CN115267162A - Mental disease linear discrimination model and diagnosis equipment based on multi-protein combination - Google Patents

Abstract

The invention discloses a serum protein model constructed by Linear Discriminant Analysis (LDA), wherein the model is one or more of serum brain-derived neurotrophic factor (BDNF), VGF, double-tail-C gene homolog 1 (BICC 1), cortisol and/or C-reactive protein (CRP). The invention also discloses application of the serum protein model reagent in preparing mental disease diagnosis products. The invention also discloses an enzyme-linked immunosorbent assay kit. The present inventors have found that there is a difference in the expression of BDNF, VGF, BICC1, cortisol and CRP in control serum of depression, schizophrenia, bipolar disorder, panic disorder and non-psychotic disorder. The accuracy of the invention reaches 96.5 percent when the five proteins are combined by the LDA method to distinguish the depression, schizophrenia, bipolar disorder, panic disorder and non-mental disease contrast. Therefore, the multi-protein combined model constructed by the five serum proteins by using LDA has higher clinical application value in the diagnosis of mental diseases.

Description

Mental disease linear discrimination model and diagnosis equipment based on multi-protein combination

Technical Field

The invention relates to the field of biological science, in particular to a mental disease linear discrimination model and diagnosis equipment based on multi-protein combination.

Background

According to the WHO International psychiatric Association, more than 1/3 of the world's population suffers from at least one mental disorder in their lifetime. Mental diseases such as depression, schizophrenia, bipolar disorder and panic disorder are commonly high in the global scope and are an important component of the loss of healthy life of the global disabilities. Since the etiology and pathological mechanism of mental disorders are unknown, the diagnosis of mental disorders has heretofore relied primarily on the subjective identification by psychiatrists of the disease's major clinical symptoms. However, due to the non-specificity of symptoms, there is often overlap of symptoms between various diseases, so that the misdiagnosis rate of mental diseases is always high, and the effective treatment of patients is delayed and even the treatment outcome is worsened. Therefore, the search for biomarkers related to the diagnosis of these diseases and the development of diagnostic methods that help to improve the accuracy of the diagnosis of mental diseases are important problems to be solved in clinical practice.

Disclosure of Invention

Aiming at the defects of the prior art, the invention provides a mental disease linear discrimination model and diagnosis equipment based on multi-protein combination.

The purpose of the invention can be realized by the following technical scheme:

a mental disease diagnostic apparatus comprising:

the kit is a reagent for detecting one or more of BDNF, VGF, BICC1, cortisol and/or CRP;

and the data processing module is configured with a linear discrimination model or algorithm and is used for analyzing and processing the detection value of the kit.

Optionally, the agent comprises a specific antibody to BDNF, VGF, BICC1, cortisol and/or CRP protein.

Optionally, the reagent for detecting the content of BDNF, VGF, BICC1, cortisol and/or CRP protein by using an immunological method

Optionally, the specific antibody comprises a monoclonal antibody or a polyclonal antibody.

Optionally, the psychiatric disorder comprises depression, schizophrenia, bipolar disorder, or panic disorder.

Optionally, the kit comprises an enzyme-linked immunosorbent assay kit or an immunoblotting kit.

The invention has the beneficial effects that:

the application combines serum brain-derived neurotrophic factor (BDNF), VGF, double-tail-C gene homolog 1 (BICC 1), cortisol and/or C-reactive protein (CRP) into a polyprotein model by an LDA method to serve as a biomarker for specifically diagnosing mental diseases, and provides a brand-new way for detecting depression, schizophrenia, bipolar disorder and panic disorder; the method adopts a polyprotein model containing BDNF, VGF, BICC1, cortisol and CRP constructed by an LDA method as an index to diagnose depression, schizophrenia, bipolar disorder, panic disorder and panic disorder in a mixed population containing depression, schizophrenia, bipolar disorder, panic disorder and non-mental disease control, wherein the area value AUC under an ROC curve is up to 1.0, and the overall accuracy is up to 96.5%; the sensitivity and specificity of the diagnosis of depression, schizophrenia, bipolar disorder and panic disorder are respectively as high as 90.0 percent and 99.0 percent, 98.0 percent and 98.0 percent, 100 percent and 100 percent, 96.0 percent and 100.0 percent, and the diagnosis value of the mental diseases is very good. AUC has certain accuracy when being 0.7-0.9, and AUC has higher accuracy when being more than 0.9.

Drawings

The invention will be further described with reference to the accompanying drawings.

FIG. 1 is a graph of the quantification of Zhou Xieqing BDNF, VGF, BICC1, cortisol and CRP proteins in addition to a control of depression, schizophrenia, bipolar disorder, panic disorder, and non-psychiatric disease;

fig. 2 is a visual scattergram of a serum BDNF, VGF, BICC1, cortisol and CRP-based multi-protein model constructed by the LDA method to differentiate depression, schizophrenia, bipolar disorder, panic disorder, and non-psychotic control groups;

figure 3 is a graph of the results of a 5 fold cross validation using a subject working profile analysis to distinguish depression, schizophrenia, bipolar disorder, panic disorder, and non-psychotic controls from mixed populations comprising depression, schizophrenia, bipolar disorder, panic disorder, and non-psychotic controls, respectively.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without any inventive step, are within the scope of the present invention.

Materials, reagents and the like used in the following examples are commercially available unless otherwise specified. The experimental procedures in the examples, in which specific conditions are not specified, are generally carried out under conventional conditions or conditions recommended by the manufacturer.

Serum samples of 50 patients with depression, 50 schizophrenia, 55 bipolar disorder, 50 panic disorder patients and 50 non-psychotic disease controls who had not been administered antipsychotic drugs, mood stabilizers, benzodiazepines, etc. within two weeks were collected. In particular, the diagnosis of patients with depression, schizophrenia, bipolar disorder and panic disorder was diagnosed and confirmed by three clinically experienced third-class psychiatrists (chief/assistant chief physicians, chief therapists and high-age hospitalizers) according to the criteria for single or recurrent depressive disorder, paranoid schizophrenia, bipolar disorder (manic episode, depressive episode or mixed episode) and panic disorder diagnosis in the handbook of diagnosis and statistics of psychiatric disorders (fourth edition) (DSM-IV) recruited by the major hospitals in subsidiary middle school of southeast university, the third hospital in lakehou city and the third hospital in huaian city, respectively, and the samples of healthy controls were from the recruited social staff.

The tests and evaluations that were made included: serum BDNF, VGF, BICC1, cortisol and CRP protein concentration assays; scales to assess the Severity of the condition in each group of patients individually include the Positive And Negative Symptoms Scale (PANSS), the Positive symptoms Scale (the Scale for Assessment Positive Symptom, SAPS), the Hamilton Depression Scale (17-item Hamilton Depression Scale, HAMD-17), the Young Mania Scale (Young manic Scale, YMRS), the Hamilton Anxiety Scale (Hamilton Anxiety Scale, HAMA), the Panic Disorder Severity Scale (PDSS).

Serum BDNF, VGF, BICC1, cortisol and CRP protein concentrations were measured using an ELISA kit using the following detailed information: BDNF and cortisol kit: r & D Systems, mutlukent Mah, arda Sk, USA; VGF and BICC1 kit: hermes Criterion Biotechnology, vancouver, B.C., canada; CRP kit: rayBiotech, norcross, GA, USA. The lower detection limits of the BDNF, VGF, BICC1, CRP and cortisol ELISA kits were <20pg/mL, 35 pg/mL, 25pg/mL, 34pg/mL and 0.2ng/mL, respectively.

The above kits provide necessary but not all reagents required to detect the concentrations of BDNF, VGF, BICC1, cortisol and CRP proteins, respectively: the kit comprises a 96-well enzyme label plate, 2 bottles of human BDNF/VGF/BICC 1/cortisol/CRP protein standard, 1 bottle of 20-fold 25ml washing solution, 2 bottles of biotin-labeled anti-human BDNF/VGF/BICC 1/cortisol/CRP, 1 bottle of 200ul avidin, 1 bottle of 12ml buffer solution, 1 bottle of 8ml stop solution, 1 bottle of 30ml color development solution A and 1 bottle of 15ml stop solution. In addition, the additional reagents and materials required are as follows: enzyme-linked immunosorbent assay instrument, pipettor, pipette, graduated cylinder, absorbent paper, distilled water or deionized water, data analysis and drawing software, etc.

All required items are prepared before the detection starts. The specific detection steps are as follows:

(1) Determining the number of the holes of the enzyme label plate coated with the antibody required by the detection, wherein a plurality of holes are required to be arranged for each sample, standard substance and blank;

(2) Adding a sample: after the BDNF/VGF/BICC 1/cortisol/CRP protein standard substance is diluted by using the diluent A in a multiple ratio, 50ul of each standard substance with corresponding concentration is sequentially added into a row of enzyme-labeled plate holes coated with BDNF/VGF/BICC 1/cortisol/CRP antibodies in advance, and 1-hole blank comparison is set (only the diluent A is added into each blank comparison hole, no sample is added, namely the concentration of the standard substance is 0). 50ul of the sample (serum samples from 50 depression, 50 schizophrenia, 55 bipolar disorder, 50 panic disorder patients and 50 non-psychotic disease controls) was added to the remaining wells of the microplate and gently mixed. Incubation at 37 ℃ for 45 min;

(3) Preparing liquid: diluting the washing liquor concentrated by 20 times with distilled water for later use;

(4) Cleaning: pouring out the liquid in the holes of the enzyme-labeled plate, spin-drying, filling washing liquid into each hole, standing for 30 seconds, discarding, and repeatedly washing for 4 times;

(5) Adding biotin-labeled anti-human BDNF/VGF/BICC 1/cortisol/CRP: adding 50ul of biotin-labeled anti-human BDNF/VGF/BICC 1/cortisol/CRP into all enzyme-labeled plate holes, and incubating for 30 minutes at 37 ℃;

(6) Cleaning: repeating operation 4;

(7) Adding avidin: 50ul of avidin was added to all wells of the microplate and mixed gently. Incubation at 37 ℃ for 15 min;

(8) Cleaning: repeating operation 4;

(9) Color development: adding 50ul of the chromogenic solution A and the chromogenic solution B into all enzyme label plate holes, and incubating for 15 minutes at 37 ℃;

(10) And (3) terminating the reaction: 50ul of stop solution was added to all wells of the microplate and the reaction was stopped (color immediately changed from blue to yellow);

(11) And (3) analysis: the optical density (O.D. value) was measured at 450nm with an enzyme-labeled detector within 15 minutes after addition of the stop solution with the blank well as zero. And performing series dilution on the standard protein of the BICC1 at a known concentration, drawing a standard curve after measuring the OD value, and calculating the content of BDNF/VGF/BICC 1/cortisol/CRP in the measured sample according to the standard curve.

The demographic and clinical characteristics of the subjects are shown in table 1, and the results of 5-fold cross-validation ROC curve analysis for distinguishing depression, schizophrenia, bipolar disorder, panic disorder and non-psychotic control from mixed populations including depression, schizophrenia, bipolar disorder, panic disorder and non-psychotic control according to serum BDNF, VGF, BICC1, cortisol and CRP concentrations and a polyprotein model constructed based on the five proteins by the LDA method are shown in table 2. The results of the concentrations of BDNF, VGF, BICC1, cortisol, CRP protein in the serum of different mental disease patients and non-mental disease controls are shown in fig. 1, the visual scattergram for distinguishing the depression, schizophrenia, bipolar disorder, panic disorder and non-mental disease control groups by the multi-protein model based on serum BDNF, VGF, BICC1, cortisol and CRP constructed by the LDA method is shown in fig. 2, and the ROC graph for distinguishing the depression, schizophrenia, bipolar disorder, panic disorder and non-mental disease control groups in the mixed group containing depression, schizophrenia, bipolar disorder, panic disorder and non-mental disease controls by the multi-protein model based on serum BDNF, VGF, BICC1, cortisol and CRP constructed by the serum BDNF, BICC1, cortisol and CRP is shown in fig. 3.

Wherein, A-E in FIG. 1 are comparisons of serum BDNF, VGF, BICC1, CRP, and cortisol concentrations in depression, schizophrenia, bipolar disorder, panic disorder, and non-psychotic disease control groups, respectively; each psychiatric disorder is abbreviated as follows: MDD, depression; SZ, schizophrenia; BPD, bipolar disorder; PD, panic disorder, HC, non-psychiatric disease control;

compared with the non-mental disease control group,^*is represented by P<0.05，^**Represents P<0.001；

In comparison with the group of panic disorders,^#represents P<0.05，^##Represents P<0.001；

In contrast to the group of bipolar disorders,^&is represented by P<0.05，&&Represents P<0.001；

In comparison to the schizophrenic group, $ indicates P <0.05 and $ indicates P <0.001.

In FIG. 3A-E are ROC plots for the serum BDNF, VGF, BICC1, CRP and cortisol based, respectively, multi-protein LDA model to distinguish (A) depression, (B) schizophrenia, (C) bipolar disorder, (D) panic disorder and (E) non-psychotic controls in mixed populations comprising depression, schizophrenia, bipolar disorder, panic disorder and non-psychotic controls, respectively.

As can be seen in fig. 1, the concentrations of serum BDNF, VGF, BICC1, CRP and cortisol in different psychiatric disorders vary. Among them, serum BDNF levels were significantly lower in all disease groups than in HC group (fig. 1A). In the disease group, MDD group had higher serum BDNF levels than SZ, BPD, and PD groups, SZ group was higher than BPD and PD groups, and BPD group was lower than PD group (fig. 1A). Serum VGF levels were significantly different between any two of the five groups (fig. 1B). Specifically, serum VGF levels were lower in the MDD group than in the SZ, BPD and HC groups and higher in the PD group, with the SZ group lower than in the BPD and HC groups and higher than in the PD group, with the BPD group higher than in all other study groups, and with the PD group lower than in all other study groups (fig. 1B). Similarly, the levels of BICC1 were significantly increased in all disease groups compared to the HC group, with serum BICC1 levels lower in the MDD and SZ groups than in the BPD and PD groups, and serum BICC1 levels higher in the BPD group than in the PD group (fig. 1C). Serum CRP and cortisol levels were higher in the MDD and BPD groups than in the SZ and HC groups and lower in the PD group; the PD group had higher serum CRP and cortisol levels than the other four study groups, while the SZ group was lower than the other three disease groups (fig. 1D and 1E). In addition, serum CRP levels were also significantly higher in SZ group than in HC group (fig. 1D and 1E), but serum cortisol was not significantly different from HC group.

Fig. 2 shows that the serum BDNF, VGF, BICC1, cortisol and CRP-based polyprotein model constructed by the LDA method can well distinguish different psychiatric patients and non-psychiatric controls from each other in a mixed population containing depression, schizophrenia, bipolar disorder, panic disorder, and non-psychiatric controls. Specifically, the discrimination of the BPD group is excellent, and the BPD group can be completely discriminated from the HC, MDD, SZ, and PD groups; the discrimination of the PD group from the BPD, HC and SZ groups is also good; the MDD group and the PD group, the MDD group and the SZ group, and the SZ group and the HC group are mixed, but the overall discrimination is better.

Figure 3 shows that the serum BDNF, VGF, BICC1, cortisol and CRP based polyprotein LDA model has a high accuracy in distinguishing any of the groups in the mixed population containing depression, schizophrenia, bipolar disorder, panic disorder and non-psychotic disease controls. AUC after 5-fold cross validation reaches 1.0, and the overall accuracy reaches 96.5%. The area under the ROC curve (AUC), sensitivity, specificity, positive predictive value and overall classification accuracy when the multi-protein LDA model and single serum BDNF, VGF, BICC1, cortisol and CRP are used for distinguishing different mental diseases and non-mental diseases in a mixed population are shown in table 2. The area under the ROC curve is between 1.0 and 0.5, with AUC >0.5, the closer the AUC is to 1, indicating better diagnostic results. AUC has lower accuracy at 0.5-0.7, certain accuracy at 0.7-0.9, and higher accuracy at more than 0.9. AUC =0.5, indicating that the diagnostic method was completely ineffective and of no diagnostic value. AUC <0.5 does not fit the real situation, and rarely occurs in practice. In addition, sensitivity and specificity are also important indicators for the evaluation of diagnostic biomarkers. Studies indicate that clinically useful biomarkers or tests for correct diagnosis and classification of disease both sensitivity and specificity reach at least 80% (Schneider B, prunovic d. Novel biomarkers in major expression. Curr Opin psychiatry.2013.26 (1): 47-53.). It can be seen that a single serum protein index does not allow diagnosis of each disease in a mixed population, and the multi-protein LDA model consisting of serum BDNF, VGF, BICC1, cortisol and CRP can become a clinically useful objective diagnostic test.

TABLE 1 demographic and clinical characteristics of the subjects

Note: MDD, depression; SZ, schizophrenia; BPD, bipolar disorder; PD, panic disorder; HC, healthy control; BMI, body mass index; PANSS, positive and negative symptom scale; SAPS, positive symptom rating scale; HAMD-17, 17 hamilton depression rating scale; HAMA, hamilton anxiety rating scale; YMRS, young's manic rating scale; PDSS, panic disorder severity scale. Continuous variables are expressed as means ± standard deviation. In comparison with the HC group,^*represents P<0.05，^**Is represented by P<0.001; in comparison with the group of PDs,^#represents P<0.05，^##Is represented by P<0.001; in comparison to the BPD group,^&represents P<0.05，^&&Represents P<0.001; compared with the SZ group, the method has the advantages that,^$is represented by P<0.05，^$$Is represented by P<0.001。^aAnalyzing single-factor variance;^bchecking a chi square;^cLulskar-Wallace analysis. P<0.05 represents a statistical difference.

Table 2 results of ROC curve analysis of single serum proteins cross-validated by 5 fold and polyprotein models constructed by LDA to diagnose different psychiatric and non-psychiatric controls in mixed populations

Note: a, the positive class refers to a group needing to be separated from the mixed sample, and the corresponding negative class is the mixed sample of the rest groups except the positive class; LDA, linear discriminant analysis; BDNF, brain-derived neurotrophic factor; BICC1, bicaudal C homolog 1; CRP, C-reactive protein; MDD, depression; SZ, schizophrenia; BPD, bipolar disorder; PD, panic disorder; HC, healthy control; AUC, area under curve; PPV, positive predictive value.

In the description herein, references to the description of "one embodiment," "an example," "a specific example" or the like are intended to mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, schematic representations of the terms used above do not necessarily refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.

The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the specification and illustrated only to illustrate the principle of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the present invention, which fall within the scope of the invention as claimed.

Claims (6)

1. A mental disease diagnosis apparatus characterized by comprising:

the kit is a reagent for detecting one or more of BDNF, VGF, BICC1, cortisol and/or CRP;

and the data processing module is configured with a linear discrimination model or algorithm and is used for analyzing and processing the detection value of the kit.

2. The diagnostic device for mental disorders according to claim 1, wherein said agent comprises an antibody specific to BDNF, VGF, BICC1, cortisol and/or CRP protein.

3. The diagnostic apparatus for mental disorders according to claim 1, wherein the agent for measuring the content of BDNF, VGF, BICC1, cortisol and/or CRP protein is an agent using an immunological method.

4. The diagnostic apparatus for mental disorders according to claim 2, wherein said specific antibody comprises a monoclonal antibody or a polyclonal antibody.

5. The psychiatric disease diagnosis apparatus according to claim 1, wherein the psychiatric disease includes depression, schizophrenia, bipolar disorder or panic disorder.

6. The psychiatric disease diagnosis apparatus according to claim 1, wherein said kit comprises an enzyme-linked immunosorbent assay kit or an immunoblotting kit.

Images