Abstract:

Objective: To explore the expression of SEMA4A in different Mycobacterium tuberculosis infection states and evaluate its diagnostic value for active tuberculosis in children. Methods: The study prospectively collected 645 children who underwent flexible bronchoscopy in Tianjin Children’s Hospital from April 2020 to October 2025, and specimens were stored to establish a specimen bank. Two hundred and sixty-five children who met the inclusion and exclusion criteria were classified into four groups: active pulmonary tuberculosis (APTB) group, latent tuberculosis infection (LTBI) group, non-tuberculous respiratory inflammatory disease control (IDC) group, and non-infectious and non-tuberculosis control (NINTC) group. Frequency matching was used for balanced grouping, and a total of 200 cases were finally included, 50 cases per group. The concentrations of SEMA4A from serum and bronchoalveolar lavage fluid (BALF) were measured by ELISA. Level of SEMA4A mRNA from BALF was detected by qPCR, and SEMA4A protein expression by Western Blot. Receiver operating characteristic (ROC) curves were constructed to evaluate the diagnostic values. Results: The serum SEMA4A concentrations of the four groups measured by ELISA were: APTB group (5.505 (1.713, 7.101) pg/ml) was significantly higher than 3.909 (1.164, 5.146) pg/ml in the LTBI group, 3.061 (0.670, 4.249) pg/ml in the IDC group and 2.350 (1.103, 3.153) pg/ml in the NINTC group (Z=2.038, P=0.042; Z=2.684, P=0.007; Z=3.984, P<0.001). Overall inter-group difference among the four groups was statistically significant (H=9.327, P=0.025). qPCR detected average relative expression of SEMA4A mRNA in BALF of the APTB group was 6.112±0.440, higher than 4.124±0.272 in the LTBI group, 2.086±0.517 in the IDC group and 1.000±0.630 in the NINTC group. The LTBI group also had higher expression level compared with the IDC group and NINTC group, with all differences statistically significant (t=2.798, P=0.008; t=4.228, P<0.001; t=5.068, P<0.001; t=2.475, P=0.012; t=3.225, P=0.002). There was a significant difference among the four groups (F=128.387, P<0.001). Western Blot tested average relative expression of SEMA4A in BALF was 2.527±0.138 in the APTB group, which was higher than 1.917±0.384 in the LTBI group, 1.619±0.275 in the IDC group and 1.000±0.370 in the NINTC group. The LTBI group showed higher expression than the IDC group and NINTC group, and the differences were statistically significant (t=2.566, P=0.014; t=3.936, P<0.001; t=4.637, P<0.001; t=2.194, P=0.029; t=2.984, P=0.005). Significant inter-group difference was observed among the four groups (F=94.979, P<0.001).ROC curve analysis revealed that the AUC (95%CI) values of SEMA4A detected by ELISA, qPCR and Western Blot for active tuberculosis diagnosis were 0.776 (0.684-0.869), 0.848 (0.767-0.929) and 0.821 (0.736-0.906), respectively. Conclusion: Detection of SEMA4A expression can serve as an auxiliary indicator for the diagnosis of active tuberculosis in children, and qPCR assay of BALF exhibits the optimal diagnostic efficiency.

Key words: Mycobacterium infections, Carrier state, Membrane glycoproteins, Children, Semaphorin 4A