HMGB1-activatied NLRP3 inflammasome induces thrombocytopenia in heatstroke rat.

Background: Thrombocytopenia, an early common complication in heatstroke (HS), has been widely considered as a mortality predictor of HS. The mechanism underlying thrombocytopenia in HS remains unknown. It is not known whether NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome is activated in HS platelet, which, in turn, induces platelet activation and thrombocytopenia. This study tried to clarify the activation of the NOD-like receptor signaling pathway under HS conditions and investigate its roles in mediating HS-induced thrombocytopenia. Methods: Rat HS models were established in a certain ambient temperature and humidity. Platelets, isolated from blood, were counted and CD62P, an index of platelet activation, was measured by flow cytometry in all rats. The colocalization of NLRP3 inflammasome in platelet was detected by confocal fluorescence microscopy. Mitochondrial-derived reactive oxygen species (ROS) was detected using the molecular probes. Plasma HMGB1 and IL-1ß levels were measured by ELISA. Results: Platelet activation, showed by upregulated CD62P, and thrombocytopenia were observed in HS rats. HS activated the NLRP3 inflammasome, which was induced by elevated levels of ROS, while the upregulated CD62P and thrombocytopenia triggered by NLRP3 inflammasome were attributed to the high mobility group box protein 1 (HMGB1) inplasma. Moreover, inhibition of the NOD-like receptor signaling pathway in rats with HS suppressed platelet activation and the decline of platelet count. Similar results were obtained when the receptor toll-like receptor 4 (TLR4)/advanced glycation end product (RAGE) was blocked. Conclusions: The NOD-like receptor signaling pathway induces platelet activation and thrombocytopenia in HS rats. These findings suggested that the NLRP3 inflammasome might be the potential target for HS treatment.

Circ_0000284 Promoted Acute Pancreatitis Progression through the Regulation of miR-10a-5p/Wnt/ß-Catenin Pathway.

Circular RNAs (circRNAs) have been found to be involved in the progression of acute pancreatitis (AP). The objective of our study was to investigate the effects of circ_0000284 on caerulein-induced AR42J cell injury. To mimic AP in vitro, rat pancreatic acinar AR42J cells were treated with caerulein. The expression of circ_0000284 and miR-10a-5p was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). Enzyme-linked immunosorbent assay (ELISA) was employed to determine the content of inflammatory cytokines interleukin (IL)-1ß, IL-6, IL-8 and tumor necrosis factor α (TNF-α). Western blotting was applied to analyze the levels of Wnt/ß-catenin pathway-related and apoptosis-related proteins. Cell viability and apoptosis were monitored by Counting Kit-8 (CCK-8) assay and flow cytometry, respectively. The target connection between circ_0000284 and miR-10a-5p was verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. AP induced inflammation in patients, and caerulein treatment increased apoptosis and inflammation in AR42J cells. Circ_0000284 was upregulated in serum of AP patients and caerulein-induced AR42J cells, while Wnt/ß-catenin pathway was inactivated. Knockdown of circ_0000284 could decrease apoptosis and inflammation in caerulein-induced AR42J cells, which was attenuated by miR-10a-5p inhibition or Wnt signaling pathway antagonist Dickkopf-related protein 1 (DKK1). MiR-10a-5p was sponged by circ_000028 and was downregulated in caerulein-induced AR42J cells. Circ_0000284 depletion could protect caerulein-induced AR42J cells from apoptosis and inflammation by upregulating miR-10a-5p expression and activating Wnt/ß-catenin pathway, underscoring a potential target for AP therapy.

MicroRNA-204 may participate in the pathogenesis of hypoxic-ischemic encephalopathy through targeting KLLN.

Hypoxic-ischemic encephalopathy (HIE) is a common neonatal disease that can lead to high neonatal mortality rates. Previous studies have indicated that microRNAs (miRs) may be involved in the pathogenesis of HIE; however, the specific mechanisms underlying their involvement require further investigation. The aim of the present study was to investigate the roles of miR-204 and its target gene killin p53 regulated DNA replication inhibitor (KLLN) in HIE using rat HIE models. Brain injury was induced by surgery and incubation of hypoxic incubator brain using 10-day-old pup rats. On day 3, rats were sacrificed, and the infarct size of the brain was determined using a tetrazolium chloride assay. Terminal deoxynucleotidyl transferase UTP nick-end labeling staining was performed to detect the cell death rate in the brain tissue. Following this, the brain tissues were collected, and reverse transcription-quantitative polymerase chain reaction, western blot analysis and immunohistochemistry assays were performed to examine the expression levels of miR-204 and KLLN. Furthermore, neurons were cultured and transfected with miR-204 inhibitors or mimics, and the effect of miR-204 on the proliferation and apoptosis of neurons was examined using MTT and flow cytometric assays. Finally, a dual-luciferase reporter assay was performed to confirm whether KLLN is a direct target of miR-204. The expression of miR-204 was significantly downregulated and the expression of KLLN was significantly increased in the brain tissue of HIE rats (P<0.001). In addition, the transfection with miR-204 inhibitors significantly decreased the proliferation rates and significantly increased the apoptosis rate of neurons; however, transfection with miR-204 mimics prompted the opposite results. The dual-luciferase reporter assay also confirmed that KLLN is a direct target of miR-204. Taken together, the results of the present study demonstrated that miR-204 was downregulated in HIE and that miR-204 may serve important roles in the pathogenesis of HIE through targeting KLLN.

Mesenteric Lymph Duct Ligation Alleviating Lung Injury in Heatstroke.

To explore the roles of mesenteric lymph on lung injury in heatstroke (HS), HS rat model was prepared in a prewarmed incubator. Vascular endothelium injury biomarkers (circulating endothelial cell [CEC] as well as von Willebrand factor [vWF] and thrombomodulin [TM]), proinflammatory factors (tumor necrosis factor-α [TNF-α], interleukin-1ß [IL-1ß], IL-6, and high mobility group box 1), and coagulant markers (activated partial thromboplastin time, prothrombin time, D-Dimer, and platelet count) were tested in HS and HS with mesenteric lymph duct ligation (LDL) rats. In addition, lung histopathology; arterial blood gas; Evans Blue dye (EBD) and protein lung permeability; intralung inflammatory parameters including bronchoalveolar lavage fluid (BALF) TNF-α, IL-1ß, and IL-6 levels; myeloperoxidase (MPO) activity; and vWF immune staining were analyzed. LDL prolonged HS onset time but not HS survival time. LDL significantly attenuated endothelial cell injury for decreased CEC counts as well as plasma vWF and TM concentrations; downregulated systemic inflammation for decreased plasma TNF-α, IL-1ß, IL-6, and high mobility group box 1 levels; and ameliorated coagulant disorders for decreased activated partial thromboplastin time, prothrombin time, and D-Dimer levels as well as increased platelet counts. LDL also significantly reduced acute lung pathological injury; improved lung function indexes including arterial blood PaO2, pH, PaCO2, and lactic acid; decreased BALF TNF-α, IL-1ß, and IL-6 levels and lung MPO activity; improved EBD and protein lung permeability; and inhibited lung vascular endothelium vWF expression. However, all of these parameters were not recovered to the normal states. In summary, LDL developed protection roles systemically and alleviated lung injury in HS rats which indicated that modulating mesenteric lymph flow may have some potential benefits in HS.

Anti-Programmed Cell Death (PD)-1 Immunotherapy for Malignant Tumor: A Systematic Review and Meta-Analysis.

This systematic review and meta-analysis evaluated anti-programmed cell death (PD)-1 immunotherapy (nivolumab or pembrolizumab) for overall efficacy, safety, and effective dose relative to standard chemotherapy or other conventional drugs in the treatment of malignant tumors. We searched the following databases, PubMed, Medline, Embase, Cochrane, Wangfang Data, Weipu, and China National Knowledge Infrastructure, and the reference lists of the selected articles for randomized controlled trials (RCTs) of anti-PD-1 therapies in humans. The outcome measures were overall survival, treatment response, and adverse events. Only four randomized controlled trials met our inclusion criteria. Three of these evaluated responses to nivolumab, whereas one tested pembrolizumab. The result of our analysis suggested that nivolumab may improve the overall response rate in treating melanoma relative to chemotherapy and has few associated adverse events. Similarly, in metastatic melanoma patients, nivolumab had a significant advantage over dacarbazine in terms of 1-year survival, progression-free survival, and objective response rate. Regarding dose levels of nivolumab for patients with metastatic renal cell carcinoma, the outcomes in response to 2 and 10 mg/kg were similar, but both had significant advantages over 0.3 mg/kg. In addition, pembrolizumab showed similar outcomes in response to 2- and 10-mg/kg treatment. Anti-PD-1 immunotherapy appears to be safe and effective for patients with melanoma or metastatic renal cell carcinoma. Our meta-analysis is limited, but additional clinical trials are warranted to verify this preliminary evidence of positive outcomes and before anti-PD-1 therapy can be recommended for routine clinical use.

EICOSAPENTAENOIC ACID ENHANCES HEATSTROKE-IMPAIRED INTESTINAL EPITHELIAL BARRIER FUNCTION IN RATS.

OBJECTIVE: Dysfunction of the intestinal barrier plays an important role in the pathological process of heatstroke. Omega-3 (or n-3) polyunsaturated fatty acids, including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), help protect the intestinal mucosal barrier. This study assessed if pretreating rats with EPA or DHA could alleviate heat stress-induced damage to the intestinal barrier caused by experimental heatstroke. METHODS: Male Wistar rats were pregavaged with either EPA, DHA, corn oil, or normal saline (all 1 g/kg) for 21 days before the heatstroke experiment (control rats were not exposed to heat). Experimental rats were exposed to an ambient temperature of 37°C and 60% humidity to induce heatstroke, and then they were allowed to recover at room temperature after rapid cooling. Survival time of rats was monitored after heatstroke. Horseradish peroxidase flux from the gut lumen and the level of plasma D-lactate were measured to analyze intestinal permeability at 6 h after heatstroke. Plasma endotoxin levels were determined using a limulus amoebocyte lysate assay. Expressions of the tight junction (TJ) proteins occludin and ZO-1 were analyzed by Western blot and localized by immunofluorescence microscopy. Tight junction protein morphology was observed by transmission electron microscopy. Fatty acids of ileal mucosa were analyzed using gas chromatography-mass selective detector. RESULTS: Eicosapentaenoic acid significantly increased survival time after heatstroke. Eicosapentaenoic acid significantly decreased intestinal permeability and plasma endotoxin levels. Eicosapentaenoic acid effectively attenuated the heatstroke-induced disruption of the intestinal structure and improved the histology score, whereas DHA was less effective, and corn oil was ineffective. Pretreatment with EPA also increased expression of occludin and ZO-1 to effectively prevent TJ disruption. Eicosapentaenoic acid pretreatment enriched itself in the membrane of intestinal cells. CONCLUSIONS: Our results indicate that EPA pretreatment is more effective than DHA pretreatment in attenuating heat-induced intestinal dysfunction and preventing TJ damage. Enhanced expression of TJ proteins that support the epithelial barrier integrity may be important for maintaining a functional intestinal barrier during heatstroke.

Association of ADIPOQ polymorphisms with obesity risk: a meta-analysis.

BACKGROUND: The association between ADIPOQ polymorphisms and the risk of obesity remains controversial. We perform a comprehensive meta-analysis to clarify the current understanding of this association. METHODS: We searched for relevant studies in PubMed, Embase and Cochrane library before February 2014. The strengths of the association between ADIPOQ polymorphisms and obesity risk were estimated by odds ratios (OR) with 95% confidence intervals (CI). RESULTS: Eighteen case-control studies analyzing four SNPs (rs17300539, rs266729, rs1501299 and rs2241766) of ADIPOQ gene were eligible for the present meta-analysis. The pooling results showed that rs17300539 (2GG+GA vs. 2AA+GA: OR=0.78, 95%CI=0.69-0.89) and rs1501299 (2GG+GA vs. 2AA+GA: OR=0.89, 95%CI=0.80-0.98) were associated with obesity risk in Caucasian ethnicity. The rs266729 were associated with obesity risk in Asian ethnicity (2CC+CG vs. 2GG+GCG: OR=0.77, 95%CI=0.65-0.92). However, there were no associations between rs2241766 and the obesity risk (P>0.05). No publication bias was found among these studies (all P>0.05). CONCLUSIONS: This study suggests that ADIPOQ rs17300539 and rs1501299 are associated with risk of obesity in Caucasian ethnicity, and the rs266729 is associated with obesity risk in Asian ethnicity. However, there is no association between rs2241766 and obesity risk.