Design and subjects
This was a prospective, observational study. 68 patients were enrolled in Intensive Care Unit of the first affiliated hospital, China Medical University from October 1, 2013 to February 28, 2014. These patients included 49 SS and 19 SWS. 6 healthy controls were selected in the meanwhile. The Ethical Committee of the First Affiliated Hospital of China Medical University approved this study and the informed consents were signed by patients’ family members.
Sepsis and septic shock were diagnosed according to the Surviving Sepsis Campaign guidelines committee 2012(10). Exclusion criteria were pregnancy, or age < 18 years.
Variables recorded
In all the patients, age, sex, Acute Physiology and Chronic Health Evaluation Ⅱ (APACHE Ⅱ) score, Sepsis-related Organ Failure Assessment (SOFA) score, sCD40L, International Society on Thrombosis and Haemostasis (ISTH) score, Japanese Association for Acute Medicine (JAAM) score, platelets, prothrombin time international normalized ratio (PT-INR), activated partial thromboplastin time (aPTT), fibrinogen, D-Dimer, Fibrinogen degradation product (FDP), leukocytes, lactate, site of infection were recorded. Survival at 28 days was used as the endpoint of our study.
Blood samples
For patients, blood samples were drawn on the day 1, 2 and 3 at ICU admission. For healthy controls fasting blood samples were taken at 8:00 am. Serum blood samples were centrifuged at 1500 g for 15 minutes. Temperature was kept at 4℃ in all steps after blood collection. Aliquots were stored at -80℃ for further analysis. Samples were thawed only once.
Soluble CD40L determination
Serum sCD40L concentration was detected by enzyme-linked immunosorbent assays((ELISA) Uscn Life Science Inc, China). Each sample was measured and made duplicate. The detection limits was 6.1 pg/mL. The intra-test variability among the duplicate items of all samples was less than 10%.
Statistical analysis
Continuous variables were analyzed as medians and inter-quartiles (IQ) 25% and 75% (Q25-Q75), and categorical variables as frequencies and percentages. Significant differences of continuous variables between groups were analyzed using Mann-Whitney U test, and categorical variables were calculated by chi-square test and Fisher’s exact test, as appropriate. Receiver operating characteristic ( ROC) curves were used for analyzing serum sCD40L levels at days 1, 2 and 3 and selecting cut off values. Survival analysis was performed with Kaplan-Meier method curves. Multiple logistic regression analysis was applied to predict 28 days mortality. To determine the association between serum levels of sCD40L and other continuous variables at days 1, 2 and 3, Spearman's rank correlation coefficient was used. All p values <0.05 were considered statistically significant. SPSS 20.0 (SPSS Inc., Chicago, IL, USA) was used for all statistical analysis.