Preliminary Study On Expression Of MicroRNA In The Peripheral Blood Of Children With Sepsis

Background: Sepsis is a systemic inflammatory response syndrome(SIRS) caused by infection, the progress often rapidly, and the number ofpatients is increasing year by year. If early and continued SIRS can not takemeasures to controll effectively, especially in pediatric patients with sepsis,which can be developed into septic shock, multiple organ dysfunctionsyndrome (MODS), has become the critically cause of death in pediatricintensive care unit (PICU), while increased the financial burden on familiesof the patients. Although the diagnosis and treatment of sepsis has achievedgreat progress, the mortality rate was still higher in the last few years. It isimportant particularly to diagnosis and treatment early on sepsis. It isdifficult to identify early sepsis and SIRS caused by non-infectious factorsbecause clinical manifestations are often confused with each other. Thetraditional infection diagnostic indicators such as temperature, white bloodcell count, erythrocyte sedimentation rate are difficult to meet clinicalneeds as a result of bad sensitivity and specificity. The same does forC-reactive protein (CRP), procalcitonin (PCT) and other indicators. It is necessary to find ideal biomarkers to distinguish sepsis and SIRS and madediagnosis earlier, predict the prognosis of the disease.MicroRNA (miRNA) is a kind of small non-coding RNA, which iswidely involved in the regulation of cell growth, development,differentiation and apoptosis in all aspects of a physiological state and isclosely related to the occurrence of disease development. Previous studieshave shown that miRNA expression of serum, plasma or tissue is differentin different disease, It has confirmed miRNA levels in serum or plasma canbe used as biological markers in many diseases (such as cancer, acutemyocardial infarction, rheumatoid arthritis, diabetes, kidney disease,inflammatory bowel disease, drug-inducedliver damage, etc.) in recentyears. As a regulator of post-transcriptional gene，miRNA associated withsignaling pathways related sepsis and immune function regulation. Thisstudy is observed expression level of inflammation-related miRNA in theperipheral blood of pediatric patients with sepsis.Objective: To establish the method to extract circulating miRNA fromthe plasma and CD14+monocytes (MC) in peripheral blood. Comparisondifferential expression of circulating miRNA levels in sepsis, SIRS andhealthy control group. At the same time we make correlation analysisbetween circulating miRNA which expression differences with otherinflammatory markers levels such as TNF-α, IL-10in sepsis and exploresepsis biomarker for early diagnosis which sensitivity and specificity better than CRP, PCT.Methods: A total of35cases of sepsis patients which hospitalized inChongqing Medical University Shenzhen Children’s Hospital PICU fromAugust2012to March2013,23males and12females, with an average ageof1.82±1.17years. They divided into general sepsis group (n=19) andsevere sepsis group (n=16) according to the severity of disease. We select15patients suffer surgical trauma as SIRS group and15healthy childrenwho made medical examination in our hospital the same time as the controlgroup. Six miRNA(miR-21, miR-125b, miR-132, miR-146a, miR-155,miR-223) closely related to the inflammatory response were enrolled,expression levels of miRNA in plasma and CD14+MC in patients withsepsis, SIRS and healthy control group was detected using real-timequantitative PCR (qRT-PCR) technology. The levels of TNF-α and IL-10inplasma was detected by CBA technology. Sensitivity and specificity werecompared between the expressions of miRNA with clinical laboratoryindicators of CRP, PCT using the ROC area under the curve (AUC).Results: There was no statistically significant difference between ageand gender (P>0.05). We successfully extracted miRNA from plasma andCD14+MC in peripheral blood samples. MiR146a, miR-223in plasma andmiR-146a in peripheral blood CD14+MC were upregulated in sepsiscompared with SIRS group and the control group, while miR-223downregulated in CD14+MC, the differences were statistical significance (P<0.05). The level of TNF-α in SIRS group is increased compared withthe control group and TNF-α levels elevated on the general sepsis groupthan severe sepsis group. The IL-10, IL-10/TNF-α level in sepsis wereincreased than SIRS group and the healthy control group. There arenegatively correlated between IL-10, IL-10/TNF-α with miR-223levels inCD14+MC and positively correlated with miR-146a levels in plasma andCD14+MC. AUC of miR-223in CD14+MC and miR-146a in plasma(0.750and0.813) are greater than CRP (0.555) and PCT (0.683). Thefomer sensitivity is62.86%and specificity is80%, the latter sensitivity is51.43%and specificity is100%.Conclusions: MiRNA in plasma and CD14+MC of peripheral bloodcould successfully extract and detect by qRT-PCR method. MiR-146a,miR-223in plasma and CD14+MC expressed differentially in sepsisrelative to SIRS and healthy children, it may be used as biomarkers forearly diagnosis in children with sepsis.