RNF20 deletion causes inflammation in model of sepsis through the NLRP3 activation.

Aim: Sepsis is an extremely complex, threatening and difficult-to-treat disease, which can occur at any age and under any underlying disease. RNF20 regulate NF-kappaB (NF-κB) signaling pathway and the transcription of inflammatory factors of target genes. Therefore, it is of great significance to study the function of RNF20 in the clinical treatment of sepsis and its underlying mechanisms.Methods: C57BL/6 mice were subjected to cecal ligation and puncture (CLP) surgery. THP-1 cells were induced with Lipopolysaccharide for 4 h.Results: RNF20 gene, mRNA expression and protein expression were reduced in patients with sepsis and mice with sepsis. Based on RNF20 deletion (RNF20-/-) mice, these were found to be increased inflammation reactions in RNF20-/- mice. However, the RNF20 human protein reduced inflammation reactions in mice with sepsis. In vitro model of sepsis, over-expression of RNF20 inhibited inflammation reactions by inducing Vitamin D Receptor (VDR), while down-regulation of RNF20 promoted inflammation reactions through the suppression of VDR. RNF20 protein was interlinked with VDR protein, and VDR protein was also interlinked with NLRP3. Furthermore, VDR promoted NLRP3 ubiquitination and reduced NLRP3 function in vitro model of sepsis.Conclusion: These studies demonstrate that RNF20 suppressed inflammation reactions in models with sepsis through NLRP3 inflammasome and NLRP3 ubiquitination by activating VDR.

Frequency and mortality of sepsis and septic shock in China: a systematic review and meta-analysis.

BACKGROUND: Sepsis, a life-threatening organ dysfunction induced by infection, is a major public health problem. This study aimed to evaluate the frequency and mortality of sepsis, severe sepsis, and septic shock in China. METHODS: We Searched MEDLINE, Embase, PubMed, and Cochrane Library from 1 January 1992 to 1 June 2020 for studies that reported on the frequency and mortality of sepsis, severe sepsis, and septic shock conducted in China. Random effects models were performed to estimate the pooled frequency and mortality of sepsis, severe sepsis, and septic shock. RESULTS: Our search yielded 846 results, of which 29 studies were included in this review. The pooled frequency of sepsis was estimated at 33.6% (95% CI 25.9% to 41.3%, I2 = 99.2%; p < 0.001), and the pooled mortality of sepsis, severe sepsis and septic shock were 29.0% (95% CI 25.3%-32.8%, I2 = 92.1%; p = 0), 31.1% (95% CI 25.3% to 36.9%, I2 = 85.8%; p < 0.001) and 37.3% (95% CI 28.6%-46.0%, I2 = 93.5%; p < 0.001). There was significant heterogeneity between studies. With a small number of included studies and the changing definition of sepsis, trends in sepsis frequency and mortality were not sufficient for analysis. Epidemiological data on sepsis in the emergency department (ED) are severely lacking, and more research is urgently needed in this area is urgently needed. CONCLUSIONS: Our findings indicated that the frequency and mortality of sepsis and septic shock in China were much higher than North America and Europe countries. Based on our results, an extremely high incidence and mortality of sepsis and septic shock in China's mainland requires more healthcare budget support. Epidemiological data on sepsis and septic shock in ED are severely lacking, and more research is urgently needed in this area. Trial registration This systematic review was conducted according to the statement of the preferred reporting items for systematic review (PROSPERO CRD42021243325) and the meta-analysis protocols (PRISMA-P).

Bioinformatics Analysis for Multiple Gene Expression Profiles in Sepsis.

BACKGROUND This work aimed to screen key biomarkers related to sepsis progression by bioinformatics analyses. MATERIAL AND METHODS The microarray datasets of blood and neutrophils from patients with sepsis or septic shock were downloaded from Gene Expression Omnibus database. Then, differentially expressed genes (DEGs) from 4 groups (sepsis versus normal blood samples; septic shock versus normal blood samples; sepsis neutrophils versus normal controls and septic shock neutrophils versus controls) were respectively identified followed by functional analyses. Subsequently, protein-protein network was constructed, and key functional sub-modules were extracted. Finally, receiver operating characteristic analysis was conducted to evaluate diagnostic values of key genes. RESULTS There were 2082 DEGs between blood samples of sepsis patients and controls, 2079 DEGs between blood samples of septic shock patients and healthy individuals, 6590 DEGs between neutrophils from sepsis and controls, and 1056 DEGs between neutrophils from septic shock patients and normal controls. Functional analysis showed that numerous DEGs were significantly enriched in ribosome-related pathway, cell cycle, and neutrophil activation involved in immune response. In addition, TRIM25 and MYC acted as hub genes in protein-protein interaction (PPI) analyses of DEGs from microarray datasets of blood samples. Moreover, MYC (AUC=0.912) and TRIM25 (AUC=0.843) had great diagnostic values for discriminating septic shock blood samples and normal controls. RNF4 was a hub gene from PPI analyses based on datasets from neutrophils and RNF4 (AUC=0.909) was capable of distinguishing neutrophil samples from septic shock samples and controls. CONCLUSIONS Our findings identified several key genes and pathways related to sepsis development.

Neuropilin-1highCD4⁺CD25⁺ Regulatory T Cells Exhibit Primary Negative Immunoregulation in Sepsis.

Regulatory T cells (Tregs) appear to be involved in sepsis-induced immune dysfunction; neuropilin-1 (Nrp-1) was identified as a surface marker for CD4(+)CD25(+)Tregs. In the current study, we investigated the negative immunoregulation of Nrp-1(high)CD4(+)CD25(+)Tregs and the potential therapeutic value of Nrp-1 in sepsis. Splenic CD4(+)CD25(+)Tregs from cecal ligation and puncture (CLP) mouse models were further segregated into Nrp-1(high)Tregs and Nrp-1(low)Tregs; they were cocultured with CD4(+)CD25(-) T cells. The expression of forkhead/winged helix transcription factor-3 (Foxp-3), cytotoxic T-lymphocyte associated antigen-4 (CTLA-4), membrane associated transforming growth factor-ß (TGF-ß(m+)), apoptotic rate, and secretive ability [including TGF-ß and interleukin-10 (IL-10)] for various types of Tregs, as well as the immunosuppressive ability of Tregs on CD4(+)CD25(-) T cells, were determined. Meanwhile, the impact of recombinant Nrp-1 polyclonal antibody on the demethylation of Foxp-3-TSDR (Treg-specific demethylated region) was measured in in vitro study. Sepsis per se markedly promoted the expression of Nrp-1 of CD4(+)CD25(+)Tregs. Foxp-3/CTLA-4/TGF-ß(m+) of Nrp-1(high)Tregs were upregulated by septic challenge. Nrp-1(high)Tregs showed strong resilience to apoptosis and secretive ability and the strongest immunosuppressive ability on CD4(+)CD25(-) T cells. In the presence of lipopolysaccharide (LPS), the recombinant Nrp-1 polyclonal antibody reduced the demethylation of Foxp-3-TSDR. Nrp-1(high)Tregs might reveal primary negative immunoregulation in sepsis; Nrp-1 could represent a new potential therapeutic target for the study of immune regulation in sepsis.

[Construction and validation of a nomogram for predicting the prognosis of patients with septic shock in department of emergency medicine].

OBJECTIVE: To construct a nomogram model for predicting the 28-day mortality of patients with septic shock in the emergency medicine department and to validate the predictive efficacy. METHODS: Based on the database of the emergency medicine department of Chu Hsien-I Memorial Hospital of Tianjin Medical University, Tianjin Medical University General Hospital and the Second Hospital of Tianjin Medical University, the data of 913 patients with septic shock admitted to the emergency medicine department from January 2017 to October 2020 were collected, including baseline demographic information and clinical characteristics, laboratory indices, and the main endpoints (28-day mortality). The patients were divided into a training set and a validation set based on simple random sampling. All significant variables from the one-way binary Logistic regression analysis of the training set were included in the multivariate Logistic regression analysis to analyze the risk factors for 28-day mortality in patients with septic shock and to construct a column-line graphical model. The predictive efficacy of the nomogram model was assessed using calibration curves and receiver operator characteristic curve (ROC curve). RESULTS: A total of 860 patients with septic shock meeting the criteria were finally enrolled, including 472 in the training set and 388 in the validation set. The 28-day mortalities were 52.5% (248/472) and 54.1% (210/388) for the training and validation sets, respectively. In the training set, age, respiratory rate (RR), the levels of C-reactive protein (CRP), D-dimer, white blood cell count (WBC), neutrophil count (NEU), neutrophil/lymphocyte ratio (NLR), monocyte/lymphocyte ratio (MLR), mean platelet volume (MPV), and platelet count (PLT) in the death group were significantly higher than those in the survival group, and the levels of base remaining (BE), lymphocyte count (LYM), hemoglobin (Hb) and the proportion of chronic obstructive pulmonary diseases (COPD) were significantly lower than those in the survival group (all P < 0.05). Multifactorial Logistic regression analysis showed that NLR [odds ratio (OR) = 0.023 0, 95% confidence interval (95%CI) was -0.204 4 to 0.113 0], MPV (OR = 0.179 8, 95%CI was -0.877 6 to 0.172 7), Hb (OR = 0.007 8, 95%CI was 0.010 3 to 0.040 8), procalcitonin (PCT; OR = 1.957 0, 95%CI was 1.243 0 to 3.081 0), and D-dimer (OR = 0.000 1, 95%CI was -0.000 4 to 0.000 1) were independent predictors of 28-day mortality in patients with septic shock in the emergency department (all P < 0.05). A column-line graph model was established based on the above variables, and the ROC curves showed that the area under the ROC curve (AUC) of the nomogram model in the training set and validation set for predicting the 28-day mortality of patients with septic shock was 0.907 (95%CI was 0.864 to 0.940) and 0.822 (95%CI was 0.781 to 0.863), respectively. The calibration curves showed good agreement between the predicted and observed results for both the training and validation sets. CONCLUSIONS: The nomogram model constructed based on NLR, MPV, Hb, PCT and D-dimer has significant clinical value in predicting the 28-day mortality of patients with septic shock in the emergency medicine department.

[Complement 5a regulates the function of dendritic cells to induce pathogenic polarization of regulatory T cell/helper T cell 17 in sepsis].

OBJECTIVE: To explore the mechanism of complement 5a (C5a) in the pathogenesis of sepsis. METHODS: SPF male C57BL/6J mice were selected and divided into sham operation group (Sham group), cecal ligation and puncture (CLP) group and CLP+anti-C5A monoclonal antibody intervention group (CLP+anti-C5a group) according to random number table with 20 mice in each group. A CLP model was reproduced to induce sepsis, and those in the Sham group only underwent laparotomy without ligation and perforation. In the CLP+anti-C5a group, 0.15 mg of anti-C5a monoclonal antibody was injected intraperitoneally immediately after CLP, and in the Sham group and CLP group were given equal amount of normal saline. The cumulative survival rate was analyzed by Kaplan-Meier method. Serum levels of tumor necrosis factor-α (TNF-α), interleukins (IL-12, IL-4), and interferon-γ (IFN-γ) were measured 24, 48 and 72 hours after operation by enzyme linked immunosorbent assay (ELISA). Immunohistochemical staining was used to observe the expression of C5a receptor (C5aR) in lung and kidney tissues 48 hours after operation. The proportions of dendritic cell (DC), regulatory T cell (Treg) and helper T cell 17 (Th17) in splenic mononuclear cells 48 hours after operation were analyzed by flow cytometry. RESULTS: The 7-day cumulative survival rate of mice in the CLP group was significantly lower than that in the Sham group (30.00% vs. 100.00%; Log-Rank test: χ2 = 47.470, P < 0.001), and the peripheral blood inflammatory mediators TNF-α, IL-12 and IL-4 were increased 24 hours after operation, followed by a significant decreasing at 48 hours, and then gradually increased at 72 hours. IFN-γ gradually increased 24 hours after operation and lasted for 72 hours. Immunohistochemistry showed that a large number of C5aR was expressed in pulmonary and renal endothelial cells 48 hours after operation in the CLP group. Compared with the Sham group, the proportion of DC [(1.80±0.30)% vs. (6.90±1.20)%, P < 0.05] and Treg [(0.38±0.02)% vs. (4.00±0.50)%, P < 0.05] in splenic mononuclear cells was down-regulated in the CLP group, the proportion of Th17 was up-regulated [(0.83±0.08)% vs. (0.32±0.03)%, P < 0.05], and disorder of immune function was found. After anti-C5A monoclonal antibody intervention, the 7-day cumulative survival rate increased significantly compared with the CLP group (54.54% vs. 30.00%; Log-Rank test: χ2 = 28.090, P < 0.001); TNF-α, IL-12 and IFN-γ were further increased, while IL-4 was significantly decreased; the expression of C5aR in lung and kidney tissues were significantly decreased, and the expression of mature DC cells [(5.10±1.20)% vs. (1.80±0.30)%, P < 0.05] and Treg [(2.58±0.05)% vs. (0.38±0.02)%, P < 0.05] in spleen were significantly increased compared with the CLP group, and Th17 was significantly decreased [(0.54±0.05)% vs. (0.83±0.08)%, P < 0.05]. CONCLUSIONS: It is preliminarily concluded that anti-C5A monoclonal antibody may improve the prognosis of sepsis by improving the polarization of mature DC and T cells in the spleen, and C5a plays an important role in the immune regulation of sepsis cells.

The effect of miR-21-5p on the MAP2K3 expressions and cellular apoptosis in the lung tissues of neonatal rats with hyperoxia-induced lung injuries.

OBJECTIVE: To explore the effect of miR-21-5p on the MAP2K3 expressions and cellular apoptosis in the lung tissues of neonatal rats with hyperoxia-induced lung injuries (HILI). METHODS: Twenty Sprague-Dawley neonatal rats were assigned to the normal group, and 120 rats were used to create a HILI model and were divided into the following six groups of 20 rats each: the model group, the miR-21-5p NC group, the miR-21-5p agomir group, the oe-NC group (MAP2K3 overexpression NC), the oe-MAP2K3 group, and the miR-21-5p agomir+oe-MAP2K3 group. RESULTS: miR-21-5p can target MAP2K3. Compared with the normal rats, the rats with HILI had lower miR-21-5p expression levels and higher MAP2K3 expression levels in the lung tissues (both P<0.05). Unlike the normal group, the other groups all presented different degrees of lung injuries, lower Bcl-2 expression levels, higher cellular apoptosis rates, and higher expression levels of cleaved caspase-3, Bax, IL-6, and TNF-α (all P<0.05). Compared with the model and the miR-21-5p NC groups, the miR-21-5p agomir group had better results in terms of the aforementioned markers; compared with the oe-NC group, the oe-MAP2K3 group had worse results in terms of these markers (all P<0.05). Moreover, we found that the protective effects of miR-21-5p overexpression on the lung tissues of HILI rats can be partially blocked by MAP2K3 overexpression. CONCLUSION: miR-21-5p can inhibit MAP2K3 expression and reduce cellular apoptosis in HILI, thereby exerting protective effects on neonatal rats with HILI.

Thyroid hormone disorders: a predictor of mortality in patients with septic shock defined by Sepsis-3?

Decreased serum thyroid hormone levels and their prediction of mortality in septic patients are still controversial, especially with the evolution of the definition of sepsis. This study aimed to assess the ability of thyroid hormone disorders to predict the early mortality of patients with septic shock defined by Sepsis-3. Sixty-three adult patients with septic shock admitted to a university hospital emergency intensive care unit (EICU) were studied. Serum free T3 (FT3), free T4 (FT4), thyroid stimulating hormone (TSH), C-reactive protein (CRP), procalcitonin (PCT), and lactate levels were determined and compared with survival status and organ dysfunction. Among the 63 patients studied, lower serum FT3 and FT4 levels were significantly associated with higher sequential organ failure assessment (SOFA) scores. Patients with septic shock with lower levels of FT3 (≤ 1.70 pmol/L) and FT4 (≤ 9.99 pmol/L) had significantly increased 28-day mortality. There was no significant difference in the serum TSH level between the survivor and nonsurvivor groups. The areas under the receiver operating characteristic curves for FT3 and FT4 levels were associated with 28-day mortality (0.92 and 0.89, respectively) and were higher than that for SOFA (0.82), CRP (0.65) and lactate (0.59). The decrease in serum levels of FT3 and FT4 in patients with septic shock is associated with the severity of organ dysfunction and 28-day mortality. Early detection of serum FT3 and FT4 levels could help clinicians to identify patients at high risk of clinical deterioration.

Recombinant human ulinastatin improves immune dysfunction of dendritic cells in septic mice by inhibiting endoplasmic reticulum stress-related apoptosis.

Urinary trypsin inhibitor (UTI), also known as ulinastatin, has been reported to protect multiple organs against inflammation- and/or injury-induced dysfunction. In the present study, we aimed to investigate the immunomodulation effects of a recombinant human ulinastatin (urinary trypsin inhibitor, UTI) (rhUTI) on splenic dendritic cells (DCs) in cecal ligation and puncture (CLP)-induced septic mice. CLP mice were treated with rhUTI intramuscularly at 0, 12, and 24 h after procedure. Splenic CD11c+ DCs were isolated and accessed with flow cytometry for apoptotic or phenotypic analysis. Protein markers and cytokines were determined with Western blotting or ELISA. Treatment with rhUTI could markedly upregulate levels of costimulatory molecules (CD80, CD86) and MHC-II on surface of the splenic DC in CLP mice. The apoptotic rate of splenic DCs was decreased in CLP mice after rhUTI treatment. The survival rate of septic mice was increased after treatment with rhUTI. In addition, protein level of markers in endoplasmic reticulum stress (ERS)-related apoptotic pathways (including GRP78, caspase-12, and CHOP) were obviously down-regulated in the rhUTI-treated group when compared with the CLP group. These results indicate that rhUTI protects CLP-induced sepsis in mice by improving immune response of splenic DCs and inhibiting the excessive ERS-mediated apoptosis.

miR-183-5p acts as a potential prognostic biomarker in gastric cancer and regulates cell functions by modulating EEF2.

BACKGROUND: Gastric cancer (GC) is the fourth most prevalent malignant tumor and the second leading cause of cancer-related death around the world. Aberrant proliferation and metastasis are the mainspring of death in patients with GC. However, the specific mechanism of gastric cancer is far from being fully elucidated. Accumulating evidence revealed that miRNA played a significant role in the tumorigenesis and development. METHODS: The level of miR-183-5p was detected in 102 GC patients by using qRT-PCR. The prognostic value of miR-183-5p in GC was evaluated. Cell function assays (CCK-8 and transwell assays) were conducted to assess the role of miR-183-5p in proliferation and metastasis in GC. Dual luciferase report assay and western blot were performed to validate this potential target regulated by miR-183-5p in GC. RESULTS: miR-183-5p was down-regulated in GC tissues and cell lines. Remarkable pertinence was obtained between miR-183-5p level and TNM stage, tumor size, invasion depth, and lymph node metastasis. TNM stage, differentiation and miR-183-5p level were independent causes impacting on the overall survival in GC in multivariate analysis. GC individuals with high miR-183-5p level would experience a relatively better survival prognosis. Upregulation of miR-183-5p restrained GC cell proliferation and migration. EEF2 may be a potential target gene regulated by miR-183-5p in GC. CONCLUSION: miR-183-5p acts as a potential prognostic biomarker in gastric cancer and regulates cell functions by modulating EEF2.