examination situation are in table 1.
3.2E-selects element and micro-hemorrhage correlation analysis
Serum levels of E-selectin when the micro-hemorrhage patient of while in hospital generation is admitted to hospital is apparently higher than the serum levels of E-selectin numerical value that micro-hemorrhage patient does not occur.Logistic regretional analysis show, proofreading and correct after blood pressure, HDL and fibrinogen, E-Selectin still with micro-hemorrhage positive correlation (OR:1.12,95%CI:1.03-1.17).If take 12 as truncation points, diagnostic sensitivity is 60%, and specificity is 74%.
3.3 adiponectins and micro-hemorrhage correlation analysis
Serum adiponectin when the micro-hemorrhage patient of while in hospital generation is admitted to hospital is starkly lower than the Serum adiponectin numerical value that micro-hemorrhage patient does not occur.Logistic regretional analysis show, proofreading and correct after blood pressure, HDL and fibrinogen, adiponectin still with micro-hemorrhage negative correlation (OR:0.54,95%CI:0.36-0.82).If take 300 as truncation points, diagnostic sensitivity is 72%, and specificity is 83%.
3.4S100B with micro-hemorrhage correlation analysis
Serum S100B when the micro-hemorrhage patient of while in hospital generation is admitted to hospital is apparently higher than the S100B numerical value that micro-hemorrhage patient does not occur.Logistic regretional analysis show, proofreading and correct after blood pressure, HDL and fibrinogen, S100B still with micro-hemorrhage positive correlation (OR:2.1,95%CI:1.8-4.3).If take 100 as truncation points, diagnostic sensitivity is 85%, and specificity is 94%.
3.5sRAGE and micro-hemorrhage correlation analysis
Serum sRAGE when the micro-hemorrhage patient of while in hospital generation is admitted to hospital is starkly lower than the serum sRAGE numerical value that micro-hemorrhage patient does not occur.Logistic regretional analysis show, proofreading and correct after blood pressure, HDL and fibrinogen, sRAGE still with micro-hemorrhage negative correlation (OR:0.32,95%CI:0.24-0.48).If take 900 as truncation points, diagnostic sensitivity is 92%, and specificity is 88%.
3.6 2 kinds of couplings detect index and micro-hemorrhage correlation analysis
Each index truncation points is the same.
If coupling adiponectin+E-Selectin, diagnostic sensitivity is 79%, and specificity is 60%.
If coupling adiponectin+S100B, diagnostic sensitivity is 88%, and specificity is 74%.
If coupling adiponectin+sRAGE, diagnostic sensitivity is 96%, and specificity is 68%.
If coupling E-Selectin+S100B, diagnostic sensitivity is 89%, and specificity is 54%.
If coupling E-Selectin+sRAGE, diagnostic sensitivity is 94%, and specificity is 62%.
If coupling S100B+sRAGE, diagnostic sensitivity is 97%, and specificity is 69%.
3.7 3 kinds of couplings detect index and micro-hemorrhage correlation analysis
Each index truncation points is the same.
If coupling adiponectin+E-Selectin+S100B, diagnostic sensitivity is 95%, and specificity is 45%.
If coupling adiponectin+E-Selectin+sRAGE, diagnostic sensitivity is 97%, and specificity is 51%.
If coupling adiponectin+S100B+sRAGE, diagnostic sensitivity is 96%, and specificity is 63%.
If coupling E-Selectin+S100B+sRAGE, diagnostic sensitivity is 96%, and specificity is 79%.
3.8 4 kinds of couplings detect index and micro-hemorrhage correlation analysis
Each index truncation points is the same.
If coupling adiponectin+E-Selectin+S100B+sRAGE, diagnostic sensitivity is 98%, and specificity is 39%.
The basic clinical data comparison of table 1. patient
* with do not send out bleeding group comparison micro-, P<0.05.
Claims (5)
1. for predicting that a micro-hemorrhage reagent occurs acute cerebral infarction patients, it is characterized in that by detecting any one in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE, or multiple reagent composition.
2. according to claim 1 for predicting that micro-hemorrhage reagent occurs acute cerebral infarction patients, it is characterized in that by detecting any one in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE, or multiple ELISA reagent composition.
3. according to claim 2 for predicting that micro-hemorrhage reagent occurs acute cerebral infarction patients, the ELISA reagent that it is characterized in that detecting adiponectin comprises human adiponectin specific antibody, human adiponectin protein standard substance, the HRP compound of Streptavidin coupling, the benzidine reaction substrate of biotin coupling; The ELISA reagent that detects E-Selectin comprises people's E-Selectin specific antibody, people's E-Selectin protein standard substance, the HRP compound of Streptavidin coupling, the benzidine reaction substrate of biotin coupling; The ELISA reagent that detects S100B comprises people S100B specific antibody, people's S100B standard items, the HRP compound of Streptavidin coupling, the benzidine reaction substrate of biotin coupling; The ELISA reagent that detects sRAGE comprises people sRAGE specific antibody, people sRAGE protein standard substance, the HRP compound of Streptavidin coupling, the benzidine reaction substrate of biotin coupling.
4. any one in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE or multiple application in the micro-hemorrhage reagent of preparation prediction acute cerebral infarction patients generation.
5. there is micro-hemorrhage combinations of biomolecules in prediction acute cerebral infarction patients, it is characterized in that by any two kinds or two or more composition in adiponectin, E-Selectin, S100B and solubility Advanced Glycation End Product Receptors sRAGE.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN113433254A (en) * | 2021-08-27 | 2021-09-24 | 宝枫生物科技（北京）有限公司 | Biomarker for diagnosing cerebral infarction of patient with leukoencephalopathy and application of biomarker |
| CN113447600A (en) * | 2021-08-27 | 2021-09-28 | 宝枫生物科技（北京）有限公司 | Biomarker for diagnosing cerebral infarction of patient with leukoencephalopathy and application of biomarker |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113433254A (en) * | 2021-08-27 | 2021-09-24 | 宝枫生物科技（北京）有限公司 | Biomarker for diagnosing cerebral infarction of patient with leukoencephalopathy and application of biomarker |
| CN113447600A (en) * | 2021-08-27 | 2021-09-28 | 宝枫生物科技（北京）有限公司 | Biomarker for diagnosing cerebral infarction of patient with leukoencephalopathy and application of biomarker |
| CN113447600B (en) * | 2021-08-27 | 2021-11-02 | 宝枫生物科技（北京）有限公司 | Biomarker for diagnosing cerebral infarction of patient with leukoencephalopathy and application of biomarker |
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