Differential Expression Pattern of Exosome Long Non-Coding RNAs (lncRNAs) and MicroRNAs (miRNAs) in Vascular Endothelial Cells Under Heat Stroke.

BACKGROUND Heat stroke is a life-threatening disease which is characterized by a high body temperature and multiple organ dysfunction syndrome. Vascular endothelial cell injury is a main feature of heat stroke. Little is known about the long noncoding RNA (lncRNA) and microRNA (miRNA) expression alternation in endothelial cell exosomes related to heat stroke. The aim of this study was to explore the changes of lncRNAs and miRNAs expression pattern in exosomes derived from vascular endothelial cells under heat stroke temperature conditions. MATERIAL AND METHODS Cultured medium exosomes from HUVECs (human vascular endothelial cells) either under normal temperature or heat stroke temperature conditions were harvested; then RNA was extracted and the lncRNAs and miRNAs were analyzed by high throughput sequencing. RESULTS Ten significantly upregulated and 10 downregulated lncRNAs were identified in exosomes derived from heat stroke temperature treated cells. Furthermore, GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) analyses were used to evaluate the signaling pathway of differential expressions in lncRNAs. Finally, the interaction network of lncRNAs-miRNAs-mRNA was uncovered using ceRNA (competing endogenous RNA) principle via prediction software. CONCLUSIONS These results indicate that the identified lncRNAs and miRNAs in endothelial cell exosomes might serve as non-invasive biomarkers for heat stroke.

Peripheral T-lymphocyte and natural killer cell population imbalance is associated with septic encephalopathy in patients with severe sepsis.

Septic encephalopathy (SE) is a diffuse cerebral dysfunction resulting from a systemic inflammatory response, and is associated with an increased risk of mortality. The pathogenesis of SE is complex and multifactorial, but unregulated immune imbalance may be an important factor. The current retrospective study examined the clinical data of 86 patients with severe sepsis who were admitted to the Intensive Care Unit at Zhongshan Hospital, Xiamen University (Xiamen, China) from January, 2014 to January, 2015. The patients were assigned to SE and non-SE patient groups according to the presence or absence of SE. The proportion of T-lymphocyte subsets and natural killer (NK) cells in the immune cell population, representing the function of the immune system, were analyzed for their association with SE and compared with other clinical predictors and biomarkers. The incidence of SE in the patients was 39.5%, and this group demonstrated higher mortality rates (38 vs. 10% in non-SE patients; P=0.001). Univariate analysis revealed that the SE patients reported a lower percentage of cluster of differentiation 4+(CD4+) T-lymphocytes (51.67±7.12 vs. 60.72±3.70% in non-SE patients; P<0.01), a lower CD4+/cluster of differentiation 8+(CD8+) ratio (1.59±0.32 vs. 1.85±0.26% in non-SE patients; P<0.01) and a higher percentage of NK cells (11.80±1.44 vs. 9.19±2.36% in non-SE patients; P<0.01). Using a binary logistic regression model, the Acute Physiology and Chronic Health Evaluation II score and the percentage of CD4+ T-lymphocytes were demonstrated to be independently associated with SE (respectively, P=0.012 and OR, 4.763; P=0.005 and OR, 0.810). An area under the curve analysis of a receiver operating characteristic curve of the two indicators revealed that these were equally powerful measures in prediction of SE (Z=1.247, P>0.05). The present results confirm that SE leads to higher mortality in patients with severe sepsis, and demonstrate that immune imbalance is important in the development of SE. The proportion of CD4+ T-lymphocytes present were revealed in the current study to be a powerful predictor of SE in patients with severe sepsis.

Diagnosis of overt disseminated intravascular coagulation in critically Ill adults by Sonoclot coagulation analysis.

Disseminated intravascular coagulation (DIC) diagnosis is hampered by the limited availability of reliable clinical or laboratory tests. Currently available tests are time consuming and expensive. We investigated whether coagulation and platelet function analyses using the Sonoclot system were suitable for overt DIC diagnosis in critically ill adults. This was an observational diagnostic study performed in 498 patients presenting with an underlying disorder associated with DIC. Overt DIC patients were identified according to an International Society on Thrombosis and Hemostasis (ISTH) score of >5. Coagulation and platelet parameters were analyzed using the Sonoclot system, and compared with ISTH as the gold standard. Receiver operating characteristic curves and area under the curves were used to evaluate the value of the Sonoclot parameters. There were no differences for age or gender between the groups. Significant correlations were observed between activated clotting time (ACT) and ISTH score (r = 0.7; P < 0.001), clot rate (CR) and ISTH score (r = 0.5; P < 0.001), platelet function (PF) and ISTH score (r = -0.6; P < 0.001), and PF and platelet count (r = 0.5; P < 0.001). An ACT cut-off value of 213.5 s alone or combined with CR presented good sensitivity (76.7 and 86.8 %, respectively) and specificity (96.2 and 93.3 %, respectively). Sonoclot analysis can be performed using a point-of-care device that effectively discriminates low and high ISTH scores, and that effectively predicts coagulation dysfunction in patients with overt DIC.

Serum procalcitonin predicting mortality in exertional heatstroke.

BACKGROUND: The aim of this study was to test if Procalcitonin PCT value at the time of admission is a predictor of mortality and/or a diagnostic marker of concomitant infection in exertional heatstroke. METHODS: 68 patients with exertional heatstroke admitted to the multidisciplinary intensive care unit were studied. Serum PCT was detected by means of a specific and ultrasensitive immunoluminometric assay within 2 h of admission. The Acute Physiology and Chronic Health Evaluation (APACHE) II score was evaluated within 24 h of admission. RESULTS: There was no significant difference in PCT levels between concomitant infection and non-infection patients (p=0.712). Elevated PCT level in exertional heatstroke patients was associated with a more critical pathological state. PCT values in patients with multiple organ dysfunction syndrome (MODS) were significantly higher than those without MODS (p=0.007.). PCT values were also positively correlated with APACHE II scores (r=0.588, p=0.016). PCT values in non-survivors were higher than in survivors at univariate regression analysis (p=0.017). After adjusting for confounders, PCT concentration also remained an independent determinant of mortality (OR 2.98; 95% CI 1.02 to 4.41; p=0.039). Receiver operating characteristic curve for PCT concentration was located above the reference line, which shows an association with mortality. The area under the curve for PCT concentration (0.705; 95% CI 0.547 to 0.862) was statistical significantly (p=0.019). As a predictor of mortality, PCT value was inferior to APACHE II score. CONCLUSIONS: PCT value at the time of admission is an independent predictor of mortality, but maybe not a good indicator of concomitant infection in exertional heatstroke.

Early elevated HMGB1 level predicting the outcome in exertional heatstroke.

BACKGROUND: Heatstroke is generally considered as a syndrome of hyperthermia associated with systemic inflammation leading to multiorgan dysfunction. High mobility group box-1 protein (HMGB1) has recently been identified as a late mediator of systemic inflammation inducing multiorgan dysfunction. Elevation of plasma HMGB1 in heatstroke has been observed in animals, but there is no data available about its changes in heatstroke patients. The objectives of this study are to observe the time course of plasma HMGB1 changes and assess its prognostic value in patients with exertional heatstroke. METHODS: Blood samples were taken from the patients with exertional heatstroke. Plasma HMGB1 level was detected by the enzyme-linked immunosorbent assay. C-reactive protein level was measured using a fully automated IMMAGE Immunochemistry System. Secreted HMGB1 in the culture supernatant of peripheral blood monocyte was assessed by immunoblotting. Acute Physiology and Chronic Health Evaluation II score was evaluated within 24 hours of admission. RESULTS: HMGB1 released into circulation at early stage, with peak levels occurring within 6 hours to 13 hours postheatstroke. Plasma HMGB1 levels remained markedly elevated in the following 6 days postheatstroke when compared with healthy volunteers (p<0.005). Positive correlation (r=0.798, p<0.001) was found between Acute Physiology and Chronic Health Evaluation II score and HMGB1 level at admission. HMGB1 levels at admission between survivors and nonsurvivors were significantly different (p<0.001). Receiver operating curve analysis showed that at a level of 47 ng/mL, HMGB1 level at admission indicated lethality with 77.4% sensitivity and 84.2% specificity. CONCLUSIONS: HMGB1 level at admission is an indicator of the severity of illness and a useful mortality predictor in exertional heatstroke.

Pathological changes in the lung and brain of mice during heat stress and cooling treatment.

BACKGROUND: Heatstroke often leads to multiple organ dysfunction syndrome (MODS) with a death rate of 40% or a neurological morbidity of 30%. These high rates in patients with heatstroke are largely due to the progression of heat stress to MODS, resulting in no specific treatment available. This study aimed to develop a mouse model of heat stress and determine the pathological changes in the lung and brain during heat stress and cooling treatment. METHODS: A mouse model of heat stress was established in a pre-warmed incubator set at 35.5 ± 0.5°C and with a relative humidity of 60% ± 5%. Rectal temperature was monitored, and at a temperature of 39 °C, 40 °C, 41 °C, or 42 °C, the mice were sacrificed. The remaining animals were removed from the incubator and cooled at an ambient temperature of 25 ± 0.5 °C and a humidity of 35% ± 5% for 12 or 24 hours at a temperature of 41 °C or for 6 hours at a temperature of 42 °C. The control mice were sham-heated at a temperature of 25 ± 0.5 °C and a humidity of 35% ± 5%. The lungs and brains of all animals were isolated. Hematoxylin and eosin staining and light microscopy were performed to detect pathological changes. RESULTS: All mice demonstrated a uniform response to heat stress. A low degree of heat stress induced marked pathological changes of the lungs. With the rise of the temperature to 42°C, progressively greater damage to the lungs with further congestion of the lung matrix, asystematic hemorrhage of alveolar space, abscission of alveolar epithelial cells, and disappearance of pulmonary alveolus tissue structure were detected. However, absorption of congestion and hemorrhage as well as recovery of pulmonary alveolus tissue structure was observed following cooling treatment at an ambient temperature. With a low degree of heat stress, the brain only showed moderate edema. Neuronal denaturation and necrosis were detected at a temperature of 42°C. Interestingly, the lesions in the brain were further aggravated at 42 °C regardless of cooling treatment, but recovery was observed after cooling treatment at 41 °C. CONCLUSIONS: The pathological changes of the lungs and brain of mice showed distinctive lesions following heat stress and cooling treatment, and they were correlated with the time and duration of cooling treatment. The results of this study are helpful for further study of the mechanisms linking heatstroke.

[Screening and identifying differentially expressed proteins in LST-R1 cells].

BACKGROUND & OBJECTIVE: Colorectal laterally spreading tumor (CLST) rarely invades deeply but always laterally spreads along the colorectal mucosa, therefore, CLST could be used as a comparative model in studying the invasion and metastasis of colorectal cancer (CRC). This study was to identify differentially expressed proteins between a CLST cell line LST-R1 and two colorectal carcinoma cell lines SW480 and LoVo using proteomic technology. METHODS: Total proteins of LST-R1, SW480 and LoVo cells were isolated by two-dimensional electrophoresis (2-DE). Differentially expressed protein spots were analyzed with Melanie 3 software. The peptide mass fingerprints (PMFs) of differently expressed proteins were analyzed by MALDI-TOF mass spectrum. Subsequently matched proteins were searched through protein databases. RESULTS: Using pH4-7 IPG gels with 250 microg protein loading, the numbers of protein spots in 2-DE maps were 1285+/-51 in LST-R1 cells, 1184+/-47 in SW480 cells, and 1124+/-54 in LoVo cells; when with 150 microg protein loading, the numbers were 989, 935 and 893, respectively. The distribution and levels of these proteins in 2-DE maps of LST-R1, SW480 and LoVo cells were analogical which indicated CLST also expresses the protein profile of common colorectal tumors. In 2-DE maps, 96+/-7 differential protein spots were detected between LST-R1 cells and SW480 cells with 50+/-6 only expressed or obviously over-expressed in LST-R1 cells and 47+/-5 in SW480 cells; 108+/-10 differential protein spots were detected between LST-R1 cells and LoVo cells with 56+/-8 only expressed or obviously over-expressed in LST-R1 cells and 52+/-11 in LoVo cells. Nineteen differentially expressed proteins were identified among LST-R1, SW480 and LoVo cells. CONCLUSION: Nineteen differentially expressed proteins are possibly involved in laterally spreading of CLST and adhesion and invasion of CRC.

[Expression of tissue inhibitor of metalloproteinase-1 in colorectal carcinoma and its clinical implications].

OBJECTIVE: To investigate the expression and clinical implication of tissue inhibitor of metalloproteinase-1 (TIMP-1) in colorectal carcinoma. METHODS: TIMP-1 expression in 54 colorectal carcinoma was observed by SP immunohistochemical method, and the results were analyzed in relation to the clinical data of patients. RESULTS: TIMP-1 was localized on the membrane and in the cytoplasm of the enteric epithelial cells, and its expression rate was 100% in normal tissue but only 59.6% (31/52) in colorectal carcinoma tissues. In addition, the expression rate of TIMP-1 was higher in the tumor tissues without lymph node metastasis than in tissues with lymph node metastasis (P<0.05). CONCLUSION: The expression of TIMP-1 is inversely correlated to lymph node metastasis of colorectal carcinoma, and decreased TIMP-1 expression may play a role in the progression of colorectal carcinoma.

[Endoscopic and histopathological features of serrated adenoma of large intestine:an analysis of 71 cases].

OBJECTIVE: To explore the endoscopic and histopathological morphology of large intestinal serrated adenomas (SA). METHODS: The endoscopic and pathological data of 71 SA patients, diagnosed in the Digestive Endoscopy Center, Nanfang Hospital from January 2002 to July 2005, were analyzed retrospectively. RESULTS: Forty-seven of the 71 serrated adenomas were protruded (sessile 23, semipedunculated 5, pedunculated 23) and 24 were superficial (flat 16, laterally spreading 8). The mean sizes of the protruded and superficial SA were 10.5 mm and 16.6 mm, respectively, and both of them were frequently located in the sigmoid and rectum. Histopathologically, SA contained tubular glands in 53, tubulovillous glands in 9 and villous glands in 9 cases. Mild dysplasia was found in 47 SAs, moderate dysplasia in 22 SAs, and canceration foci in 2 SAs. The dysplasia of SAs (<10 mm) was significantly better than that of SAs (>or= 10 mm) (P< 0.01). Most IV and III L pit SAs presented villous glands (64%) and tubular glands (68%), respectively. 40% of hyperplastic polyps-like SAs, composed of tubular glands,showed II pit pattern. Atypia in II pit SAs was similar to that in IIIL pit SAs, but was worse than that in IV pit SAs. CONCLUSION: Polyps with II pit pattern possibly are SAs sometimes. SA with the common characters of neoplastic polyps,should be regarded as a new potential precancerous lesion.

HBsAg loading on dendritic cells in patients with chronic hepatitis B: expressions of phenotypic molecules.

BACKGROUND: The antigen reducing ability of dendritic cells (DCs), a kind of antigen presenting cells (APCs) initiating immune response, is associated with the specific immune tolerance of chronic hepatitis B (CHB) patients. However, the dysfunction of DCs can be possibly reversed by the stimulation of antigen peptides. In this study, DCs were cultured from peripheral blood monocytes (PBMCs) in patients with CHB in vitro, and the expression of phenotypic molecules on DCs loaded by different concentrations of HBsAg was observed. METHODS: Forty patients with CHB were divided randomly into 4 groups (10 patients in each group). PBMCs were isolated, and DCs were cultured after addition of granulocyte/macrophage colony-stimulating factor (GM-CSF) and interleukin 4 (IL-4). On the 9th day, DCs of the experimental groups were loaded at HBsAg concentrations of 2.5 mg/L, 5 mg/L and 10 mg/L for 24 hours, whereas those of the control group were not loaded. An electron microscope was used to analyze the morphological changes of the DCs. The expression of phenotypic molecules on DCs in different groups was detected with flow cytometry. RESULTS: A combination of GM-CSF and IL-4 produced DCs from PBMCs in patients with CHB after being cultured for 9 days, whose morphological changes were tested by an electron microscope. The expression of phenotypic molecules on DCs in the control group was as low as CD83 (8.02+/-3.99)%, CD80 (8.77+/-2.06)%, and MHC-DR (14.05+/-2.66)%. Loaded by different concentrations of HBsAg, the up-regulation of phenotypic molecules on DCs was found, with CD83 (18.35+/-2.93)%, CD80 (42.63+/-7.15)% and MHC-DR (47.49+/-6.59)% in 2.5 mg/L HBsAg loading group, CD83 (17.88+/-3.12)%, CD80 (45.24+/-10.93)% and MHC-DR (47.07+/-8.52)% in 5 mg/L HBsAg loading group and CD83 (16.74+/-2.86)%, CD80 (44.59+/-6.99)% and MHC-DR (48.59+/-7.42)% in 10 mg/L HBsAg loading group, respectively. Compared with the control group, the phenotypic molecules in the experimental groups were all different significantly (P<0.01), but among them, there were no differences (P>0.05). CONCLUSIONS: DCs cultured from PBMCs in the patients with CHB under the conditions of GM-CSF and IL-4 present on the typical dendritic morphology but are immature for expressing low phenotypic molecules. Loaded by different concentrations of HBsAg, the immature DCs can differentiate to mature DCs for expressing increasing phenotypic molecules.