Application of CTRP1 as a biomarker in the preparation of early diagnosis products for pediatric sepsis

Abstract

The invention proposes the application of CTRP1 as a biomarker in the preparation of an early diagnosis product for childhood sepsis, and belongs to the field of medical technology; the serum CTRP1 level is detected by an ELISA method, and a human complement C1q tumor necrosis factor-related protein 1 (C1QTNF1 / CTRP1) enzyme-linked immunosorbent assay (ELISA) kit is used to perform qualitative or quantitative analysis according to the depth of color; and the serum CTRP1 expression levels of children with infectious diseases in various age groups are divided into 29 days-≤1 year old, 1 year-≤6 years old, and 7 years-<18 years old according to age; and it is used for the early diagnosis of childhood sepsis; the invention applies serum CTRP1 to the early diagnosis of childhood sepsis for the first time, and the detection method has the advantages of high sensitivity, strong specificity, simple and quick operation, etc., and is low in price and suitable for popularization and application.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and in particular relates to the application of CTRP1 in the preparation of a product for diagnosing sepsis in children. Background Art

[0002] Sepsis is an organ dysfunction secondary to infection. Due to its high morbidity, high mortality rate and huge medical burden, it has always been a hot topic in the field of critical care medicine. Early diagnosis and appropriate treatment can greatly improve the prognosis of the disease. About 50% of sepsis cases occur in children. The clinical manifestations of pediatric sepsis are highly heterogeneous due to factors such as age, underlying diseases, primary infection sites, and primary infection bacteria, which brings great difficulties to its early diagnosis and treatment. Therefore, in order to effectively improve the prognosis of children with sepsis, it is urgent to continue to explore highly sensitive and specific biological markers or evaluation indicators with high clinical feasibility, improve the early identification rate of children with severe sepsis, further develop new and effective treatment measures, carry out stratified and individualized treatment, and establish an effective prognosis evaluation system.

[0003] The pathogenesis of sepsis is very complex and is related to a series of physiological and pathological processes such as infection, inflammation, immunity, coagulation and tissue damage. The inflammatory factors released by the infection involve multiple cells and multiple systems throughout the body through the neuro-endocrine immune regulation system, forming a huge and complex network. The cascade effect is amplified and mutually restricted, so clinical diagnosis and treatment are very difficult. The current treatment method is to give effective antibacterial treatment combined with comprehensive treatment of changes in various organ systems, but the effect is not good and the mortality rate remains high. Therefore, there is an urgent need to find rapid diagnostic biomarkers for pediatric sepsis and new targets for effective treatment.

[0004] Inflammatory cytokines play a pivotal role in the development of sepsis. Complement C1q / tumor necrosis factor-related protein 1 (CTRP1) is a recently discovered adipose-derived cytokine, with a total of 15 members (CTRP1-15). This family is highly conserved and widely expressed in human and mouse tissues. It has been shown to have a wide range of effects on metabolism, inflammation, tumor metastasis, blood vessels, and ischemic injury diseases. CTRP1 is a secreted glycoprotein with a relative molecular mass of 35,000 and is composed of 281 amino acids. It can be expressed in a variety of tissues such as fat, heart, placenta, liver, muscle, kidney, prostate, and ovary, and its expression level in children and adults is different.

[0005] So far, there have been no reports internationally on the relationship between CTRP1 and childhood sepsis. Summary of the invention

[0006] In view of the problems in the prior art, the present invention provides the use of CTRP1 as a biomarker in the preparation of an early diagnosis product for childhood sepsis.

[0007] In order to achieve the above purpose, the technical solution adopted in this application is:

[0008] The invention discloses an application of CTRP1 as a biomarker in the preparation of an early diagnosis product for sepsis in children, wherein the product comprises a kit.

[0009] Preferably, the reagents in the kit include one or more of CTRP1 standard, sample dilution buffer, biotin-labeled antibody working solution, antibody dilution buffer, SABC working solution, SABC dilution buffer, TMB substrate solution, and stop solution.

[0010] Preferably, the kit is used by performing ELISA to detect the expression level of CTRP1 in a biological sample.

[0011] Preferably, the biological sample comprises plasma, serum or blood.

[0012] Preferably, the child is between 29 days and 18 years old.

[0013] Preferably, the diagnosis is based on: compared with infection without sepsis, the expression level of serum CTRP1 is significantly decreased when sepsis is present.

[0014] Preferably, the ELISA method for detecting serum CTRP1 expression levels is divided into 29 days-≤1 year old, 1 year old-≤6 years old, and 7 years old-<18 years old according to age.

[0015] Compared with the prior art, the advantages and positive effects of the present invention are:

[0016] The application of CTRP1 in the preparation of a product for diagnosing childhood sepsis is to detect serum CTRP1 levels by ELISA, using a human complement C1q tumor necrosis factor-related protein 1 (C1QTNF1 / CTRP1) enzyme-linked immunosorbent assay (ELISA) kit to perform qualitative or quantitative analysis based on the depth of color; and the serum CTRP1 expression levels of children with infectious diseases in various age groups are divided into 29 days-≤1 year old, 1 year old-≤6 years old, and 7 years old-<18 years old according to age; for the early diagnosis of childhood sepsis; the present invention is the first to apply serum CTRP1 to the diagnosis of childhood sepsis, and the detection method has the advantages of high sensitivity, strong specificity, simple and quick operation, etc., and is low in price and suitable for popularization and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0018] Figure 1 The predictive efficacy of serum CTRP1 levels for sepsis in children (age 29 days-18 years);

[0019] Figure 2 The predictive efficacy of serum CTRP1 levels for sepsis in infants (age 29 days-≤1 year);

[0020] Figure 3 The predictive efficacy of serum CTRP1 levels for sepsis in young children (1-≤6 years old);

[0021] Figure 4 Predictive efficacy of serum CTRP1 levels for sepsis in older children (7-<18 years old). DETAILED DESCRIPTION

[0022] In order to more clearly understand the above-mentioned purpose, features and advantages of the present invention, the present invention is further described below in conjunction with the accompanying drawings and embodiments. It should be noted that the embodiments of the present application and the features in the embodiments can be combined with each other without conflict.

[0023] In the following description, many specific details are set forth to facilitate a full understanding of the present invention. However, the present invention may also be implemented in other ways than those described herein. Therefore, the present invention is not limited to the specific embodiments of the following disclosure.

[0024] Example 1, use of CTRP1 as a biomarker in the preparation of a product for early diagnosis of sepsis in children, the product comprising a kit;

[0025] The reagents in the kit include CTRP1 standard, sample dilution buffer, biotin-labeled antibody working solution, antibody dilution buffer, SABC working solution, SABC dilution buffer, TMB substrate solution, and stop solution;

[0026] 1. Case collection and grouping

[0027] The clinical data of 220 children with infection who visited the Children's Intensive Care Unit of Shandong Provincial Hospital from January 2021 to December 2023 were collected, and 100 children who underwent healthy physical examinations during the same period were included. According to the clinical symptoms and diagnosis of the included subjects, they were divided into a normal control group (100 cases), an infection non-sepsis group (80 cases), and an infection and sepsis group (140 cases). Based on the diagnosis of childhood sepsis and the growth and development rules of children, all the above subjects were divided into a ≤1 year old group, a 1-≤7 year old group, and a 7-<18 year old group. Serum CTRP1 and other related laboratory indicators were collected from all subjects within 24 hours of admission. SPSS 26.0 software was used for data analysis to compare the clinical data, laboratory indicators, and serum CTRP1 levels of children in each group. The receiver operating characteristic curve (ROC curve) was used to compare the diagnostic value of different indicators in childhood sepsis.

[0028] A total of 125 infected children (aged 29 days-<18 years) were selected as the research subjects. According to whether they developed sepsis, the children were divided into the sepsis group (n=85) (median age was 4.5 years, including 50 males) and the non-sepsis group (n=40) (median age was 5.5 years, including 22 males). The diagnosis of pediatric sepsis was based on the "2012 Surviving Sepsis Campaign: Guidelines for the Diagnosis and Treatment of Severe Sepsis and Septic Shock". Children were divided into 29 days-≤1 year old, 1-≤6 years old, and 7-<18 years old groups according to different age groups.

[0029] 2. Use the above kit to perform ELISA to detect serum CTRP1 expression level

[0030] The detection instruments include a microplate reader and a 37°C incubator.

[0031] The application principle of the kit is based on the sandwich enzyme-linked immunosorbent assay technology to detect the solid phase carrier of the antigen or antibody in the biological sample and the enzyme label of the antigen or antibody; wherein the antigen or antibody on the surface of the solid phase carrier still retains its immunological activity, and the enzyme-labeled antigen or antibody retains both its immunological activity and the enzyme activity;

[0032] The biological sample is serum, and the serum collection method is: collecting peripheral blood from the research subject, and collecting serum by centrifugation;

[0033] During the measurement, the specimen to be tested (the antibody or antigen to be measured) reacts with the antigen or antibody on the surface of the solid phase carrier; the antigen-antibody complex formed on the solid phase carrier can be separated from other substances in the liquid by washing, and then the enzyme-labeled antigen or antibody is added and also bound to the solid phase carrier through reaction. At this time, the amount of enzyme on the solid phase is in a certain ratio to the amount of the substance to be tested in the specimen. After the enzyme reaction substrate is added, the substrate is catalyzed by the enzyme to become a colored product. The amount of the product is directly related to the amount of the substance to be tested in the specimen, so qualitative or quantitative analysis can be performed based on the depth of color.

[0034] 3. Experimental Data Processing

[0035] (1) As shown in Table 1, for children of all ages with infectious diseases (29 days < 18 years old), serum CTRP1 levels were significantly decreased when they were complicated with sepsis compared with those without sepsis (170.77 vs 209.33, P < 0.05).

[0036] Table 1 Comparison of serum CTRP1 levels in normal controls, children with non-septic infections and children with sepsis aged 29 days to <18 years

[0037]

[0038] (2) As shown in Tables 2-4, when the infection was complicated with sepsis, the serum CTRP1 levels of children aged 29 days to ≤1 year, 1 to ≤6 years, and 7 to <18 years were significantly lower than those of children without sepsis (219.10 vs 347.20; 146.15 vs 210.26; 145.17 vs 189.56, all P < 0.05).

[0039] Table 2 Comparison of serum CTRP1 levels in normal controls, non-septic infected and septic children aged 29 days to ≤ 1 year

[0040]

[0041] Table 31 - Comparison of serum CTRP1 levels in normal controls, non-septic infected children, and septic children aged ≤6 years

[0042]

[0043] Table 47 - Comparison of serum CTRP1 levels in normal controls, children with non-septic infections and children with sepsis aged <18 years

[0044]

[0045] It should be noted that, VS sepsis group, *P<0.05 **P<0.05.

[0046] (3) If Figure 1 As shown, the threshold value of serum CTRP1 for diagnosing infection and sepsis in children (29 days < 18 years old) is 198.74 ng / ml. When the level is lower than 198.74 ng / ml, sepsis in children can be diagnosed (area under the ROC curve 0.721, 95% CI 0.622-0.789, sensitivity 68.6%, specificity 71.1%).

[0047] (4) If Figure 2As shown, for children aged 29 days to ≤1 year, the threshold of serum CTRP1 for diagnosing infection with sepsis was ≤301.59 ng / ml (area under the ROC curve was 0.877, 95% CI was 0.75-1.00, sensitivity was 93.8%, and specificity was 75%).

[0048] (5) Figure 3 As shown, for children aged 1-≤6 years, the threshold of serum CTRP1 for diagnosing infection with sepsis was ≤176.34 ng / ml (area under the ROC curve was 0.804, 95% CI was 0.677-0.929, sensitivity was 59%, and specificity was 41.9%).

[0049] (6) If Figure 4 As shown, in children aged 7-<18 years, the threshold of serum CTRP1 for diagnosing infection and sepsis was ≤130.44 ng / ml (area under the ROC curve was 0.658; 95% CI was 0.543-0.816; sensitivity was 41%; specificity was 48%).

[0050] According to the above experimental data, serum CTRP1 expression level can be used for the early diagnosis of sepsis in children; and the diagnostic threshold of sepsis in different age groups is different.

[0051] The above description is only a preferred embodiment of the present invention and does not limit the present invention in other forms. Any technician familiar with the profession may use the technical content disclosed above to change or modify it into an equivalent embodiment with equivalent changes and apply it to other fields. However, any simple modification, equivalent change and modification made to the above embodiment based on the technical essence of the present invention without departing from the content of the technical solution of the present invention still falls within the protection scope of the technical solution of the present invention.

Claims

1. Use of CTRP1 in plasma, serum or blood as a biomarker in the preparation of an early diagnosis product for sepsis in children, characterized in that: The products include kits.

2. The use of CTRP1 in plasma, serum or blood as a biomarker in the preparation of a product for early diagnosis of sepsis in children according to claim 1, characterized in that: The reagents in the kit include one or more of CTRP1 standard, sample dilution buffer, biotin-labeled antibody working solution, antibody dilution buffer, SABC working solution, SABC dilution buffer, TMB substrate solution, and stop solution.

3. The use of CTRP1 in plasma, serum or blood as a biomarker in the preparation of a product for early diagnosis of sepsis in children according to claim 2, characterized in that: The kit is used as follows: ELISA is performed to detect the expression level of CTRP1 in biological samples.

4. The use of CTRP1 in plasma, serum or blood as a biomarker in the preparation of a product for early diagnosis of sepsis in children according to any one of claims 1 to 3, characterized in that: The children ranged in age from 29 days to 18 years.

5. The use of CTRP1 in plasma, serum or blood as a biomarker in the preparation of a product for early diagnosis of sepsis in children according to claim 4, characterized in that: The diagnosis is based on the fact that compared with infection without sepsis, the expression level of serum CTRP1 is significantly decreased when sepsis is present.

6. The use of CTRP1 in plasma, serum or blood as a biomarker in the preparation of a product for early diagnosis of sepsis in children according to claim 3, characterized in that: When ELISA was used to detect CTRP1 in serum, the CTRP1 threshold for diagnosing infection and sepsis in children aged 29 days to ≤1 year was ≤301.59 ng / ml; the CTRP1 threshold for diagnosing infection and sepsis in children aged 1 year to ≤6 years was ≤176.34 ng / ml; and the CTRP1 threshold for diagnosing infection and sepsis in children aged 7-<18 years was ≤130.44 ng / ml.

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