Mycobacterium tuberculosis Infection Induces Low-Density Granulocyte Generation by Promoting Neutrophil Extracellular Trap Formation via ROS Pathway.

The roles and characteristics of low-density granulocytes (LDGs) have recently attracted attention; however, the mechanism of the formation of LDGs is yet unclear. In one of our previous studies, the frequency of LDGs was significantly elevated in the peripheral blood of tuberculosis patients, and in situ activation contributed to the generation of LDGs upon Mycobacterium tuberculosis infection. However, the underlying molecular mechanisms are yet to be elucidated. In the present study, the release of neutrophil extracellular traps (NETs) and the levels of ROS were regulated before the normal-density granulocytes (NDGs) to be infected with M. tuberculosis, and the conversion of NDGs to LDGs was monitored subsequently as well. The results showed that tuberculosis-related LDGs spontaneously released high levels of NETs. Promoting the release of NETs led to increase in the conversion of NDGs to LDGs in M. tuberculosis infection, while inhibiting the release of NETs suppressed this conversion after the infection. The M. tuberculosis infection significantly increased the ROS levels in neutrophils and the conversion of NDGs to LDGs. Scavenging ROS or blocking the ROS generation of M. tuberculosis-infected NDGs significantly suppressed the release of NETs and blocked the generation of LDGs. Moreover, inhibiting the formation of NETs without affecting the levels of ROS significantly decreased the conversion of NDGs to LDGs after M. tuberculosis infection. Overall, this study demonstrated that M. tuberculosis could induce the generation of LDGs by promoting the release of NET via ROS pathway.

Salivary Circular RNAs Hsa_Circ_0001874 and Hsa_Circ_0001971 as Novel Biomarkers for the Diagnosis of Oral Squamous Cell Carcinoma.

BACKGROUND/AIMS: Recent studies have demonstrated that circular RNAs (circRNAs) can serve as potential molecular markers for disease diagnosis. However, little is known about their diagnostic potential for oral squamous cell carcinoma (OSCC). This study aimed to determine the expression of circRNAs in the saliva of OSCC patients to identify novel biomarkers for OSCC screening. METHODS: Microarray screening of circRNA was performed to identify differentially expressed circRNAs in saliva from 3 OSCC patients compared with 3 healthy controls. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to validate the results, and the association between these confirmed salivary circRNAs and clinicopathological features was analyzed using the chi-squared test. A receiver operating characteristic (ROC) curve was constructed to evaluate the diagnostic value of the circRNAs identified. Preoperative expression and postoperative expression (1 month after the surgery) of hsa_circ_0001874 and hsa_circ_0001971 was also determined. RESULTS: Our results indicated 12 upregulated and 20 downregulated circRNAs in the saliva from the OSCC patients compared with that from the healthy controls. Among the differentially expressed circRNAs, hsa_circ_0001874, hsa_circ_0001971, and hsa_circ_0008068 were upregulated and hsa_circ_0000140, hsa_circ_0002632, and hsa_circ_0008792 were downregulated in the OSCC group versus the healthy group. Clinical data indicated that salivary hsa_circ_0001874 was correlated with TNM stage (P=0.006) and tumor grade (P=0.023) and that hsa_circ_0001971 was correlated with TNM stage (P=0.019). The combination of hsa_circ_0001874 and hsa_circ_0001971 showed an area under the ROC curve of 0.922 (95% confidence interval, 0.883-0.961; P< 0.001). The risk score based on the combination of hsa_circ_0001874 and hsa_circ_0001971 also discriminated patients with OSCC from patients with oral leukoplakia (P< 0.001). Moreover, the expression levels of salivary hsa_circ_0001874 and hsa_circ_0001971 were clearly decreased in the postoperative samples compared with preoperative samples (P< 0.001). CONCLUSIONS: This is the first study to demonstrate the potential of salivary hsa_circ_0001874 and hsa_circ_0001971 as biomarkers for the diagnosis of OSCC.

Microarray Expression Profile of Circular RNAs in Peripheral Blood Mononuclear Cells from Active Tuberculosis Patients.

BACKGROUND/AIMS: Dysregulated expression of circular RNAs (circRNAs) was demonstrated to be implicated in many diseases. Here, we aimed to determine circRNA profile in peripheral blood mononuclear cells (PBMCs) from active tuberculosis (TB) patients to identify novel biomarkers for TB. METHODS: Expression profile of circRNAs in PBMCs from 3 active pulmonary TB patients and 3 healthy controls were analyzed by microarray assay. Six circRNAs were selected for validation using real time-quantitative PCR (qRT-PCR) in 40 TB patients and 40 control subjects. Receiver operating characteristic (ROC) curve was constructed to evaluate their values in TB diagnosis. Hsa_circRNA_001937 was chosen for further evaluation in an independent cohort consisting of 115 TB, 40 pneumonia, 40 COPD, 40 lung cancer patients and 90 control subjects. An eight-month follow up was performed in 20 newly diagnosed TB patients to investigate the expression change of hsa_circRNA_001937 after chemotherapy. RESULTS: We revealed and confirmed that a number of circRNAs were dysregulated in TB patients. Of the six studied physio circRNAs, the levels of hsa_circRNA_001937, hsa_circRNA_009024 and hsa_ circRNA_005086 were significantly elevated and hsa_circRNA_102101, hsa_circRNA_104964 and hsa_circRNA_104296 were significantly reduced in PBMCs from TB patients as compared to healthy controls. ROC curve analysis suggested that hsa_circRNA_001937 has the largest area under the curve (AUC = 0.873, P<0.001). Hsa_circRNA_001937 was significantly increased in patients with TB compared with patients with pneumonia, COPD and lung cancer. Hsa_ circRNA_001937 was correlated with TB severity (r = 0.4053, P = 0.010) and its expression significantly decreased after treatment. CONCLUSION: This study identified a set of deregulated circRNAs in active TB PBMCs, our data also suggest that hsa_circRNA_001937 can be used as a potential diagnostic biomarker of TB.

[Expression of microRNA-31 and its clinicopathologic significance in oral squamous cell carcinoma].

OBJECTIVE: To investigate the expression of microRNA-31 and its association with clinicopathologic features in oral squamous cell carcinoma (OSCC). METHODS: The expression level of microRNA-31 in 62 cases of OSCC and matched non-tumor adjacent tissue specimens was examined using stem-loop real-time PCR. The relationship between the expression of microRNA-31 and its clinicopathologic features of OSCC was analyzed. RESULTS: The expression of microRNA-31 was significantly higher in the tumor tissues than that in the adjacent tissues (P < 0.05).Up-regulated microRNA-31 expression was associated with the lymph node metastasis (P < 0.05) and cell differentiation (P < 0.05) in OSCC patients.No significant association was found between the expression of microRNA-31 and gender, age, lymph node metastasis, tumor size and location.Receiver operator characteristic curve (ROC) of microRNA-31 about cell differentiation resulted in a diagnostic sensitivity of 70.4% and specificity of 89.5%. CONCLUSIONS: The up-regulated level of microRNA-31 expression may be related to the pathogenesis of OSCC.

Performance of the microscopic observation drug susceptibility assay in pyrazinamide susceptibility testing for Mycobacterium tuberculosis.

BACKGROUND: Drug susceptibility assay is very important in tuberculosis therapy. Pyrazinamide is a first line antituberculosis drug and diagnosis of its resistance in Mycobacterium tuberculosis (M. tuberculosis) is difficult and time consuming by conventional methods. In this study, we aimed to evaluate the performance of the microscopic observation drug susceptibility (MODS) assay in the detection of pyrazinamide resistance in M. tuberculosis relative to the conventional Wayne assay and Lowenstein-Jensen (LJ) proportion method. METHODS: M. tuberculosis clinical isolates (n = 132) were tested by the MODS and the Wayne assay: the results were compared with those obtained by the LJ proportion method. Mutations in the gene were identified by direct sequencing of the pncA genes of all isolates in which pyrazinamide resistance was detected by any of the three methods. RESULTS: Compared to the LJ results, the sensitivity and specificity of the MODS assay were 97.8% and 96.5% respectively; the sensitivity and specificity of the Wayne assay were 87.0% and 97.7% respectively. Mutations in the pncA gene were found in 41 of 46 strains that were pyrazinamide resistant (3 tests), in 1 of the 4 strains (LJ only), in 42 of 48 strains (at least 1 test), but no mutations in 1 strain sensitive according to the MODS assay only. The MODS assay, Wayne assay and LJ proportion method provided results in a median time of 6, 7 and 26 days respectively. CONCLUSIONS: MODS assay offers a rapid, simple and reliable method for the detection of pyrazinamide resistance in M. tuberculosis and is an optimal alternative method in resource limited countries.