MicroRNAs combined with the TLR4/TDAG8 mRNAs and proinflammatory cytokines are biomarkers for the rapid diagnosis of sepsis.

The early diagnosis and treatment of sepsis are of particular importance to patient survival. To obtain novel biomarkers that serve as prompt indicators of sepsis, the current study screened the differentially expressed microRNAs (DEMs) that were associated with sepsis susceptibility. The correlation between the elucidated DEMs and the inflammatory response was also examined. The present study included 40 patients with sepsis and 40 healthy controls. RNA­sequencing technology and bioinformatics analysis were applied to screen the DEMs between the two cohorts. The expression of these DEMs was subsequently verified by performing reverse transcription­quantitative PCR (RT­qPCR). In addition, IL­6, IL­21, C­X­C motif chemokine ligand­8 (CXCL8) and monocyte chemoattractant protein­1 (MCP­1) levels, along with T­cell death­associated gene 8 (TDAG8) and toll­like receptor 4 (TLR4) mRNA expression levels were assessed. The association between microRNA (miRNA/miR)­3663­3p and the secretion of various proinflammatory cytokines or TDAG8 and TLR4 mRNA expressions were subsequently evaluated by linear correlation analysis. The results revealed 305 DEMs (P<0.05; fold change >2) between patients with sepsis and healthy controls. Among these, the top 18 up­ and downregulated miRNAs were selected for RT­qPCR verification. In addition, the serum content of IL­6, IL­21, CXCL8 and MCP­1, and the expression of TDAG8 and TLR4 mRNAs were significantly increased in patients with sepsis compared with healthy controls. Moreover, in patients with sepsis, a positive correlation was identified between miR­3663­3p and the secretion of inflammatory cytokines or TDAG8 and TLR4 mRNA expression. A positive correlation was also elucidated between TDAG8 and TLR4 mRNA expression and proinflammatory cytokine/chemokine secretion. Receiver operating characteristic curve analysis of miR­3663­3p expression, IL­6, IL­21, CXCL8 and MCP­1 secretion and TDAG8 and TLR4 mRNA expression demonstrated that miRNA analysis may be invaluable for the diagnosis of sepsis. Collectively, the results determined that miR­3663­3p may be a potentially powerful diagnostic and predictive biomarker of sepsis and that the combined and simultaneous detection of several biomarkers, including proteins, miRNAs and mRNA may be a reliable approach for the fast diagnosis and early identification of sepsis.

Effect of Src tyrosine kinase on a rat model of asthma.

Src tyrosine kinase is a protein encoded by the Src gene. The present study aimed to determine the role of Src protein kinase in asthma using small interfering RNA (siRNA) technology. Several Src siRNAs were designed and the most effective siRNA pair was selected. A rat model of asthma was established using ovalbumin, and the rats were treated with Src siRNA, empty vector or phosphate-buffered saline (PBS). A non-asthmatic control group was also established. The rats were clinically observed and Src mRNA and protein levels were measured by reverse transcription-quantitative PCR and western blot analysis, respectively. Pathological observation of the lung tissue, counting of white blood cells (WBCs) and eosinophils (EOSs) and analysis of the concentrations of IL-5, IL-33 and IFN-γ in the bronchoalveolar lavage fluid were performed. The expression levels of Src mRNA in the control, PBS, empty vector and siRNA groups were 110±30.7x103, 253±55.4x103, 254±41.3x103 and 180±50.9x103, respectively. Histochemical analysis of the lung tissue of rats in the siRNA group exhibited a relatively complete lung structure and little damage to the alveolar cavity. Src protein expression and IL-5, IL-33 levels, WBC and EOS levels were positively correlated with Src mRNA expression, while the IFN-γ concentration was negatively correlated with Src mRNA expression. These results indicate that Src knockdown inhibits the release of tracheal inflammatory factors and significantly alleviates asthma in rats. In conclusion, the present study utilized a gene transfer technique to interfere with the expression of Src in rats, which decreased the levels of IL-5, IL-33, WBCs and EOSs and increased the level of IFN-γ; these changes effectively ameliorated the condition of the trachea in asthmatic rats.

[Effects of hypoxia on the expression of HMGB1 and related inflammatory factors in human pulmonary artery smooth muscle and pulmonary artery endothelial cells].

OBJECTIVE: To investigate the effects of hypoxia on the expressions of high mobility group box-1(HMGB1), HMGB1 receptors and inflammatory factors in human pulmonary artery smooth muscle cell( HPASMC) and human pulmonary artery endothelial cells (HPAEC).The effects of HMGB1 on the proliferation and migration activity of the two kinds of cells were detected. METHODS: HPASMC and HPAEC were cultured under hypoxic conditions (Hypoxia at 1% oxygen, Hypoxia group) and normoxic conditions(Control group) . The mRNA levels of HMGB1, Toll-like receptors 2,4,9 (TLR2 , TLR4, TLR9), the receptor of advanced glycation end products(RAGE) , CD24 and IL-6 ,TNF-α,CXCL8 were detected by real-time PCR. Cell proliferation was measured by MTS. Cell migration was analysed by wound healing test. RESULTS: Compared with control group, the expressions of HMGB1 mRNA and RAGE mRNA in both HPASMC and HPAEC were increased significantly (P＜0.05 and 0.01). Meanwhile, the expressions of CD24 mRNA in HPAEC and IL-6 mRNA in HPASMC of hypoxia group were increased significantly (P＜0.05). MTS results showed that HMGB1 inhibited the proliferation of HPAEC at 345 pmol/L significantly (P＜0.01). HMGB1 had no effect on the proliferation of HPASMC. Wound healing test showed that HMGB1 had no significant effect on HPASMC and HPAEC. CONCLUSION: HMGB1 was produced by HPAEC and HPASMC induced by hypoxia. HMGB1 induces endothelial barrier dysfunction by inhibiting HPAEC proliferation. Hypoxia stimulates HPASMC to produce inflammatory cytokines.

[Expressions of G2A and OGR1 in peripheral blood cells of patients with hypoxia induced pulmonary hypertension].

OBJECTIVE: To detect the expression changes of proton-sensing receptor G2 accumulation (G2A) and ovarian cancer G protein-coupled receptor 1(OGR1) in human peripheral blood cells in hypoxic pulmonary hypertension patients(HPH). METHODS: Thirty-one patients with HPH were enrolled for HPH group(16 men and 15 women,age:(65.19±5.86) and thirty healthy persons were enrolled for the control group (NC group). The peripheral blood samples were collected and the mRNA expressions of G2A and OGR1 were determined by using real-time fluorescent quantitative PCR. The serum levels of tumor necrosis factor-α (TNF-α) were detected by using enzyme-linked immunosorbent assay (ELISA). RESULTS: PaCO2 was increased significantly in HPH group than that of the NC group (P<0.05). Forced expiratory volume in 1 sencond(FEV1)pro% and FEV1/forced vital capacity(FVC)in HPH group were significant lower than those of the NC group(P<0.05). The expressions of peripheral blood G2A mRNA and TNF-α in HPH group were increased dramatically than those of the NC group(P<0.05). The expressions of OGR1 mRNA in peripheral blood had no difference between HPH group and NC group. The expressions of G2A and TNF-α in HPH group were positively related to pulmonary artery pressure significantly. CONCLUSIONS: The expression of proton-sensing receptor G2A and the level of TNF-α are increased in peripheral blood cells of patients with pulmonary hypertension.The expressions of TNF-α,G2A and pulmonary artery pressure have positive correlation in the HPH group.