CN113655143A - Serum organic acid molecule prediction model for early warning of severe acute pancreatitis and application - Google Patents
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Abstract
The invention discloses a serum organic acid molecule prediction model for early warning of severe acute pancreatitis, and belongs to the technical field of biological detection. The prediction model is a composition of serum organic acid molecules, the composition is nervonic acid and glyceric acid, and the moderate-severe acute pancreatitis and the severe acute pancreatitis are distinguished within 48 hours of patients suffering from the diseases by detecting the content change of the nervonic acid and the glyceric acid in the serum. The acute pancreatitis severity prediction model has the advantages that the sensitivity of diagnosis of severe acute pancreatitis is 92.30%, the specificity is 84.70%, the false positive rate is 15.30%, the false negative rate is 7.70%, and early warning of severe acute pancreatitis is realized.
Description
Technical Field
The invention relates to a serum organic acid molecule prediction model for early warning of severe acute pancreatitis and application thereof, and belongs to the technical field of biological detection.
Background
Acute pancreatitis is a common digestive system emergency, and the onset is acute and the progress is fast. Its main pathological manifestations are pancreatic acinar cell damage, necrosis and systemic inflammatory response. The incidence rate of acute pancreatitis is as high as 13-45/10 ten thousands of people, and the incidence rate is on the rise year by year, and the high diagnosis and treatment cost brings heavy burden to patients and society. In 2012, the latest atlanta classification classified acute pancreatitis into Mild Acute Pancreatitis (MAP), Moderate Severe Acute Pancreatitis (MSAP), and Severe Acute Pancreatitis (SAP) according to the severity of the acute pancreatitis. The mild acute pancreatitis is a self-limiting disease with better prognosis, but about 20-30% of patients still have the disease continuously progress to severe acute pancreatitis, and the fatality rate is as high as 10-30%. Treatment strategies currently exist primarily with early effective fluid resuscitation and reasonable critical support. Therefore, the early warning of the disease severity degree and the selection of a reasonable treatment strategy have important clinical significance.
According to the atlanta criteria of 2012, whether patients had developed organ failure (Marshall score) or local or systemic complications after admission was differentiated between MAP and SAP, and whether organ failure exceeded 48h was differentiated between MSAP and SAP, which resulted in a 48h waiting time window for diagnosis of SAP. Early stages are primarily concerned with the presence of organ functional impairment/failure; the severity evaluation indexes such as DBC, Ranson, APACHE II, Balthazar CT grading, MCTSI, CRP, bedside severity evaluation index (BISAP), harmless score (HAPS) and the like can be simultaneously combined. However, the methods for evaluating the severity of the disease mainly depend on years of clinical practice experience accumulation of clinicians, have strong subjectivity and more collected parameters, and are not beneficial to data updating and clinical wide practice. It is worth noting that treatment in the early stage of pancreatitis significantly affects prognosis, however, existing evaluation systems all require 48 hours for accurate diagnosis, and early warning of severe acute pancreatitis is difficult to achieve. Indexes such as CRP, PTC and the like have poor specificity and only have limited reference value. Therefore, the search for new markers closely related to the severity and progression of the disease will have great significance in the early diagnosis and treatment of the severe acute pancreatitis.
Metabolomics is an important branch of system biology. The technology is a technology for characterizing, identifying and quantifying metabolites existing in the life process of an organism, and the scientific basis is that the metabolic phenotype is a direct embodiment of the biological phenotype, and the qualitative and quantitative information of the metabolites in the biological body fluid under a certain time-space node can reflect the current physiological state of the organism. Metabolomics allows the quantitative detection of specific metabolic pathways and their endogenous small molecule metabolites. The substance is qualitatively and quantitatively determined by adopting a specific sample pretreatment method, chromatographic and mass spectrum conditions and using a standard substance as a reference. Therefore, the sensitivity, specificity and repeatability of detection are greatly improved, the metabolite concentration information is ensured not to be lost, and the method has high flux, clinical operability and data mining performance. At present, researches prove that the combination of different circulating metabolites as a diagnosis or prognosis marker has good application prospect in a plurality of diseases.
Disclosure of Invention
In view of the defects of the prior art, the invention aims to provide a prediction model for realizing early warning of severe acute pancreatitis through serum organic acid molecule detection, and aims to solve the problem that no effective early warning prediction model for acute pancreatitis exists clinically at present.
The technical scheme of the invention is as follows:
the utility model provides a serum organic acid molecule prediction model of early warning severe acute pancreatitis, prediction model is the composition of serum organic acid molecule, the composition is nervonic acid and glyceric acid, and through detecting the content change of nervonic acid and glyceric acid in the serum, sick 48h in the patient, distinguish severe acute pancreatitis and severe acute pancreatitis in the middle of.
Further, in the above technical solution, the detecting content changes of nervonic acid and glyceric acid in serum is:
compared with healthy people, the expression of nervonic acid in the serum of a patient with severe acute pancreatitis is up-regulated by more than 2.8 times, and the expression of glyceric acid is up-regulated by more than 2.1 times;
compared with healthy people, the expression of nervonic acid in the serum of patients with moderate and severe acute pancreatitis is up-regulated by 1.6-2.8 times, and the expression of glyceric acid is up-regulated by 0.8-2.1 times.
Further, in the above technical scheme, the method for detecting content changes of nervonic acid and glyceric acid in serum is a liquid chromatography-mass spectrometry combined method.
An application of a serum organic acid molecule prediction model for early warning of severe acute pancreatitis is used for distinguishing severe acute pancreatitis from moderate acute pancreatitis within 48 h.
Further, in the above technical solution, the application method includes:
the method comprises the steps of collecting serum of a patient, detecting content change of nervonic acid and glyceric acid in the serum by adopting a liquid chromatography-mass spectrometry metabonomics analysis technology, and early warning severe acute pancreatitis if the content change of nervonic acid and glyceric acid in the serum is up-regulated by more than 2.8 times and the content change of glyceric acid in the serum is up-regulated by more than 2.1 times in normal conditions.
Advantageous effects of the invention
According to the prediction model of the severe acute pancreatitis of the serum organic acid molecules, the diagnosis sensitivity of the prediction model of the severity of the acute pancreatitis is 92.30%, the specificity is 84.70%, the false positive rate is 15.30%, the false negative rate is 7.70%, and the early warning of the severe acute pancreatitis is realized.
According to the atlanta standard of 2012, there is a 48h waiting time window for the differentiation of moderate to severe acute pancreatitis and severe acute pancreatitis: namely, the AP patient is accompanied by organ failure, the moderate-severe degree is obtained within 48h, and the severe degree is obtained if the AP patient is not recovered within 48 h. The invention can early warn patients with severe transformation risk by detecting the serum content of the Nervonic acid and the Glyceric acid, and select a proper treatment strategy, thereby having important clinical significance.
Drawings
FIG. 1 shows a chromatogram (A) and a mass spectrum (B) of Nervonic acid.
FIG. 2 shows a chromatogram (A) and a mass spectrum (B) of Glyceric acid.
FIG. 3 shows the abundance change of Nervonic acid in different groups.
FIG. 4 shows the abundance change of Glyceric acid in different groups.
FIG. 5 is a ROC curve analysis of Nervonic acid and Glycric acid to distinguish MSAP from SAP.
Detailed Description
The following non-limiting examples will allow one of ordinary skill in the art to more fully understand the present invention, but are not intended to limit the invention in any way.
Example 1
The invention provides a serum organic acid molecule prediction model for early warning of severe acute pancreatitis and application thereof, and the invention is further described in detail below in order to make the purpose, technical scheme and effect of the invention clearer and more clear. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
The invention provides a serum organic acid molecule severe acute pancreatitis detection kit, wherein the biomarkers for predicting severe acute pancreatitis are Nervonic acid (Nervonic acid) and Glyceric acid (Glyceric acid).
The severe acute pancreatitis prediction model comprises Nervonic acid and Glyceric acid which are small molecules metabolized by 2 organic acid molecules. Nervonic acid expression is up-regulated, while Glyceric acid expression is up-regulated, both molecules have a synergistic effect as markers.
In other words, the invention determines a prediction model of the severe acute pancreatitis of the serum organic acid molecules, and the biomarker for predicting the acute pancreatitis is the combination of 2 serum organic acid molecules. The serum organic acid molecules Nervonic acid and Glyceric acid are detected, the sensitivity, the specificity and the AUC of the detection to diagnosis of severe acute pancreatitis are 83.3%, 89.5% and 0.915 respectively, the pathological development condition and the disease severity of acute pancreatitis can be accurately reflected, and 48h early warning to severe acute pancreatitis is realized.
The invention detects whether 2 organic acid molecular members forming the severe acute pancreatitis prediction model exist in serum or not by a liquid chromatography-mass spectrometry metabonomics analysis technology. The invention takes the combination of the expression differential serum organic acid molecules as a prediction model of the severe acute pancreatitis, is used for predicting the severity of diseases, and can greatly improve the specificity and the accuracy of diagnosis of the severe acute pancreatitis. The serum is collected quickly and simply, and compared with the imaging detection method of acute pancreatitis, such as CT and MRI detection, the serum has no toxic or side effect on patients and has the effect of quantitative detection.
Compared with the existing scale detection and imaging diagnosis used clinically, the method for detecting the change of the expression level of the organic acid molecules in the serum of the acute pancreatitis has the advantages of reduced human subjectivity, higher accuracy and quantitative detection effect.
The present invention will be described in detail below with reference to specific examples.
Reagent: chromatographic grade methanol, acetonitrile, formic acid, ammonia were purchased from Fisher Science (Fair Lawn, USA). Chromatographic grade ammonium bicarbonate was purchased from Sigma-Aldrich (st. louis, USA). Ultrapure water was prepared using a MILI-Q purified water system (Merck KGaA, Darmstadt, Germany).
The experimental steps are as follows:
1. 50 μ L of serum samples were placed in 96-well plates and 200 μ L of methanol/acetonitrile (1: 1, v/v) was added.
2. Vortex and shake for 5 minutes, stand at 4 ℃ for 10 minutes and centrifuge at 5010g in a plate centrifuge at 4 ℃ for 20 minutes.
3. After 200. mu.L of the supernatant was transferred to a new 96-well plate, it was lyophilized in a freeze-dryer (about 2 to 3 hours, based on lyophilization)
4. And (4) sealing and storing at low temperature for mass spectrometry.
5.80. mu.L acetonitrile-water (1:3, v/v) was redissolved, vortexed for 3 minutes, and centrifuged at 5010g for 20 minutes at 4 ℃.
1. Taking 70 mu L of supernatant, and loading for detection.
An Acquity HSS C18 column (Waters Co., USA,1.8 μm, 2.1X 100mm) was used, with water as mobile phase A and acetonitrile/methanol (1: 1, v/v) as mobile phase B, both containing 0.4% ammonium bicarbonate buffer salt and 0.1% ammonia. The mobile phase gradient was: the mobile phase B gradient increased linearly from 2% to 100% over 10 min, and the column was washed and equilibrated for an additional 5 min. The flow rate, the amount of sample introduced, and the column temperature were 0.4mL/min, 5. mu.L, and 50 ℃ respectively.
Using a heated electrospray ionization source (ESI-), the various parameters were as follows: sheath gas 45 arb; an assist gas 10 arb; the heater temperature was 355 ℃; the capillary temperature is 320 ℃; S-Lens RF level 55%. The metabolome extracts were performed in full scan mode (FS) with a resolution of 70000FWHM, a maximum injection time of 200ms, and an automatic gain control target (AGC) of 1 × 106. The data acquisition uses a scanning range of 70-1000 m/z.
2. As a result:
after the serum of 39 patients with mild, 20 patients with moderate severe acute pancreatitis and 19 patients with severe acute pancreatitis is analyzed by an ultra 3000 type ultra-high performance liquid chromatograph and a QOxctive high resolution mass spectrometer (Thermo Scientific, USA), the expression difference of Nervonic acid and Glycic acid in a severe acute pancreatitis group is obvious compared with that in a mild or moderate acute pancreatitis group, and the Nervonic acid content in an SAP group is increased by 1.728 times than that in an MSAP group; meanwhile, the content of the Glyceric acid in the SAP group is increased by 2.651 times compared with that in the MSAP group. Compared with healthy people, the expression of nervonic acid in the serum of a patient with severe acute pancreatitis is up-regulated by more than 2.8 times, and the expression of glyceric acid is up-regulated by more than 2.1 times; compared with healthy people, the expression of nervonic acid in the serum of patients with moderate and severe acute pancreatitis is up-regulated by 1.6-2.8 times, and the expression of glyceric acid is up-regulated by 0.8-2.1 times. Establishing a disease prediction model by utilizing SSPS 19.0 statistical analysis software through binary Logistic regression analysis, drawing an ROC curve,
in conclusion, the invention provides a prediction model of severe acute pancreatitis of serum organic acid molecules, and the prediction model has the diagnostic sensitivity of 83.3 percent, the specificity of 89.5 percent and the AUC of 0.915 to severe acute pancreatitis. According to the invention, whether the expression difference metabolites exist in serum is detected through a non-targeted metabonomics analysis technology, and the combination of the expression difference metabolites is used as an acute pancreatitis severity prediction model for clinical diagnosis and disease severity prediction, so that the specificity and accuracy of early diagnosis of severe acute pancreatitis can be greatly improved. The serum is collected quickly and simply, and compared with the imaging detection method of acute pancreatitis, such as CT and MRI detection, the serum has no toxic or side effect on patients and has the effect of quantitative detection. Compared with the prior scale detection and imaging diagnosis used clinically, the method for detecting the change of the expression level of the organic acid molecules in the serum of the acute pancreatitis has the advantages of artificial subjectivity reduction and relative accuracy.
It is to be understood that the invention is not limited to the examples described above, but that modifications and variations may be effected thereto by those of ordinary skill in the art in light of the foregoing description, and that all such modifications and variations are intended to be within the scope of the invention as defined by the appended claims.
Claims (5)
1. The utility model provides a serum organic acid molecule prediction model of early warning severe acute pancreatitis, its characterized in that, the prediction model is the composition of serum organic acid molecule, the composition is nervonic acid and glyceric acid, and through detecting the content change of nervonic acid and glyceric acid in the serum, sick 48h in the patient, distinguish severe acute pancreatitis and severe acute pancreatitis in the middle of.
2. The model for predicting organic acid molecules in serum according to claim 1, wherein the content of nervonic acid and glyceric acid in the serum is measured as:
compared with healthy people, the expression of nervonic acid in the serum of a patient with severe acute pancreatitis is up-regulated by more than 2.8 times, and the expression of glyceric acid is up-regulated by more than 2.1 times;
compared with healthy people, the expression of nervonic acid in the serum of patients with moderate and severe acute pancreatitis is up-regulated by 1.6-2.8 times, and the expression of glyceric acid is up-regulated by 0.8-2.1 times.
3. The model of claim 1, wherein the method for detecting the content of nervonic acid and glyceric acid in serum is a combination of liquid chromatography-mass spectrometry.
4. An application of a serum organic acid molecule prediction model for early warning of severe acute pancreatitis is characterized by being used for distinguishing moderate and severe acute pancreatitis from severe acute pancreatitis within 48 h.
5. The application according to claim 4, wherein the application method comprises:
the method comprises the steps of collecting serum of a patient, detecting content change of nervonic acid and glyceric acid in the serum by adopting a liquid chromatography-mass spectrometry metabonomics analysis technology, and early warning severe acute pancreatitis if the content change of nervonic acid and glyceric acid in the serum is up-regulated by more than 2.8 times and the content change of glyceric acid in the serum is up-regulated by more than 2.1 times in normal conditions.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20160282351A1 (en) * | 2013-10-28 | 2016-09-29 | Salivatech Co., Ltd. | Salivary biomarkers for cancers, methods and devices for assaying the same, and methods for determining salivary biomarkers for cancers |
| US20200390777A1 (en) * | 2017-12-19 | 2020-12-17 | Puretech Lyt, Inc. | Lipid prodrugs of mycophenolic acid and uses thereof |
| CN113136369A (en) * | 2020-01-20 | 2021-07-20 | 温州医科大学附属第一医院 | Cell modified with lipoxygenase gene and preparation method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20160282351A1 (en) * | 2013-10-28 | 2016-09-29 | Salivatech Co., Ltd. | Salivary biomarkers for cancers, methods and devices for assaying the same, and methods for determining salivary biomarkers for cancers |
| US20200390777A1 (en) * | 2017-12-19 | 2020-12-17 | Puretech Lyt, Inc. | Lipid prodrugs of mycophenolic acid and uses thereof |
| CN113136369A (en) * | 2020-01-20 | 2021-07-20 | 温州医科大学附属第一医院 | Cell modified with lipoxygenase gene and preparation method and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| JOHANNIS P.KAMERLING ET AL: "Determination of the configurations of lactic and glyceric acids from human serum and urine by capillary gas—liquid chromatography", 《JOURNAL OF CHROMATOGRAPHY B: BIOMEDICAL SCIENCES AND APPLICATIONS》 * |
| LITING SUN ET AL: "Metabolomic profiling of ovary in mice treated with FSH using ultra performance liquid chromatography/mass spectrometry", 《BIOSCI REP》 * |
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