Application of CCL28 as biomarker and therapeutic target in sepsis

Abstract

The invention belongs to the technical field of biological medicine, and particularly discloses application of CCL28 as a biomarker and a therapeutic target in sepsis. The invention particularly relates to application of a CCL28 gene or a detection reagent for the expression quantity of the CCL28 gene in preparation of a kit for diagnosis or prognosis monitoring of sepsis. The expression of the novel immunomodulatory factor CCL28 in a sepsis patient is obviously improved, and the expression level of the novel immunomodulatory factor CCL28 is in obvious negative correlation with poor prognosis of sepsis. Through CLP induction of a sepsis mouse model, the invention finds that the expression level of CCL28 in sepsis mouse blood, abdominal cavity lavage fluid (PLF), liver and kidney is significantly increased. By adopting the recombinant CCL28 for treatment, the bacterial load in the mice with sepsis can be remarkably reduced, and the multi-organ injury of the mice with sepsis can be reduced, so that the survival rate of the mice with sepsis can be remarkably improved. In conclusion, the invention explores a novel immunomodulatory protein CCL28, and the novel immunomodulatory protein CCL28 not only can be used as a diagnosis and prognosis biomarker of sepsis, but also can be used as a novel target spot for treating sepsis.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to the application of CCL28 as a biomarker and therapeutic target in sepsis. Background Art

[0002] The pathogenesis of sepsis involves complex inflammatory imbalances and immune dysfunction. In the early stages of sepsis, bacteria and their components bind to pattern recognition receptors (TRRs) and NODs on host cells, initiating intracellular signaling systems that synthesize and release multiple inflammatory mediators, leading to a cascade of inflammatory responses. This is accompanied by a compensatory anti-inflammatory response, resulting in immune imbalance and organ dysfunction in multiple systems, including the cardiovascular, respiratory, urinary, nervous, and hematologic systems. Therefore, immune imbalance is key to the development and progression of sepsis. Finding early and rapid diagnostic and therapeutic targets from the perspective of host immune regulation is crucial for improving the survival and prognosis of patients with sepsis.

[0003] Currently, the diagnosis of sepsis is primarily based on the diagnostic criteria for Sepsis 3.0, jointly released by the American Society of Critical Care Medicine and the European Society of Intensive Care Medicine in 2016: infection combined with a Sequential Organ Failure Assessment (SOFA) score ≥ 2. Clinically, early diagnosis and initial screening for sepsis rely on procalcitonin (PCT), C-reactive protein (CRP), and pathogen testing. However, these methods have the following drawbacks: 1) PCT can be used to guide antibiotic use and has high specificity for systemic bacterial infections, but elevations are smaller in localized infections, viral infections, and intracellular bacterial infections such as Mycoplasma pneumoniae. It has no diagnostic value for skin and soft tissue infections. Furthermore, PCT levels can be elevated to varying degrees in pathological conditions such as postoperative surgery, cardiogenic shock, and acute graft-versus-host disease, resulting in low specificity and lacking independent diagnostic capability. 2) CRP, as a common diagnostic marker for inflammation, has poor specificity. 3) Clinical microbiological testing (microscopy and culture) remains the gold standard for diagnosing pathogenic infections, but it is time-consuming, subjective, and often contains numerous interfering substances. 4) The SOFA score involves a comprehensive evaluation of multiple body systems, posing significant challenges to its early diagnosis. Currently, the treatment of sepsis still relies on traditional approaches such as antibiotics, mechanical ventilation, and fluid resuscitation. Medical treatment has not achieved breakthroughs, resulting in a lengthy treatment process and poor prognosis. Therefore, there is an urgent need to identify new biomarkers for the early diagnosis and treatment of sepsis to improve early detection, survival, and prognosis. Summary of the Invention

[0004] In view of the shortcomings of the existing technology, the purpose of the present invention is to provide the application of CCL28 as a biomarker and therapeutic target in sepsis, which solves the problems in the existing technology.

[0005] The purpose of the present invention can be achieved through the following technical solutions:

[0006] A first aspect of the present invention relates to the use of a detection reagent for the CCL28 gene or its expression level in the preparation of a kit for diagnosing or prognostic monitoring of sepsis.

[0007] Furthermore, the reagent is used to measure the expression level of mucosa-associated epithelial cell chemokine in a body fluid sample.

[0008] Furthermore, the method for detecting the expression level of the mucosa-associated epithelial cell chemokine by the reagent is at least one selected from the group consisting of chemiluminescence, ELISA, immunoturbidimetry, immunofluorescence, and colloidal gold method.

[0009] Furthermore, the body fluid sample is selected from at least one of serum, plasma, whole blood, urine, cerebrospinal fluid, pleural effusion, ascites, and joint fluid.

[0010] A second aspect of the present invention relates to a kit for diagnosing or prognostic monitoring of sepsis, comprising a detection reagent for the CCL28 gene or its expression level.

[0011] The third aspect of the present invention relates to the use of CCL28 recombinant protein, gene, enhancer and overexpression vector thereof in the preparation of a drug for treating sepsis.

[0012] The fourth aspect of the present invention relates to a drug for treating sepsis, comprising a recombinant protein of a mucosa-associated epithelial cell chemokine, a gene, a potentiator, and an overexpression vector thereof. Furthermore, the vector is a virus or liposome capable of carrying the recombinant protein of a mucosa-associated epithelial cell chemokine.

[0013] Furthermore, the drug has at least one of the following effects: (I) enhancing the body's ability to clear bacteria in sepsis; (II) alleviating multiple organ damage caused by sepsis; and (III) exerting an immune protective effect in sepsis.

[0014] Furthermore, the synergist is selected from agonists, upregulators or stabilizers.

[0015] Furthermore, the carrier is a virus or liposome capable of carrying the mucosa-associated epithelial cell chemokine recombinant protein.

[0016] Beneficial effects of the present invention:

[0017] 1) The present invention provides a mucosal-associated epithelial cell chemokine that can be used as a diagnostic indicator for sepsis.

[0018] 2) The present invention provides mucosal-associated epithelial cell chemokines that can be used as an indicator for evaluating the prognosis of sepsis.

[0019] 3) The present invention provides the use of mucosal-associated epithelial cell chemokine recombinant proteins, genes, enhancers and overexpression vectors thereof in the preparation of drugs for improving and / or treating sepsis, providing a new idea and option for the treatment of sepsis. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] 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, for ordinary technicians in this field, other drawings can be obtained based on these drawings without any creative work.

[0021] Figure 1 a in Figure 1 b in Figure 1 c in the figure respectively show the results of the detection of CCL28 expression in the peripheral blood of clinical adult sepsis patients and healthy subjects, the results of the detection of CCL28 expression in the peripheral blood of adult septic shock patients and non-septic shock patients, and the results of the detection of CCL28 expression in the peripheral blood of deceased adult sepsis patients and surviving adult sepsis patients in the embodiment of the present invention;

[0022] Figure 1 d in Figure 1 e in Figure 1 f in Figure 1 g in the figure respectively show the correlation analysis results of CCL28 expression in the peripheral blood of adult sepsis patients in the embodiment of the present invention and PCT, CRP, Creatine, and SOFA score;

[0023] Figure 2 a in Figure 2 b in Figure 2 Figures c in the figure respectively show the results of the detection of CCL28 expression in the peripheral blood of clinical pediatric sepsis patients and healthy subjects, the results of the detection of CCL28 expression in the peripheral blood of pediatric septic shock patients and non-septic shock patients, and the results of the detection of CCL28 expression in the peripheral blood of deceased pediatric sepsis patients and surviving adult sepsis patients in the embodiment of the present invention;

[0024] Figure 2 d in Figure 2 e in Figure 2 f in Figure 2 g in the figure respectively show the correlation analysis results of CCL28 expression in peripheral blood of children with sepsis in the embodiment of the present invention and PCT, CRP, Creatine, and SOFA score;

[0025] Figure 3 a in Figure 3b in the figure respectively shows a schematic diagram of the comparative analysis of the early diagnosis effect of CCL28 in adults and children for sepsis patients through ROC curve in an embodiment of the present invention;

[0026] Figure 4 a in Figure 4 b in the figure respectively shows a schematic diagram of the predictive effect of CCL28, PCT and CRP expression levels in adults and children on the 28-day mortality of sepsis patients by ROC curve comparison analysis in an embodiment of the present invention;

[0027] Figure 5 Shown is a diagram of the process of establishing a sepsis mouse model by cecal ligation and puncture (CLP) in an embodiment of the present invention;

[0028] Figure 6 Graph showing the experimental results of detecting the expression of CCL28 in blood, peritoneal lavage fluid (PLF), and lung in an animal sepsis model according to an embodiment of the present invention;

[0029] Figure 7 a in the figure is a statistical graph showing the survival rates of septic mice in the CCL28 protein-treated group and the PBS-treated group in the embodiment of the present invention;

[0030] Figure 7 b is a statistical graph of the survival rates of septic mice in the CCL28 antibody-treated group and the PBS control group;

[0031] Figure 8 Shown are graphs of bacterial loads in cardiac blood, spleen, and PLF of the CCL28 protein-treated group and the PBS-treated group in the examples of the present invention;

[0032] Figure 9 Shown is a graph showing the detection of inflammatory cell levels in PLF in the CCL28 protein-treated group and the PBS-treated group according to an embodiment of the present invention;

[0033] Figure 10 Shown are graphs showing liver / kidney function damage detection results for the CCL28 protein-treated group and the PBS-treated group in the examples of the present invention, as well as organ pathological sections of the liver, kidney, spleen, and lung. DETAILED DESCRIPTION

[0034] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts shall fall within the scope of protection of the present invention.

[0035] The present invention focuses on the application of CCL28 (Mucosae Associated Epithelia Chemokine, mucosa-associated epithelial cell chemokine, MEC) as a biomarker and therapeutic target in the diagnosis and treatment of sepsis. The human CCL28 encoding gene is located in the coding region of the short arm of chromosome 5, 5p12, with a length of 384bp. It encodes a precursor protein containing 127 amino acids, which is cleaved to form a mature protein of 105 amino acids. The mouse CCL28 gene is located at the distal end of chromosome 13, with a length of 393bp. It encodes a precursor protein of 130 amino acid residues, which can produce a mature protein of 108 amino acid residues after cleavage of the signal peptide. Mature human CCL28 is highly conserved with mouse CCL28 protein, with a homology of 83%, especially in the receptor binding domain (CCR10 / CCR3), that is, the N-terminal domain. Recent studies have confirmed that CCL28 is primarily produced by mucosal epithelial cells and has biological functions such as maintaining mucosal immune homeostasis, promoting trophoblast infiltration, and inducing apoptosis in decidual stromal cells. However, its application in sepsis has not been reported. The present invention, through a series of examples, preliminarily demonstrates that CCL28 is a novel immunomodulatory factor that can serve not only as an effective biomarker for early diagnosis and prognosis monitoring of sepsis, but also as a potential therapeutic target for sepsis. Therefore, the present invention provides a new technical approach for the diagnosis and treatment of sepsis.

[0036] Example 1

[0037] Detection of CCL28 in the blood of patients with sepsis and its correlation with SOFA score

[0038] Blood samples were collected from patients with sepsis treated at Zhongda Hospital, Southeast University, between January 2023 and December 2024 as the experimental group. Healthy subjects undergoing physical examinations during the same period served as the control group (their basic information is shown in Tables 1 and 2). An ELISA kit (purchased from Biolegend, Catalog Number: 441204) was used to detect CCL28 expression. Specific procedures were strictly followed according to the kit instructions. Statistical analysis was performed using SPSS 27.0 software, and graphics were generated using GraphPad Prism 10.0 software. Specific experimental details are as follows:

[0039] 1) Grouping of research subjects and establishment of inclusion / exclusion criteria: divided into sepsis group, septic shock group and healthy control group. Establishment of inclusion criteria: ① Sepsis patients: ICU hospitalized patients; aged 18 years or above; clinically diagnosed with sepsis, and the diagnostic criteria are in accordance with the "2021 International Guidelines for the Management of Sepsis and Septic Shock". ② Septic shock patients: ICU hospitalized patients; aged 18 years or above; clinically diagnosed with septic shock, and the diagnostic criteria are in accordance with the "2021 International Guidelines for the Management of Sepsis and Septic Shock", specifically manifested as refractory hypotension on the basis of sepsis, the need for continuous use of vasopressors to maintain a mean arterial pressure of 65 mmHg or above, and blood lactate >2 mmol / L. ③ Healthy controls: healthy physical examination subjects matched with the disease group in age and gender. Exclusion criteria: patients with malignant tumors; patients with severe blood system diseases, including but not limited to lymphoma, acute / chronic leukemia, aplastic anemia, etc.; patients with active autoimmune system diseases, including but not limited to systemic lupus erythematosus, rheumatoid arthritis, etc.; patients with chronic liver damage (score greater than 3 points) or chronic renal failure (stage 3 or above); pregnant women; patients with continuous hormone therapy (0.5 mg / kg) for more than 30 days.

[0040] 2) Sample data collection: The patient's personal information, medication status, and vital signs were recorded; the patient's acute physiology and chronic health evaluation (APACHE II) and sequential organ failure assessment (SOFA) were calculated; and the patient's 28-day survival status was recorded (as shown in Tables 1 and 2).

[0041] 3) Sample Collection and Processing: For patients in the experimental group who met the inclusion criteria, 2 mL of peripheral venous blood was collected on the first day after admission. For the control group, 2 mL of peripheral venous blood was collected on the day of physical examination. The blood was centrifuged within 1 hour (3500 rpm, 10 minutes), and the serum was separated and stored at -80°C.

[0042] 4) Sample Testing: Serum CCL28 protein expression levels were measured using ELISA. Samples were also sent to the laboratory for testing of laboratory parameters, including PCT, CRP, blood count, and liver / kidney function.

[0043] 5) Data Analysis: Nonparametric paired tests were used to compare the expression of CCL28 between patients with sepsis and healthy subjects, patients with septic shock and those without septic shock, and patients with sepsis who died and those who survived. Correlation analysis was used to determine the correlation between CCL28 and indicators such as the SOFA score. Receiver-operating characteristic (ROC) curves were used to calculate the area under the curve (AUC), sensitivity, and specificity. The cut-off value of CCL28 for diagnosing sepsis and predicting 28-day survival in patients with sepsis, as well as the corresponding Youden Index, were also calculated.

[0044] Table 1 Basic information statistics of adult sepsis patients and healthy subjects

[0045]

[0046]

[0047] Table 2 Basic information statistics of children with sepsis and healthy subjects

[0048]

[0049] The experimental results of this embodiment are as follows Figure 1 As shown in Figure 2, CCL28 expression is significantly increased in adult patients with sepsis ( Figure 1 In a), the expression level of CCL28 in the peripheral blood of patients with septic shock was significantly lower than that of patients with sepsis without shock ( Figure 1 b), and the CCL28 content in the peripheral blood of patients who died of sepsis was significantly lower than that of survivors ( Figure 1 c), with statistically significant differences. The CCL28 expression in children with sepsis was also significantly higher than that in normal children ( Figure 2 In a), the expression of CCL28 in peripheral blood of children with septic shock was significantly lower than that of children with sepsis without shock ( Figure 2 b), and the CCL28 content in children who died of sepsis was also significantly lower than that in survivors ( Figure 2 c in the figure). Further correlation analysis revealed that the expression of CCL28 in the peripheral blood of adult sepsis patients was negatively correlated with their PCT, CRP, Creatine, and SOFA scores ( Figure 1 dg in the dg), the CCL28 expression in the peripheral blood of children with sepsis was also negatively correlated with their PCT, CRP, Creatine, and SOFA scores ( Figure 2 These results suggest that CCL28 expression is elevated in sepsis and may become a novel diagnostic indicator for sepsis. Furthermore, the expression of CCL28 is negatively correlated with the severity of sepsis.

[0050] Example 2

[0051] ROC curve analysis was used to analyze the diagnostic efficacy of serum CCL28 for sepsis;

[0052] The ROC curve was drawn using SPSS27.0 software, and indicators such as the area under the ROC curve of CCL28 for diagnosing sepsis patients were analyzed.

[0053] Table 3 ROC curve analysis of the diagnostic efficacy of adult CCL28 for adult sepsis

[0054]

[0055] The results are as follows Figure 3 Figure a and Table 3 show that in adult sepsis, the area under the ROC curve of serum CCL28 for diagnosing sepsis is 0.924, which is statistically significant, indicating that CCL28 can be used as an effective early diagnostic indicator for sepsis.

[0056] Table 4 ROC curve analysis of the diagnostic efficacy of pediatric CCL28 for pediatric sepsis

[0057]

[0058] The results are as follows Figure 3 Figure b and Table 4 show that in children with sepsis, the area under the curve of serum CCL28 is 0.891, which is statistically significant, further indicating that CCL28 can be used to diagnose sepsis patients.

[0059] Example 3

[0060] ROC curve analysis was used to compare the prognostic evaluation efficacy of serum CCL28, PCT, and CRP in sepsis;

[0061] Subsequently, adult sepsis patients were divided into a survival group and a death group according to their 28-day survival status. The expression of CCL28 in their peripheral blood was detected by ELISA, and the levels of laboratory indicators PCT and CRP of sepsis patients were collected through the LIS system of the laboratory department. Then, the ROC curve was drawn using SPSS27.0 software, and the statistical differences of indicators such as the area under the ROC curve were analyzed and compared.

[0062] Table 5 ROC curve analysis of the prognostic efficacy of CCL28 / CRP / PCT in adult sepsis

[0063]

[0064] The results are as follows Figure 4 Figure a and Table 5 show that in adult sepsis, the areas under the ROC curves of serum CCL28, PCT, and CRP for predicting 28-day survival in sepsis were 0.814, 0.587, and 0.586, respectively. The area under the curve of CCL28 was significantly greater than that of PCT and CRP, which was statistically significant, indicating that CCL28 can be used to predict the prognosis of patients with sepsis.

[0065] Table 6 ROC curve analysis of the prognostic efficacy of CCL28 / CRP / PCT in children with sepsis

[0066]

[0067] The results are as follows Figure 4 Figure b and Table 6 show that in children with sepsis, the areas under the ROC curves of serum CCL28, PCT, and CRP for predicting 28-day survival in sepsis were 0.801, 0.521, and 0.517, respectively. The area under the curve of CCL28 was significantly larger than that of PCT and CRP, and was statistically significant, further indicating that CCL28 can be used to predict the prognosis of patients with sepsis.

[0068] Example 4

[0069] Cecal ligation and puncture (CLP) was used to establish an animal model of sepsis.

[0070] Experimental Animals: All mice used in this study were male, weighing 18-22 g and approximately 6-8 weeks old. They were SPF (Specific Pathogen Free)-grade and housed in the SPF laboratory at the Laboratory Animal Center of Southeast University. Wild-type C57BL / 6 mice (WT) were purchased from Jiangsu Jicui Pharmaceutical Biotechnology Co., Ltd.

[0071] Methods: Mice were anesthetized with 1.5% sodium pentobarbital intraperitoneally, fixed on an operating board, and their abdomens were shaved with pet electric clippers. The skin was disinfected and a 1 cm long incision was made in the middle of the abdomen. The cecum was ligated and punctured with a 26-gauge syringe needle. Figure 5 As shown. Finally, the wound is sutured and the skin is disinfected. (Specific reference: Daniel Rittirsch, Peter A Ward, et al. Immunodesign of experimental sepsis by cecal ligation and puncture. Nat Protoc. 2009; 4(1): 31–36). This modeling method is a classic modeling method for sepsis animal models and is currently the gold standard animal model for sepsis research.

[0072] Example 5

[0073] Detection of CCL28 levels in sepsis mouse models;

[0074] A mouse sepsis model was established using CLP (as described in Example 4). Cardiac blood, peritoneal lavage fluid (PLF), liver, and kidney samples were collected at 6 and 24 hours. CCL28 expression was measured using an ELISA kit (purchased from Solarbio, Catalog Number: SEKM-0045). Specific procedures were strictly followed according to the kit instructions. GraphPad Prism 10.0 software was used for statistical analysis and graphing.

[0075] like Figure 6 a in Figure 6 b in Figure 6 As shown in (c), compared with normal control mice, the expression levels of CCL28 in the peripheral blood, PLF, and lung of septic mice were significantly increased, with statistical differences.

[0076] The above experimental results also suggest that detecting the expression of CCL28 in peripheral blood, PLF, and lung may have potential applications in the diagnosis of sepsis.

[0077] Example 6

[0078] Survival rate experiment:

[0079] A mouse sepsis model (as described in Example 4) was established and divided into two groups (a control group and a recombinant CCL28 protein-treated group). The control group was intraperitoneally injected with 100 μL of sterile PBS buffer after model establishment, while the protein-treated group was intraperitoneally injected with 1 μg of recombinant CCL28 protein (purchased from Novus Biologicals, Catalog Number: NBP2-35169) in a volume of 100 μL after model establishment. The survival of the mice was observed and recorded twice daily until no more mice died, for 14 days.

[0080] The results are as follows Figure 7 As shown in a, treatment with exogenous CCL28 recombinant protein (1 μg / 100 μL, 1 μg CCL28 recombinant protein dissolved in 100 μL sterile PBS buffer, the same below) can increase the survival rate of septic mice from 12.5% ​​to 66%, suggesting that CCL28 recombinant protein is beneficial to the survival of septic mice.

[0081] To further verify this conclusion, in this example, CCL28 antibody (purchased from R&D Systems, Catalog Number: MAB533) was used to treat septic mice (2 μg / 100 μL, 2 μg CCL28 antibody was dissolved in 100 μL sterile PBS buffer) and a mouse sepsis model was established by CLP (as described in Example 4). The mice were divided into a control group and a CCL28 antibody-treated group. The control group was given an intraperitoneal injection of 100 μL sterile PBS after model establishment, and the survival of the mice was observed twice a day for 14 consecutive days.

[0082] The results are as follows Figure 7 As shown in b, compared with the wild-type septic mouse group, the survival rate of septic mice in the CCL28 antibody-treated group was significantly reduced, from 87.5% to 40%, which was statistically significant, further demonstrating that CCL28 plays an immune protective role in sepsis.

[0083] Example 7

[0084] Determination of bacterial load;

[0085] A mouse sepsis model was established by CLP (as described in Example 4). 24 hours later, the mice were anesthetized and fixed, and cardiac blood, PLF, and spleen were collected. The blood was then diluted 10-fold, the PLF was serially diluted 100-fold and 10,000-fold, and the spleen homogenate was diluted 10-fold. 100 μL of each diluted sample was plated and incubated at 37°C. The number of colonies on the blood agar plates was counted 18-24 hours later.

[0086] The results are as follows Figure 8 a in Figure 8 b in Figure 8 As shown in Figure c, compared with the control group, the bacterial load in the blood, PLF and spleen of septic mice in the CCL28 protein treatment group was significantly reduced, suggesting that CCL28 can enhance the body's bacterial clearance ability.

[0087] Example 8

[0088] Determination of the number of inflammatory cells in peritoneal lavage fluid;

[0089] A mouse sepsis model was established by CLP (as shown in Example 4). After 24 hours, the mice were anesthetized and fixed. PLFs were collected and aliquoted into 1 mL portions, centrifuged, washed once with sterile PBS, and resuspended in 1 mL of sterile PBS. The cells were counted using a Bovine Bowman's counting chamber.

[0090] The results are as follows Figure 9As shown, compared with wild-type septic mice, the content of inflammatory cells in PLF of septic mice in the CCL28 protein-treated group was significantly increased, and the difference was statistically significant, further indicating that CCL28 recombinant protein can enhance the body's bacterial clearance ability by increasing the recruitment of inflammatory cells.

[0091] Example 9

[0092] Biochemical and pathological examination of various organ damage

[0093] A mouse sepsis model was established by CLP (as described in Example 4). After modeling, the mice were intraperitoneally injected with 100 μL of sterile PBS buffer. 24 hours later, the mice were anesthetized and sacrificed. Cardiac blood was collected and centrifuged to obtain the supernatant. Liver / kidney injury-related indicators such as alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), creatine kinase (CK), and urea were measured. Simultaneously, the lungs, livers, spleens, and kidneys of the septic mice were removed and fixed in 4% paraformaldehyde for 24 to 48 hours. The samples were then dehydrated, paraffin-impregnated, embedded, and sectioned. The samples were then stained with H&E and examined under a microscope.

[0094] The results are as follows Figure 10 As shown in ae, compared with the control group, the levels of AST, ALT, LDH, CK and Urea in the serum of septic mice in the CCL28 protein treatment group were significantly decreased, with statistical differences, suggesting that CCL28 recombinant protein alleviated the multi-organ damage in septic mice ( Figure 10 ae in ); Pathological section results showed that the inflammation of the liver, kidney, lung and spleen of the septic mice in the CCL28 protein treatment group was significantly alleviated, and the manifestations such as inflammatory cell infiltration, protein inflammatory exudation, tissue congestion and edema, and cell necrosis were significantly improved ( Figure 10 f), further confirming that CCL28 recombinant protein can reduce multiple organ damage in sepsis.

[0095] In summary, the present invention found that the expression level of CCL28 in adult and pediatric sepsis patients was significantly upregulated compared with healthy subjects, and was negatively correlated with PCT, Creatine, CRP, and SOFA scores in sepsis patients, and can be used as a new early diagnostic biomarker for sepsis; and ROC curve analysis found that compared with PCT and CRP, CCL28 can effectively predict the 28-day survival rate of sepsis. Further in vivo animal experiments found that septic mice can cause increased expression levels of CCL28; treatment with exogenous CCL28 recombinant protein can significantly reduce bacterial load, liver / kidney and other organ damage in septic mice, and effectively improve the survival rate of septic mice. Therefore, the present invention provides CCL28 as a diagnostic, prognostic and monitoring indicator for sepsis, and provides CCL28 recombinant protein, gene, enhancer and overexpression vector thereof as new drugs for improving and / or treating sepsis.

[0096] It should be noted that, in order to specifically study the application of CCL28 and evaluate the diagnostic efficacy of CCL28, the detection results of the CCL28 encoding gene or its expression level are used as the indicators of interest in the examples of this application. However, this does not mean that CCL28 must be used as an independent marker for diagnosis when the technical solution of this application is actually implemented. To further improve the accuracy and specificity of diagnosis, those skilled in the art should understand that it is also acceptable to combine the sepsis diagnostic marker CCL28 of this application with sepsis biomarkers known in the prior art in this application.

[0097] Similarly, the CCL28 recombinant protein in the examples of the present application can be used as a new drug to improve and / or treat sepsis, but those skilled in the art should understand that genes, potentiators, and overexpression vectors thereof should also be able to achieve equivalent results. In actual drug applications, those skilled in the art should understand that the CCL28 recombinant protein of the present application can also be combined with other pharmaceutically acceptable excipients, or other commonly used sepsis treatment drugs in the art that can be used together to form a drug combination. In addition, in order to deliver CCL28 recombinant protein or genetic information encoding CCL28 recombinant protein, those skilled in the art should be able to conceive of constructing other commonly used vectors in medicine, such as plasmids, viruses, or liposomes.

[0098] Throughout this specification, references to terms such as "one embodiment," "example," or "specific example" indicate that the specific features, structures, materials, or characteristics described in conjunction with that embodiment or example are included in at least one embodiment or example of the present invention. In this specification, schematic representations of these terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.

[0099] The basic principles, main features, and advantages of the present invention are shown and described above. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are merely illustrative of the principles of the present invention. Various changes and modifications may be made to the present invention without departing from the spirit and scope of the present invention, and such changes and modifications fall within the scope of the invention as claimed.

Claims

1. Use of a detection reagent for the CCL28 gene or its expression level in the preparation of a kit for diagnosing or prognostic monitoring of sepsis.

2. Use of the detection reagent for the CCL28 gene or its expression level according to claim 1 in the preparation of a kit for diagnosing or prognostic monitoring of sepsis, characterized in that: The reagent is used to determine the expression level of CCL28 in a body fluid sample.

3. Use of the detection reagent for the CCL28 gene or its expression level according to claim 1 in the preparation of a kit for diagnosing or prognostic monitoring of sepsis, characterized in that: The method for detecting the expression level of CCL28 using the reagent is selected from at least one of chemiluminescence method, ELISA method, immunoturbidimetry, immunofluorescence method, and colloidal gold method.

4. Use of the detection reagent for the CCL28 gene or its expression level according to claim 2 in the preparation of a kit for diagnosing or prognostic monitoring of sepsis, characterized in that: The body fluid sample is selected from at least one of serum, plasma, whole blood, urine, cerebrospinal fluid, pleural effusion, ascites, and joint fluid.

5. A kit for diagnosing or prognostic monitoring of sepsis, comprising a detection reagent for the CCL28 gene or its expression level.

6. Application of CCL28 recombinant protein, gene, enhancer and its overexpression vector in the preparation of drugs for the treatment of sepsis.

7. The use according to claim 6, characterized in that: The drug has at least one of the following effects: (I) enhancing the body's ability to clear bacteria in sepsis; (II) alleviating multiple organ damage caused by sepsis; and (III) exerting an immune protective effect in sepsis.

8. A drug for treating sepsis, characterized in that: The invention comprises CCL28 recombinant protein, gene, enhancer and over-expression vector thereof.

9. The drug for treating sepsis according to claim 8, characterized in that: The potentiator is selected from an agonist, an upregulator or a stabilizer.

10. The drug for treating sepsis according to claim 8, characterized in that: The vector is a virus or liposome capable of carrying CCL28 recombinant protein.

Images