A method, system and kit for assisting in the assessment of the risk of a CTSL-based hyperthermic exposure-related neuroendocrine disorder

Abstract

The application discloses a kind of high heat exposure related neuroendocrine disorder risk auxiliary evaluation method, system and kit based on CTSL, the method includes sampling, detects CTSL level, analyzes result and obtains high heat exposure related neuroendocrine disorder risk information.The present application takes CTSL as core detection index, can be through the risk of high heat exposure related neuroendocrine disorder of ex vivo biological sample detection evaluation, it is helpful to improve the objectivity of high heat exposure related neuroendocrine disorder risk evaluation, it is favorable to early screening of high-risk population;The present application can further be combined with nervous system injury, inflammation reaction and neuroendocrine hormone related index, form risk score or risk stratification result, so as to the auxiliary evaluation and stratified management of high heat exposure related risk;The present application can also be extended to auxiliary evaluation system or kit, sample acquisition is relatively convenient, detection process is easy to standardize, with good application prospect.

Description

Technical Field

[0001] This invention relates to a method, system, and kit for auxiliary assessment of the risk of high heat exposure-related neuroendocrine disorders, and more particularly to a method, system, and kit for auxiliary assessment of the risk of high heat exposure-related neuroendocrine disorders based on CTSL. Background Technology

[0002] High heat exposure is a common risk factor in occupational health and public health management. With the increase in extreme heat weather, the health risks for people working in high-temperature environments and exposed to the outdoors continue to rise. Construction workers, sanitation workers, agricultural laborers, outdoor transport workers, military trainees, and those working in high-temperature workshops may all be exposed to high-heat environments for extended periods or repeatedly. High heat exposure can not only lead to acute heat injuries such as heatstroke and heat exhaustion, but may also cause abnormal thermoregulation, enhanced inflammatory responses, impaired central nervous system regulation, and neuroendocrine homeostasis imbalances, exhibiting a certain degree of insidiousness and cumulative effects.

[0003] Currently, risk assessment related to high heat exposure mainly relies on exposure history, changes in body temperature, clinical symptoms, routine biochemical indicators, and signs of acute heat injury. These methods are helpful for individuals who have already shown obvious signs of heatstroke or heat exhaustion, but they are insufficient for identifying the risks of early central nervous system dysfunction, increased inflammation, and neuroendocrine disorders caused by continuous or repeated high heat exposure. Especially when symptoms are not yet typical, body temperature has recovered, or routine indicators do not change significantly, existing methods are unable to provide objective, quantitative, and proactive auxiliary assessment results, and are also not conducive to early screening and stratified management of high-risk groups.

[0004] Therefore, it is necessary to establish a method that can effectively assess the risk of early central nervous system dysfunction, increased inflammation, and neuroendocrine disorders caused by continuous or repeated exposure to high heat, in order to carry out further risk stratification, health monitoring, and management of occupationally exposed populations. Summary of the Invention

[0005] Objective of the Invention: The objective of this invention is to provide a method, system, and kit for auxiliary assessment of the risk of neuroendocrine disorders related to high heat exposure, using cathepsin L (CTSL) as a biomarker. The method can obtain risk assessment information based on CTSL levels, addressing the problem of insufficient objectivity in early identification of existing assessment methods.

[0006] Technical solution: This invention provides the application of CTSL in assessing the risk of neuroendocrine disorders related to high heat exposure.

[0007] This invention screened CTSL as a candidate molecule associated with abnormal hypothalamic regulation after high-heat exposure by jointly analyzing the transcriptome and lipidome of the hypothalamus in a high-heat exposure model. Based on this, consistency verification was performed in the hypothalamus, cerebrospinal fluid, and serum of pig and mouse models. It was found that CTSL mRNA expression was decreased in the hypothalamus tissue of the high-heat exposure group, and CTSL protein levels were reduced in cerebrospinal fluid and serum, indicating that using CTSL as an auxiliary indicator for risk assessment of high-heat exposure-related neuroendocrine disorders has good biological basis and detection stability.

[0008] This invention also provides an auxiliary assessment method for the risk of high-heat exposure-related neuroendocrine disorders based on CTSL, comprising the following steps:

[0009] (1) Obtaining samples, including serum, plasma, whole blood, cerebrospinal fluid or tissue;

[0010] (2) Detect CTSL level;

[0011] (3) Analyze the test results;

[0012] (4) Obtain information on the risk of neuroendocrine disorders related to high heat exposure.

[0013] In step (1), the sample is preferably serum, plasma, or whole blood. Tissue samples can be used for CTSL expression validation or model construction.

[0014] In step (2), the CTSL level is preferably the gene transcription level or protein expression level of CTSL. For serum, plasma, whole blood or cerebrospinal fluid samples, ELISA, chemiluminescent immunoassay, immunoturbidimetry or mass spectrometry are preferred to detect the CTSL protein level; for tissue samples, qPCR, Western blot or immunoassay methods can be used to detect CTSL expression.

[0015] In step (3), the analysis preferably compares the CTSL test results with a preset reference value, a control level, or a high heat exposure stratification reference value, or inputs the CTSL test results into a risk assessment model. The reference values ​​can be derived from healthy control samples, non-high heat exposure samples, similar model control samples, or historical test data. When the CTSL level is lower than the preset reference value or control level, it indicates an increased risk of high heat exposure-related neuroendocrine disorders.

[0016] Furthermore, the risk assessment model is established using threshold comparison, scoring model, logistic regression model, ROC curve analysis model or machine learning model. The input indicators of the model include at least the CTSL level, and the output results are risk score, risk probability or risk stratification results.

[0017] In step (4), the information preferably includes risk score, risk probability, low / medium / high risk stratification results or risk threshold prompt information.

[0018] Furthermore, the method also includes detecting combined indicators, which include indicators of nervous system damage, inflammatory markers, and / or neuroendocrine hormones.

[0019] The neurological injury indicators preferably include NSE and / or S100β, the inflammatory indicators include TNF-α, IL-1β and / or IL-6, and the neuroendocrine hormone indicators include ACTH, cortisol, GnRH, LH, FSH and / or testosterone.

[0020] This invention also provides a risk-assisted assessment system for high-heat exposure-related neuroendocrine disorders based on CTSL (Continuous Transmission of Suffering and Respiratory Disorders). The system includes a data input module, a data processing module, and a result output module. The data input module receives CTSL test results. The data processing module includes a reference value and threshold management module and a risk calculation module. The result output module outputs the risk-assisted assessment results. Preferably, the data input module also receives the test results of the aforementioned combined indicators.

[0021] The present invention also provides a kit comprising reagents for detecting CTSL, the detection results being used for auxiliary assessment of the risk of high heat exposure-related neuroendocrine disorders. Preferably, the kit further comprises reagents for the aforementioned combined indicators.

[0022] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0023] 1. This invention uses CTSL as the core detection indicator, which can assess the risk of high heat exposure-related neuroendocrine disorders through in vitro biological sample detection, avoiding reliance solely on symptoms, body temperature, and routine biochemical indicators. This helps improve the objectivity of the risk assessment of high heat exposure-related neuroendocrine disorders and is beneficial for early screening of high-risk groups.

[0024] 2. This invention can further combine indicators related to nervous system injury, inflammatory response and neuroendocrine hormones to form risk scores or risk stratification results, so as to assist in the assessment and stratified management of risks related to high heat exposure;

[0025] 3. This invention can be expanded into an auxiliary evaluation system or kit. Sample acquisition is relatively convenient, and the detection process is easy to standardize, which has good application prospects. Attached Figure Description

[0026] Figure 1 Flowchart of a CTSL-based method for assisting in the risk assessment of neuroendocrine disorders associated with high-temperature exposure;

[0027] Figure 2 Schematic diagram for joint screening of candidate biomarkers for heat exposure-related neuroendocrine disorders using the transcriptome and lipidome of the hypothalamus in a porcine heat exposure model (A: Schematic diagram of hypothalamic lipidome detection in a porcine heat exposure model; B: Results of differential molecular analysis of the lipidome; C: Classification results of differential lipids; D: Schematic diagram of hypothalamic transcriptome detection in a porcine heat exposure model; E: Results of differential gene analysis of the transcriptome; F: Correlation distribution of differential molecules between the transcriptome and lipidome; G: Heatmap of correlation between differential genes and differential lipids; H: Statistics on the number of shared pathways between the transcriptome and lipidome; I: Information on shared KEGG pathways; J: Schematic diagram of the localization of CTSL candidate molecules in autophagy-related pathways).

[0028] Figure 3 Schematic diagrams showing the detection results of CTSL changes in the hypothalamus, cerebrospinal fluid, and serum in pig and mouse hyperthermia exposure models (A: pig hyperthermia exposure model and sample detection process; B: detection results of CTSL mRNA expression in pig hypothalamus; C: detection results of CTSL levels in pig cerebrospinal fluid; D: detection results of CTSL levels in pig serum; E: mouse hyperthermia exposure model and sample detection process; F: detection results of CTSL mRNA expression in mouse hypothalamus; G: detection results of CTSL levels in mouse cerebrospinal fluid; H: detection results of CTSL levels in mouse serum).

[0029] Figure 4 Schematic diagrams illustrating the relationships between CTSL and inflammatory markers, neuroendocrine hormone markers, and related phenotypes at different sample levels in pig and mouse models (A: Correlation heatmap between CTSL and neuroendocrine and inflammatory markers at different sample levels in pigs and mice; B: Comprehensive trend analysis of serum CTSL and related serum neuroendocrine markers in pigs; C: Comprehensive trend analysis of serum CTSL and related serum inflammatory markers in pigs; D: Comprehensive trend analysis of serum CTSL and related serum neuroendocrine markers in mice; E: Comprehensive trend analysis of serum CTSL and related serum inflammatory markers in mice).

[0030] Figure 5 This is a schematic diagram of a CTSL-based risk assessment system for neuroendocrine disorders related to high-temperature exposure.

[0031] Abbreviations: HS represents high heat exposure or heat stress, Con represents control, CSF represents cerebrospinal fluid, CTSL represents cathepsin L, GnRH represents gonadotropin-releasing hormone, LH represents luteinizing hormone, FSH represents follicle-stimulating hormone, NSE represents neuron-specific enolase, TNF-α represents tumor necrosis factor α, IL-1β represents interleukin-1β, and IL-6 represents interleukin-6; in the figure, r represents the Spearman correlation coefficient, and the asterisk indicates statistical significance. Detailed Implementation

[0032] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0033] Example 1

[0034] This embodiment illustrates the screening process for CTSL as an auxiliary indicator for assessing the risk of high-heat exposure-related neuroendocrine disorders.

[0035] A porcine heat exposure model was established, and hypothalamic tissue samples were collected from both the control and heat-exposed groups. Transcriptome sequencing and lipidome analysis were performed on these samples to identify differentially expressed genes and lipid molecules in the hypothalamus after heat exposure. Subsequently, the transcriptome and lipidome results were jointly analyzed, including differential molecule screening, correlation analysis, pathway enrichment analysis, and key pathway localization analysis.

[0036] The results showed that the porcine hypothalamus exhibited significant transcriptional and lipid metabolism abnormalities after high-heat exposure, with differentially expressed molecules mainly involved in autophagy, lysosomal function, and cellular homeostasis regulation. Combining pathway enrichment results and candidate molecule localization analysis, CTSL was identified as a candidate molecule associated with abnormal hypothalamic regulation after high-heat exposure.

[0037] This embodiment illustrates that CTSL is derived from the combined screening results of the hypothalamic transcriptome and lipidome of the high heat exposure model, and can be used as the core detection indicator in the risk-assisted assessment method of this invention.

[0038] Example 2

[0039] This example illustrates the changes in CTSL in tissue and body fluid samples after high-temperature exposure, serving as further verification of the screening results of Example 1.

[0040] High-heat exposure models were established in pigs and mice, respectively. Hypothalamic tissue, cerebrospinal fluid, and serum samples were collected from animals in the control and high-heat exposure groups. The expression levels of CTSL or Ctsl mRNA in hypothalamic tissue were detected by qPCR, and the CTSL protein levels in cerebrospinal fluid and serum were detected by ELISA or other immunoassay methods.

[0041] The results showed that, compared with the control group, the expression of CTSL mRNA in the hypothalamus of pigs exposed to high heat was decreased, and the levels of CTSL protein in cerebrospinal fluid and serum were reduced. Consistent changes were also observed in the mouse model, namely, the expression of CTSL mRNA in the hypothalamus decreased and the levels of CTSL in cerebrospinal fluid and serum were reduced after high heat exposure.

[0042] This embodiment demonstrates that CTSL shows a consistent decrease in the hypothalamus, cerebrospinal fluid, and serum after high-temperature exposure, exhibiting cross-model stability and a basis for body fluid detection.

[0043] Example 3

[0044] This example illustrates the relationship between CTSL and high-heat exposure-related inflammatory responses, neuroendocrine abnormalities, and related phenotypes.

[0045] In pig and mouse models of high heat exposure, serum CTSL levels, inflammatory markers, neuroendocrine hormone markers, and related phenotypic indicators were measured. The inflammatory markers included one or more of TNF-α, IL-1β, and IL-6; the neuroendocrine hormone markers included one or more of GnRH, LH, FSH, testosterone, ACTH, and cortisol. Correlation analysis was performed on these indicators.

[0046] The results showed that in both pig and mouse models, CTSL levels were generally positively correlated with neuroendocrine-related indicators such as GnRH, LH, FSH, and testosterone, and generally negatively correlated with inflammatory indicators such as TNF-α and IL-1β. These results suggest that changes in CTSL are related to increased inflammation, hormonal abnormalities, and neuroendocrine homeostasis imbalance after high-temperature exposure.

[0047] This embodiment illustrates that CTSL not only undergoes stable changes after high heat exposure, but also is associated with abnormal phenotypes related to high heat exposure, and can be used as a core detection indicator in risk-assisted assessment.

[0048] Example 4

[0049] This implementation provides an auxiliary assessment method for the risk of high-heat exposure-related neuroendocrine disorders based on CTSL, such as... Figure 1 As shown, it includes the following steps:

[0050] Step 1: Obtain in vitro biological samples from high-heat exposure model animals or test subjects. The in vitro biological samples include serum, plasma, whole blood, cerebrospinal fluid, or hypothalamic tissue.

[0051] Step 2: Detect the expression level or protein content of CTSL in the ex vivo biological samples. For serum, plasma, whole blood, or cerebrospinal fluid samples, ELISA is used to detect CTSL protein levels; for tissue samples, qPCR is used to detect CTSL expression.

[0052] Step 3: Compare the CTSL test results with the preset reference value. When the CTSL level is lower than the preset reference value or the control level, it indicates an increased risk of neuroendocrine disorders related to high heat exposure.

[0053] Step 4: Obtain auxiliary assessment information on the risk of high heat exposure-related neuroendocrine disorders based on the comparison results. This auxiliary assessment information includes risk score, risk probability, low / medium / high risk stratification results, or risk threshold information.

[0054] In some embodiments, the method further includes detecting a combination of indicators. These indicators include one or more of neurological damage indicators, inflammatory indicators, and neuroendocrine hormone indicators. The neurological damage indicators include one or more of NSE and S100β; the inflammatory indicators include one or more of TNF-α, IL-1β, and IL-6; and the neuroendocrine hormone indicators include one or more of ACTH, cortisol, GnRH, LH, FSH, and testosterone. These combined indicators can be used in conjunction with CTSL for risk scoring, risk probability calculation, or risk stratification, and can be used as optional inputs to improve the stability or stratification capability of risk assessment.

[0055] In some implementations, the risk assessment model may be established using threshold comparison, scoring models, logistic regression models, or ROC threshold models; when the sample size meets the modeling requirements, machine learning models may also be used. The model input indicators include at least the CTSL level, and may further include one or more of the combined indicators. The model output is an auxiliary assessment result for the risk of neuroendocrine disorders related to high heat exposure.

[0056] This embodiment illustrates that by detecting the CTSL level in isolated biological samples and analyzing it in conjunction with reference values ​​or risk models, auxiliary assessment information on the risk of high heat exposure-related neuroendocrine disorders can be obtained.

[0057] Example 5

[0058] This embodiment provides a risk-assisted assessment system for implementing the method of the present invention.

[0059] like Figure 5 As shown, the system includes a detection input terminal, a risk-assisted assessment system, and an application output terminal. The risk-assisted assessment system includes a data receiving module, a reference value and threshold management module, a risk calculation module, and a result generation module.

[0060] The data receiving module is used to receive CTSL test results, optional joint indicator test results, sample information, and exposure stratification information. The sample information includes sample type, test time, or sample number; the exposure stratification information includes one or more of the following: high heat exposure status, exposure frequency, exposure duration, or exposure type.

[0061] The reference value and threshold management module is used to store and retrieve CTSL reference values, high heat exposure stratification reference values, joint indicator reference ranges, and risk threshold libraries. The reference values ​​can be derived from control samples, historical test data, model animal control data, or preset risk stratification data.

[0062] The risk calculation module is used to standardize the input data, compare reference values, determine thresholds, calculate risk scores, and invoke models. The models include scoring models, logistic regression models, and ROC threshold models; when the sample size meets the modeling requirements, machine learning models may also be included.

[0063] The result generation module is used to generate risk-assisted assessment results. The results include risk scores, risk probabilities, low / medium / high risk stratification results, or risk threshold prompts.

[0064] The application output is used to record and display risk-assisted assessment results, risk stratification records, health monitoring records, and risk management reference information.

[0065] In one implementation, the CTSL test result of the sample to be tested is input into the data receiving module; the system calls the CTSL reference value or risk threshold in the reference value and threshold management module; the risk calculation module completes the comparison and scoring; and the result generation module outputs the risk auxiliary assessment result. If the joint indicator test result is input at the same time, the system can generate a risk score or risk stratification result based on the CTSL and the joint indicator.

[0066] This embodiment illustrates that the present invention can realize the reception, processing, calculation and output of CTSL detection data through a modular system, thereby completing the auxiliary assessment of the risk of high heat exposure-related neuroendocrine disorders.

[0067] Example 6

[0068] This embodiment provides a detection kit for implementing the method described in this invention.

[0069] The kit includes detection reagents for detecting CTSL. These reagents include one or more of the following: CTSL-specific antibody, standards, calibrators, quality controls, reaction buffer, washing buffer, chromogenic substrate, luminescent substrate, stop solution, or instructions for use. The kit can be used with ELISA, chemiluminescent immunoassay, immunoturbidimetry, or other immunoassay platforms.

[0070] In one embodiment, the kit is an ELISA detection kit, comprising an enzyme-labeled plate coated with CTSL capture antibody, CTSL standards, sample diluent, detection antibody, enzyme-labeled secondary antibody, washing buffer, chromogenic solution, stop solution, and instructions for use. In use, the ex vivo sample to be tested is added to the wells of the enzyme-labeled plate to bind with the CTSL capture antibody; after washing, the detection antibody and enzyme-labeled secondary antibody are added; then the chromogenic solution is added to initiate the reaction; finally, the stop solution is added and the absorbance value is read. The CTSL content in the sample is calculated based on a standard curve.

[0071] In another embodiment, the kit is a chemiluminescence detection kit, comprising a CTSL capture antibody, a CTSL detection antibody, magnetic microparticles, a luminescent substrate, calibrators, quality control materials, a reaction buffer, and instructions. In use, the sample is reacted with the antibody reagent to form an immune complex, washed, and then the luminescent substrate is added. The luminescence signal is read, and the CTSL concentration is calculated.

[0072] In some embodiments, the kit further includes reagents for detecting a combination of indicators. These indicators include one or more of NSE, S100β, IL-1β, IL-6, TNF-α, ACTH, cortisol, GnRH, LH, FSH, and testosterone.

[0073] This embodiment illustrates that CTSL can be detected using a standardized test kit, which can be used to implement the risk-assisted assessment method of the present invention.

Claims

1. Application of CTSL in assessing the risk of neuroendocrine disorders associated with high heat exposure.

2. A method for assisting the evaluation of the risk of a CTSL-based hyperthermic exposure-related neuroendocrine disorder, characterized in that, Includes the following steps: (1) Obtaining samples, including serum, plasma, whole blood, cerebrospinal fluid or tissue; (2) Detect CTSL level; (3) Analyze the test results; (4) Obtain information on the risk of neuroendocrine disorders related to high heat exposure.

3. The method of claim 2, wherein, In step (2), the CTSL level refers to the gene transcription level or protein expression level of CTSL.

4. The method of claim 2, wherein, In step (3), the analysis involves comparing the CTSL test results with preset reference values, control levels, or high heat exposure stratification reference values, or inputting the CTSL test results into a risk assessment model.

5. The method according to claim 4, characterized in that, The risk assessment model is established using threshold comparison, scoring model, logistic regression model, ROC curve analysis model or machine learning model. The input indicators of the model include CTSL level, and the output results are risk score, risk probability or risk stratification results.

6. The method according to claim 2, characterized in that, In step (4), the information includes risk score, risk probability, low / medium / high risk stratification results or risk threshold prompts.

7. The method according to claim 2, characterized in that, The method also includes detecting combined indicators, which include indicators of nervous system damage, inflammation, and / or neuroendocrine hormones.

8. The method according to claim 7, characterized in that, The neurological damage indicators include NSE and / or S100β, the inflammatory indicators include TNF-α, IL-1β and / or IL-6, and the neuroendocrine hormone indicators include ACTH, cortisol, GnRH, LH, FSH and / or testosterone.

9. A risk assessment system for high-heat exposure-related neuroendocrine disorders based on CTSL, characterized in that, The system includes a data input module, a data processing module, and a result output module. The data input module is used to receive CTSL detection results. The data processing module includes a reference value and threshold management module and a risk calculation module. The result output module is used to output risk auxiliary assessment results.

10. A reagent kit, characterized in that, The kit includes reagents for detecting CTSL, and the results are used for auxiliary assessment of the risk of high heat exposure-related neuroendocrine disorders.

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