Abstract

Objective

   To investigate the correlation between the expression of
   lipopolysaccharide-binding protein (LBP) in peripheral blood of spinal
   tuberculosis and clinical diagnosis and to evaluate its value as a
   diagnostic marker of spinal tuberculosis.

Methods

   In the experimental group, clinical history data and peripheral blood
   were collected from 100 patients with spinal tuberculosis who were
   admitted to the Department of Spine Surgery, General Hospital of
   Ningxia Medical University from May 2017 to May 2020, and peripheral
   blood was collected from 30 healthy volunteers in the control group.
   Screening of differential LBP expression by proteomics and ELISA to
   verify its expression in peripheral blood of spinal tuberculosis
   patients. t-test, Spearman analysis, linear regression and ROC curve
   were used to evaluate the diagnostic value of LBP in peripheral blood
   for spinal tuberculosis.

Results

   The expression of LBP protein in peripheral blood is significantly
   higher in patients with spinal tuberculosis than in the normal
   population; LBP assay values were significantly and positively
   correlated with CRP and ESR values (P < 0.01); the AUC of LBP in the
   diagnosis of spinal tuberculosis for pathological examination,
   bacteriological culture, T-cell spot test for tuberculosis infection
   (T-SPOT), imaging diagnosis, and acid fast bacillus were, respectively,
   0.677 (P < 0.01), 0.707 (P < 0.01), 0.751 (P < 0.01), 0.714 (P < 0.01),
   and 0.656 (P < 0.05), and there was a correlation between LBP and the
   diagnostic evaluation of spinal tuberculosis.

Conclusion

   LBP could be a new candidate biomarker for the diagnosis of spinal
   tuberculosis.

   Keywords: spinal tuberculosis, proteomics, LBP, biomarker

Introduction

   Tuberculosis due to infection by the bacterial pathogen Mycobacterium
   tuberculosis represents one of the ten most potent killers and the
   deadliest disease due to a single pathogen, more than HIV/AIDS,
   totaling 5.8 million newly diagnosed cases and approximating 1.3
   million deaths in 2020.[42]^1 Spinal tuberculosis (STB) is often
   secondary to pulmonary tuberculosis and is the most common bone and
   joint tuberculosis, accounting for approximately 50% of all bone and
   joint tuberculosis, which predisposes to serious complications such as
   spinal cord and nerve damage.[43]^2 The early clinical symptoms of STB
   patients are atypical, the onset is insidious, and the duration of the
   disease is long, which makes it difficult to confirm the diagnosis and
   makes it easy to be misdiagnosed and missed. Early diagnosis of the
   disease can reduce the risk of pain and complications. Therefore, it is
   of great significance to explore molecular markers with high
   specificity and sensitivity for the clinical diagnosis and treatment of
   STB disease. Lipopolysaccharide-binding protein (LBP) is a 60kDa
   molecular protein that belongs to type I reactive proteins and plays an
   important significant role in infectious diseases or immune
   responses.[44]^3 The literature has confirmed that LBP is expressed
   directly in human peripheral hematogenous monocytes, interacting with
   human macrophages and monocytes.[45]^4 Its specific and stable
   expression in cardiovascular and neoplastic diseases is expected to be
   the potential for detecting molecular markers of diseases.[46]^5–7
   However, the potential capabilities of LBP are not yet clear. In this
   study, we screened and validated the differential expression of LBP as
   a molecular marker in the peripheral blood of STB patients and healthy
   subjects by proteomics and analyzed the correlation of its clinical
   characteristics to evaluate the potential of LBP as a diagnostic
   biomarker for STB, hoping to provide reference and ideas for the
   diagnosis and treatment of STB.

Materials and Methods

Clinical Data

   From May 2017 to May 2020, the Department of Spine orthopedics of
   Ningxia Medical University admitted 100 exceptions to the peripheral
   blood and clinical data of patients with spinal tuberculosis were
   collected for the experimental group and 30 cases of healthy examinees,
   of which 50 cases of males and 50 cases of females in the experimental
   group, aged 18–77 years old, the average age (49.47±16.32 years old),
   the control group of 13 cases of males, 17 cases of females, age 40–72
   years old, the average age (53.39±9.67 years old). There was no
   statistical significance in the comparison of sex and age in both
   groups (P>0.05).

   All experiments involving human blood samples were conducted in
   accordance with the Declaration of Helsinki. The experimental protocol
   of this study was reviewed and approved by the Ethics Committee of the
   General Hospital of Ningxia Medical University.

   Inclusion criteria for patients with spinal tuberculosis:[47]^8 (1)
   Clinical symptoms (such as: night sweats, back pain, fatigue, etc),
   laboratory and imaging tests are comprehensively diagnosed as patients
   with spinal tuberculosis; (2) Patients with spinal tuberculosis who
   have not been treated with anti-tuberculosis before surgery; (3)
   Patients with spinal tuberculosis who have not recently taken hormones
   or immunosuppressant drugs.

   Criteria for exclusion in patients with spinal tuberculosis:[48]^8 (1)
   Patients with diabetes, malignant tumors and other infectious diseases;
   (2) Patients who may cause changes in blood routine values due to
   recent medication; (3) Women in menstrual cycles; (4) Patients with
   purulent infections and lumbar spine lesions such as Brucella; (5)
   Participants with incomplete information or refusal to participate in
   research and research.

   Inclusion criteria for healthy people: (1) Previously healthy, no
   chronic diseases such as hypertension, no history of diabetes,
   malignant tumors, tuberculosis, and no immune activating or inhibitory
   drugs have been taken in the past 1 month; (2) There is no history of
   viral and bacterial infections in the past two weeks; (3) There is no
   abnormal performance in physical examination, and routine blood tests
   show that macrophages, monocytes, granulocytes and other indicators are
   within the normal range; (4) The reagents and methods used in the blood
   routine of the experimental group and the control group are consistent.

   Criteria for exclusion of healthy people: (1) The control group has
   active pulmonary tuberculosis, diabetes, malignant tumors and
   infectious diseases; (2) Patients who may have recent medications that
   may cause changes in blood routine values; (3) Women in the menstrual
   cycle; (4) immunodeficiency diseases or other immune system diseases;
   (5) Participants have incomplete information or refuse to participate
   in surveys.

Clinical Data Collection of Patients

   We collected basic clinical data of STB patients and treatment related
   to STB diagnosis, including name, sex, age, course of disease,
   laboratory tests (blood routine, CRP, ESR, etc), bacterial culture,
   pathological examination, T-SPOT, imaging results and other examination
   results.

Sample Preparation

   All participants received fasting peripheral blood samples in the early
   morning and were stored in a vacuum anticoagulant tube. Then, the
   collected peripheral blood samples were centrifuged and packed in the
   laboratory within 1h. The centrifuge conditions were as follows: at 4℃
   for 3000 RPM/min for 10 minutes. After completion, the upper serum was
   taken and placed in a 1.5mLEP tube and stored at −80℃.

Proteomics Detection and Analysis

   Three pairs of peripheral blood of STB patients and healthy persons
   were selected as experimental groups and control groups, and Tandem
   Mass Tag quantitative proteomics detection was performed using the
   high-resolution mass spectrometer Q-Exactive plus, and the differential
   protein data were analyzed using mass spectrometry analysis software
   Mascot 2.6 and Proteome Discoverer 2.2 after mass spectrometry scanning
   to determine the unilateral edge of the preliminary screening
   differences. The preliminary filter criteria are: difference multiples
   ≥1.5 times, using Cluster 3.1, GraphPad Prism 8.0.1 software to map
   differential protein clusters and volcanoes.

Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG)
Signaling Pathway Analyses

   Metascope, a web-base resource ([49]http://metascope.org) for gene and
   protein annotation, visualization, and integration discovery, was used
   to perform GO analyses. The KEGG (KEGG Orthology And Links Annotation)
   online analysis database ([50]http://www.kegg.jp/blastkqalal) was used
   to perform KEGG pathway analyses.

ELISA Validation of LBP Differential Proteins

   The relevant ELISA experiments should be started after the collection
   of the number of samples in the experimental combination control group
   is completed. We selected 50 STB patients and 30 health examiners for
   peripheral blood to verify the differences in LBP. The ELISA kit (LBP
   antibody kit, Beijing Xinbosheng Technology Biotechnology Co, Ltd.)
   were used to measure LBP concentrations within the range of the
   standard curve. A standard curve was constructed according to the
   manufacturer’s instructions. The measurement range for the LBP kit was
   15–350 pmol/L.

Statistical Methods

   SPSS26.0 software and GraphPad Prism Version 8.0.1 software were used
   for statistical analysis and processing. Measurement data were
   expressed as X ± S; t-test was used to compare the data between the two
   groups. Curvilinear regression and Spearman test were used to analyze
   the correlation between LBP and other counts. ROC curve was used to
   evaluate the specificity and sensitivity of LBP in the diagnosis of
   spinal tuberculosis. The area under curve (AUC) ≥0.6 was considered to
   be of diagnostic value. Inspection level P<0.05 was considered
   statistically significant.

Results

Screening and Validation of LBP Proteins

   Under the preliminary proteomic screening, a total of 16 differentially
   expressed proteins were found in STB patients and healthy peripheral
   blood, of which 11 were upregulated differential proteins and 5 were
   expressed in STB peripheral blood. According to cluster analysis heat
   maps, advanced volcano maps show differential protein expression in STB
   and healthy peripheral blood ([51]Figures 1A and [52]B). Expanded
   filter criteria: the difference multiple ≥1.5, P < 0.01, the associated
   protein was related to the STB signaling pathway; we used ELISA to
   verify LBP expression in peripheral blood between STB patients and
   healthy people, and the results confirmed that LBP does have
   significant differences in the peripheral blood of STB patients and
   meets the screening criteria. ELISA verifies that LBP differential
   protein expression differs between healthy and STB peripheral blood
   ([53]Figure 1C). The ROC curve shows: LBP specificity and sensitivity
   in the peripheral blood of STB patients, AUC of 0.880 (P < 0.01)
   indicates that LBP has the potential to detect STB markers ([54]Figure
   1D).

Figure 1.

   [55]Figure 1
   [56]Open in a new tab

   Validation of LBP protein screening in peripheral blood of normal human
   and STB patients.

   Notes: (A) Differential protein clustering analysis graph. (B)
   Differential protein volcano distribution. (C) ELISA validation of
   differential protein LBP result analysis (***P < 0.001, the difference
   was statistically significant; Normal: healthy people). (D) ROC curve
   results of LBP in STB examination.

   Abbreviations: FC, fold change; STB, spinal tuberculosis; LBP,
   lipopolysaccharide-binding protein; AUC, area under the curve.

Results of LBP Analysis in GO and KEGG

   GO and KEGG analyses are important methods and tools in the field of
   bioinformatics for analyzing the functional enrichment of
   differentially expressed proteins. Thus, we used these analyses to
   identify differences between children with vs without MPP. In [57]Table
   1, the GO functional enrichment analysis of the STB group versus normal
   controls included, among others, cellular process of GO, biological
   regulation, localization, metabolic process and immune response. In
   [58]Table 2, the KEGG pathway enrichment analysis showed that among
   others, NF-κB signaling pathway, TLRs signaling pathway and Apoptosis
   may play important roles in the development of STB. These results
   indicated that multiple functions and signaling pathways of cells
   participate in the occurrence or development of STB. Meanwhile,
   significantly different proteins from the screening were associated
   with TLRs and NF-κB signaling pathways ([59]Figure 2A and [60]B).

Table 1.

   Results of GO Functional Annotation Analyses
    GO Term                    Description                  Input Number P value
   GO:0035662 Extracellular space                           16           0.001
   GO:0044548 Localization                                  5            0.001
   GO:0050544 Response to stimulus                          14           0.001
   GO:0050786 Biological Regulation                         13           0.001
   GO:0006805 Cellular process                              13           0.001
   GO:0006914 Locomotion                                    16           0.001
   GO:0006919 Multi-organism process                        15           0.001
   GO:0032119 Multicellular organismal process              9            0.001
   GO:0044057 Developmental process                         9            0.001
   GO:0051000 Growth                                        5            0.001
   GO:0051493 Metabolic process                             6            0.001
   GO:0050729 Cell killing                                  1            0.001
   GO:0001726 Signaling                                     6            0.001
   GO:0016529 Cellular component organization or biogenesis 11           0.001
   GO:0030017 Reproductive process                          11           0.001
   GO:0008017 Reproduction                                  10           0.001
   GO:0002224 Leukocyte migration                           13           0.001
   GO:0030307 Cell growth                                   15           0.001
   GO:0050832 Immune response                               8            0.001
   [61]Open in a new tab

Table 2.

   Results of KEGG Signaling Pathway Analyses
   Protein ID Definition P value Signaling Pathway
   [62]P18428 Lipopolysaccharide-binding protein 0.001 NF-kappaB Toll-like
   receptor
   [63]P05109 Protein S100-A8 0.015 IL-17
   [64]P06702 Protein S100-A9 0.044 IL-17
   [65]P05156 Complement factor I 0.023 NF-kappaB
   [66]P01009 Alpha-1-antitrypsin 0.015 Complement and coagulation
   cascades
   [67]P07339 Cathepsin D 0.072 Tuberculosis
   [68]P00450 Ceruloplasmin 0.035 Porphyrin and chlorophyll metabolism
   [69]P18206 Vinculin 0.001 Regulation of actin cytoskeleton
   A0A0J9YX35 Immunoglobulin heavy chain 0.026 PI3K-Akt
   A0A0A0MS14 Immunoglobulin heavy chain 0.009 Calcium
   [70]Open in a new tab

Figure 2.

   [71]Figure 2
   [72]Open in a new tab

   Screening of LBP-related signalling pathways by KEEGG analysis.

   Note: (A) LBP and TLRS-related signaling pathway, (B) LBP and
   NFκB-related signaling pathway.

Results of Laboratory Tests

   In the experimental examination index, the value measurements of
   healthy people and STB patients were compared and found that there was
   no statistical difference in age, WBC, NEUT%, LYM% and other aspects.
   However, the ESR and CRP measurements in STB patients were
   significantly higher than those in normal patients. As we all know, ESR
   and CRP are important test indicators for evaluating STB patients. The
   LBP measurements (102.73±23.29) in STB patients were significantly
   higher than the normal LBP measurements (81.30±15.88), and there were
   statistically significant differences between the two groups, so LBP
   may be used as an indicator to evaluate the difference between STB
   patients and normal people ([73]Table 3).

Table 3.

   General Statistics of Healthy Individuals and STB Patients
    Item (x±s)   Healthy Person (n=30) STB Patients (n=100)
   Age (years)   53.39±9.67            49.47±16.33
   Course (days)          NA           28.88±16.07
   LBP (ng/mL)   81.30±15.88           102.73±23.29
   CRP (mg/L)    1.11±0.70             24.92±23.69
   ESR (mm/h)    8.27±4.20             40.48±28.71
   WBC (*10^9/L) 5.99±1.28             6.61±1.84
   NEUT%         60.19±5.02            64.72±10.93
   LYM%          23.65±7.86            24.13±9.53
   [74]Open in a new tab

   Abbreviations: CRP, C-reactive protein; ESR, erythrocyte sedimentation
   rate; WBC, white blood cell; NEUT, neutrophilic granulocyte; LYM,
   lymphocyte.

Analysis of LBP in Clinical Diagnosis

   Differential expression in serum of STB patients was measured by ELISA
   method, and the results of patients in age, sex, course, WBC, NEUT%,
   LYM%, pathology, T-SPOT, acid fast bacillus and imaging were analyzed,
   and the results showed that STB patients had no statistical
   significance (P > 0.05) in age, sex, course, WBC, NEUT, LYM%, and ESR,
   CRP, pathology, T-SPOT. There were statistically significant results
   for acid-resistant staining and imaging (t ≤ 0.05) ([75]Table 4).

Table 4.

   Distribution of LBP Proteins in STB Clinical Phenotypes
       Item      n  LBP Value (x±s) t-Test (t)   P
   Age (years)
    ≤50          51 103.27±20.54    0.237      0.813
    >50          49 102.16±26.05
   Gender
    Men          50 101.15±23.74    −0.678     0.499
    Women        50 104.31±22.96
   Course (days)
    ≤14          23 100.81±32.89    −0.450     0.654
    >14          77 103.30±19.80
   WBC values
    Normal       91 101.66±22.61    −1.475     0.143
    Abnormal     9  113.59±28.58
   LYM %
    Normal       67 99.83±22.11     −1.792     0.076
    Abnormal     33 108.61±24.82
   NEUT %
    Normal       78 100.53±22.17    −1.801     0.075
    Abnormal     22 110.54±25.96
   ESR Values
    Normal       22 83.13±24.26
    Abnormal     78 108.26±19.90    −4.978     0.000
   CRP Values
    Normal       15 66.32±14.45
    Abnormal     85 109.15±18.06    −8.696     0.000
   [76]Open in a new tab

   Abbreviations: WBC, white blood cell; CRP, C-reactive protein; LYM,
   lymphocyte; ESR, erythrocyte sedimentation rate; NEUT%, neutrophilic
   granulocyte.

Analysis of LBP Protein Measurement Data in Laboratory Examination

   Correlation analysis of clinical counts was carried out using curve
   regression and Spearman correlation analysis, and it was found that the
   LBP protein content in patients was positively correlated with CRP and
   ESR changes, which was statistically significant (P ≤ 0.01), and there
   was no correlation with the course, age, WBC, NEUT%, LYM% values, and
   there was no statistical significance (P ≥ 0.05) ([77]Figure 3 and
   [78]Table 5). It is suggested that LBP has a synergistic effect with
   CRP and ESR in the detection of infectious or inflammatory diseases.

Figure 3.

   [79]Figure 3
   [80]Open in a new tab

   Analysis and detection results of LBP in peripheral blood of STB
   patients.

   Note: (A–G) are curve regression analyses of CRP, ESR, disease
   duration, WBC, age, NEUT%, and LYM% values correlated with measured
   data of LBP expression levels in STB patients.

Table 5.

   Analysis of Intra-Serum LBP Protein Levels and Clinical Phenotype in
   STB Patients
          Spearman Rho          CRP   ESR  Course  WBC   Age  NEUT%  LYM%
   LBP Correlation coefficient 0.544 0.345 0.101  0.260 0.037 0.035 −0.064
       P-value                 0.000 0.000 0.317  0.009 0.713 0.732 0.530
       N                       100   100   100    100   100   100   100
   [81]Open in a new tab

   Note: P < 0.01, significant correlation.

   Abbreviations: CRP, C-reactive protein; ESR, erythrocyte sedimentation
   rate; WBC, white blood cell; NEUT, neutrophilic granulocyte; LYM,
   lymphocyte.

Evaluation of LBP as a Diagnostic Biomarker

   The positive results of pathology, bacteriological culture, T-SPOT
   detection, imaging diagnosis, and acid fast bacillus were statistically
   analyzed by independent sample t-test and ROC curve analysis, and the
   results showed that the expression of LBP protein content was
   statistically significant in pathology positive, bacteriological
   positive, T-SPOT positive, imaging diagnosis, and positive
   acid-resistance staining (P < 0.01) ([82]Table 4). The results of ROC
   curve analysis showed that the AUC of the LBP protein identified ASB
   positive for STB in pathology, bacteriology, T-SPOT, imaging, and acid
   fast bacillus was 0.677, 0.707, 0.751, 0.714, and 0.656 (P < 0.01)
   ([83]Figure 4). The results showed that the effect of LBP in diagnosing
   STB was related to the diagnostic results of pathology, bacteriology,
   T-SPOT, imaging, and acid fast bacillus, and LBP had the potential to
   be used as a biomarker for diagnosing STB in peripheral blood.

Figure 4.

   [84]Figure 4
   [85]Open in a new tab

   ROC analysis of molecular biomarkers of LBP in peripheral blood of STB
   patients.

   Note: (A–E) are the ROC curve analysis of LBP combined with pathology,
   bacterial culture, T-SPOT, imaging, and antacid staining for diagnosis
   of STB.

Discussion

   STB is a common form of extrapulmonary tuberculosis, caused by
   Mycobacterium tuberculosis spreading through the circulatory system and
   localizing to the blood vessels of the bone, resulting in destructive
   bone and joint disease.[86]^9 Early symptoms and clinical
   manifestations are atypical, and the hidden course of the disease is
   the main reason for the untimely treatment of STB and the main cause of
   serious complications, therefore diagnosis is of great significance for
   STB treatment and prognosis. The lack of specific and sensitive
   diagnostic indicators in the process of STB diagnosis and treatment is
   the main reason for the delay in the diagnosis and treatment of STB
   patients. LBP has stable and specific expression characteristics in the
   peripheral blood of STB patients, which makes LBP may have special
   significance in STB diagnosis and potential as a diagnostic marker of
   STB.

   In this study, we used proteomics to screen for differentially
   expressed proteins in the peripheral blood of STB patients and healthy
   individuals, under the conditions of fold change (FC) ≥ 1.5, P<0.01,
   and relevant proteins associated with STB signaling pathway, we
   verified by ELISA whether there was differential expression in the
   peripheral blood of healthy individuals and STB patients and finally
   determined that LBP met the conditions. Studies have shown that LBP is
   a protein with high affinity for binding to the acute phase of
   glycosylation of the bacterial lipopolysaccharide part and plays an
   important role in the pathophysiological process of sepsis.[87]^10 At
   the same time, Gyula found that LBP expression in postoperative renal
   cell carcinoma patients has important implications for the detection
   and adjuvant treatment of the disease and can be used as a relevant
   biomarker for the progression of renal cell carcinoma after
   surgery.[88]^6 In addition, Huang found that LBP in osteoarthritis of
   the knee can be used as a diagnostic marker for evaluating the severity
   of osteoarthritis of the knee.[89]^11 Increased LBP levels during
   inflammation, insulin resistance, and metabolic stress suggest impaired
   cellular differentiation.[90]^12 Elevated LBP levels have a special
   significance in clinical disorders, such as LBP can be used as a
   biomarker to determine the severity of sepsis.[91]^10 Meanwhile, we
   found that LBP protein was mainly associated with TLRS signaling
   pathway and NF-κB signaling pathway by KEGG analysis. The study found
   that LBP is mainly transmembrane recognition by TLRs-related receptors,
   which plays a role in detecting and defending against pathogenic
   microorganisms in immunity, and high concentrations of LBP in the host
   are believed to play a protective role in immune defense.[92]^13 The
   classic NF-κB signaling pathway in STB pathogenesis is closely related
   to activation of TLRs receptors.[93]^14 Proteomics detection and
   confirmation of stability differences in the expression of LBP in the
   peripheral blood of STB patients, therefore, it is hypothesized that
   LBP may provide a feedback to the organism through its increased
   expression in response to infection or inflammation, regulating the
   organism’s defense function. ROC analysis showed that LBP measured the
   area under the curve in the peripheral blood of STB patients with an
   area of 0.880 (P < 0.01), confirming that changes in LBP expression in
   STB peripheral blood make there a close relationship or special
   biological function between the two. In addition, the differential
   expression of LBP in many clinical diseases has also been shown to be
   associated with the occurrence of diseases, so it is speculated that
   LBP has the potential to be used as a diagnostic biomarker when
   detecting STB. This study found differential expression in peripheral
   blood between LBP and STB patients, and the relationship between LBP
   and STB was also reported for the first time.

   After determining the value of LBP in the diagnosis of STB, we further
   analyzed its relevance to the clinical features of STB and found that
   the relative expression of LBP as a detection index in peripheral blood
   was positively correlated with ESR and CRP levels (P < 0.01). It is
   comparable to Wen,[94]^15 who reported that LBP is significantly
   correlated with elevated ESR and CRP in active disease, and has
   comparable value for disease prediction with ESR and CRP. There was no
   correlation between LBP and age, course, WBC, NEUT%, and LYM% (P>0.05),
   which was basically consistent with the evaluation index of clinical
   STB. LBP performed a curve regression analysis of the positive results
   of pathology, bacteriological culture, T-SPOT, imaging and acid fast
   bacillus confirmed by STB, and found that the AUC between the two was
   0.677, 0.707, 0.751, 0.714 and 0.656 (P < 0.01), indicating that LBP is
   valuable for STB diagnosis. Positive pathology and cell culture are the
   gold standards for the definitive diagnosis of STB, and ROC curve
   analysis was performed on STB patients with pathology and bacteriology
   positive, which found that LBP had a good correlation with pathology
   and bacteriological culture, confirming that LBP can be used for the
   diagnostic evaluation of STB. T-SPOT has a significant correlation of
   88% and 90% in the sensitivity and specificity of diagnosing spinal
   tuberculosis,[95]^16 and LBP has an area of 0.751 under the T-SPOT
   curve, indicating that LBP has a significant correlation with T-SPOT,
   which has good sensitivity and specificity for detecting STB and can be
   used as a diagnostic marker for the diagnosis of STB. MRI and CT in
   imaging have higher sensitivity and specificity in the diagnosis of
   STB,[96]^17 and there is also a good correlation in the analysis of the
   ROC curve of LBP in the diagnosis of MRI and CT, which indicates that
   LBP has high diagnostic value for STB and can be used as an evaluation
   factor for its diagnosis. The specificity and sensitivity of AFB in the
   detection of STB were 87.0% and 85.9%, respectively, and the positive
   curve regression results of LBP showed that the two were correlated,
   which had a synergistic effect in the diagnostic evaluation of STB.
   Combined with the reports that LBP as a biomarker for evaluating
   rheumatoid arthritis sensitivity, evaluating the prediction of cardiac
   insufficiency prognosis after radiation therapy for breast cancer, and
   distinguishing mastocytosis, there are significant differences in
   peripheral blood expression of LBP in many clinical diseases, which
   indicates that LBP is indeed closely related to clinical
   diseases.[97]^15^,[98]^18^,[99]^19 In this study, the statistical
   analysis of the differences and clinical phenotypes of LBP in the
   peripheral blood of STB patients was screened and found that it has the
   potential as a biomarker in the diagnosis of STB.

Conclusion

   In conclusion, this study found for the first time that LBP has stable
   differential expression in the peripheral blood of STB patients, which
   has high diagnostic value in the analysis of ESR, CRP, pathology and
   imaging indicators, and has the potential as a molecular marker for STB
   diagnosis; LBP may also play an important role in the study of STB
   pathogenesis-related mechanisms, but the specific functional mechanism
   still needs to be further explored. The shortcoming of this study is
   that the sample size collected is relatively small, LBP protein needs
   to be validated as a diagnostic biomarker for clinical use in STB with
   expanded samples.

Funding Statement

   National Natural Science Foundation of China (grant No. 81860395); Key
   R&D program of the autonomous region (grant No.: 2022BEG03099); Natural
   Science Foundation of Ningxia (No. 2020AAC03391); The National Natural
   Science Foundations of China (No.82160305); The First-Class Discipline
   Construction Project of Ningxia Medical University Clinical Medicine
   (2022); The fifth group of Ningxia Young Scientific and Technological
   Talents lifting Project (NXKJTJGC2020080).

Abbreviations

   LBP, lipopolysaccharide-binding protein; STB, Spinal tuberculosis; WBC,
   white blood cell; CRP, C-reactive protein; ESR, Erythrocyte
   Sedimentation Rate; T-SPOT, T-cell spot test for tuberculosis
   infection; ELISA, enzyme-linked immunosorbent assay; AUC, area under
   the curve; TMT, Tandem Mass Tag; AFB, acid fast bacillus; MRI, Magnetic
   Resonance Diffusion Tension Imaging; CT, computerized tomography; NEUT,
   neutrophilic granulocyte; LYM, lymphocyte.

Data Sharing Statement

   The datasets used and analyzed during the current study are available
   from the corresponding author upon reasonable request.

Ethics Approval and Consent to Participate

   Approval for the study was obtained from the ethics committee of
   General Hospital of Ningxia Medical University. The verbal consent was
   obtained from each patient before enrollment. In an outbreak
   investigation situation, the ethics committee approved this procedure
   to conduct an investigation after obtaining verbal consent. All
   procedures were performed in accordance with relevant guidelines in the
   manuscript.

Author Contributions

   Caili Lou is the first author. All authors made a significant
   contribution to the work reported, whether that is in the conception,
   study design, execution, acquisition of data, analysis and
   interpretation, or in all these areas; took part in drafting, revising
   or critically reviewing the article; gave final approval of the version
   to be published; have agreed on the journal to which the article has
   been submitted; and agree to be accountable for all aspects of the
   work.

Disclosure

   The authors declare that they have no competing interests.

References