OBJECTIVE To investigate the feasibility and application of enzyme-linked bridging assay(ELBA)method to the pharmacokinetic evaluation of antisense strand siRNA drug. METHODS Antisense strand RNAs were diluted in LNCap cell lysates from 5 to 50 000 pmol·L-1 to construct the quantification curves. We transfected the intact double-strand siRNA at a final concentration 100 nmol·L-1 targeting Polo-like kinase into the LNCap cells and investigated the specificity of ELBA quantitating the siRNA antisense strand in cell supernatant,cell lysates and RNA-induced silencing complex( RlSC). Quantification curves were constructed and validated in biological matrices such as plasma (5-25 000 pmol·L-1 )and multiple tissues(liver,heart,spleen,and kidneys)(3-6250 pmol·L-1 ). The prostate specific membrane antigen aptamer siRNA delivery system with the intact siRNA concentration of 15 nmol·kg-1 was prepared. The siRNAs were delivered into the LNCap xenogrant tumor model in C57 mice by tail vein injection. The concentration of siRNA antisense strand was determined in plasma and tissues 30 min post administration by ELBA. RESULTS The quantitative range of antisense strand siRNA in cell lysates was 5-50 000 pmol·L-1 ,and ELBA method could quantify the siRNA antisense strand concentration from cell lysates and RlSC in LNCap cells transfected with double-strand siRNA. ln addition,ELBA could specifically reflect the single antisense strand concentration instead of intact siRNA double strands in plasma. The quantification range of siRNA antisense strand using ELBA in plasma was 5-25 000 pmol·L-1 and 3-3125 pmol·L-1 in tissues. About 30 min post administration of PSMA aptamer-siRNA,the antisense strand of siRNA was distributed mainly to the tumor,liver,kidneys,blood and spleen in sequence. The distribution profile might be attributed to the target delivery and siRNA pharma-codynamics. CONCLUSION The ELBA method is successfully applied to the siRNA antisense strand pharmacokinetic evaluation,which provides an alternative for pharmacokinetic studies of siRNA-based drugs.

Dysimmunity plays a key role in diabetes, especially type 1 diabetes mellitus. Islet-specific autoantibodies (ISAs) have been used as diagnostic markers for different phenotypic classifications of diabetes. This study was aimed to explore the relationships between ISA titers and the clinical characteristics of diabetic patients.

Background: Dysimmunity plays a key role in diabetes, especially type 1 diabetes mellitus. Islet-specific autoantibodies (ISAs) have been used as diagnostic markers for different phenotypic classifications of diabetes. This study was aimed to explore the relationships between ISA titers and the clinical characteristics of diabetic patients. Methods: A total of 509 diabetic patients admitted to Department of Endocrinology and Metabolism at the Affiliated Hospital of Nantong University were recruited. Anthropometric parameters, serum biochemical index, glycosylated hemoglobin, urinary microalbumin/creatinine ratio, ISAs, fat mass, and islet β-cell function were measured. Multiple linear regression analysis was performed to identify relationships between ISA titers and clinical characteristics. Results: Compared with autoantibody negative group, blood pressure, weight, total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), visceral fat mass, fasting C-peptide (FCP), 120 minutes C-peptide (120minCP) and area under C-peptide curve (AUCCP) of patients in either autoantibody positive or glutamate decarboxylase antibody (GADA) positive group were lower.Body mass index (BMI), waist circumference, triglycerides (TGs), body fat mass of patients in either autoantibody positive group were lower than autoantibody negative group. GADA titer negatively correlated with TC, LDL-C, FCP, 120minCP, and AUCCP.The islet cell antibody and insulin autoantibody titers both negatively correlated with body weight, BMI, TC, TG, and LDL-C. After adjusting confounders, multiple linear regression analysis showed that LDL-C and FCP negatively correlated with GADA titer. Conclusion Diabetic patients with a high ISA titer, especially GADA titer, have worse islet β-cell function, but less abdominal obesity and fewer features of the metabolic syndrome.

Objective To evaluate the outcomes of acute lower extremity deep venous thrombosis (DVT) in bedridden patients by vascular ultrasound monitoring and analyze the influencing factors. Methods Bedridden patients with acute lower extremity DVT diagnosed by vascular ultrasound were recruited from March 2013 to August 2017. All patients accepted the lower extremity deep venous ultrasound after 2 weeks, and the outcomes of DVT were observed. The patients were divided into DVT improvement group and non-improvement group. The relationships between the factors including clinical characteristics, past history, laboratory examinations, medications and outcomes of DVT were analyzed. The statistically significant influencing factors were taken into the multi factor Logistic regression analysis. Results There were 134 bedridden patients with lower extremity DVT, including 85 males and 49 females, with an average age of (63.9 ± 18.1) years. DVT was improved in 32.8% (44/134) patients, and was not improved in 67.2% (90/134) patients. Anticoagulation therapy, increased international normalized ratio, and delayed thrombin time were associated with improved DVT. Non traumatic causes of bed rest and dehydration were associated with not improved DVT. Anticoagulation therapy ( P＝0.021, OR＝2.729, 95% CI 1.162-6.410) and dehydration treatment ( P＝0.032, OR＝0.240, 95% CI 0.065-0.882) were independent risk factors for improvement of lower extremity DVT. Conclusions Anticoagulation therapy may improve DVT of the lower extremities, and dehydration may aggravate the DVT of the lower extremities. Vascular ultrasound can conveniently and dynamically monitor the changes of the lower limbs DVT in bedridden patients.

Dysimmunity plays a key role in diabetes, especially type 1 diabetes mellitus. Islet-specific autoantibodies (ISAs) have been used as diagnostic markers for different phenotypic classifications of diabetes. This study was aimed to explore the relationships between ISA titers and the clinical characteristics of diabetic patients.

Background: Fatty acid-binding protein 4 (FABP4) has been demonstrated to be a predictor of early diabetic nephropathy. However, little is known about the relationship between FABP4 and diabetic retinopathy (DR). This study explored the value of FABP4 as a biomarker of DR in patients with type 2 diabetes mellitus (T2DM). Methods: A total of 238 subjects were enrolled, including 20 healthy controls and 218 T2DM patients. Serum FABP4 levels were measured using a sandwich enzyme-linked immunosorbent assay. The grade of DR was determined using fundus fluorescence angiography. Based on the international classification of DR, all T2DM patients were classified into the following three subgroups: non-DR group, non-proliferative diabetic retinopathy (NPDR) group, and proliferative diabetic retinopathy (PDR) group. Multivariate logistic regression analyses were employed to assess the correlation between FABP4 levels and DR severity. Results: FABP4 correlated positively with DR severity (r=0.225, P=0.001). Receiver operating characteristic curve analysis was used to assess the diagnostic potential of FABP4 in identifying DR, with an area under the curve of 0.624 (37% sensitivity, 83.6% specificity) and an optimum cut-off value of 76.4 μg/L. Multivariate logistic regression model including FABP4 as a categorized binary variable using the cut-off value of 76.4 μg/L showed that the concentration of FABP4 above the cut-off value increased the risk of NPDR (odds ratio [OR], 3.231; 95% confidence interval [CI], 1.574 to 6.632; P=0.001) and PDR (OR, 3.689; 95% CI, 1.306 to 10.424; P=0.014). Conclusion FABP4 may be used as a serum biomarker for the diagnosis of DR.

Objective: To study the clinical features and treatment of children with influenza, and provide evidence for clinical screening and appropriate treatment timely. Methods: Epidemiology, clinical manifestations, laboratory features and drug therapy of 978 pediatric patients with influenza in Beijing New Century International Children's Hospital in 2014 were analyzed retrospectively. Results: Among the 978 pediatric patients with influenza, 90.8%were outpatients, while 9.92% were inpatients. The incidence was the highest in winter (85.28%). The age of most cases ranged from 1 to 5 years (57.16%). The cases with type A influenza accounted for 81.29%. High fever (99.59%) and cough (85.89%) were the two main symptoms. The average count of WBC was 6.86±2.68×10⁹/L, lymphocyte percentage was lower than the proportion of neutrophils. CRP was normal (66.16%) or slightly-increased (19.00± 15.12 mg/l). Compared with type A influenza, digestive tract symptoms were more common in cases with type B (P=0.000). Analysis of 97 hospitalized cases: the main diagnosis was lower respiratory tract infection (71.13%). Nearly 23.71% cases had been detected to have combined infection with other pathogenic agents. Course of fever in cases who started taking oseltamivir after fever of 48 hours was significantly longer than those who took within 48 hours (P=0.000). Seven severe pneumonia cases were mainly in young children, cases with underlying disease or mixed infection, they were all cured by active and comprehensive therapy. Conclusions Influenza in children is characterized by high fever and cough, digestive tract symptoms frequently occurred in patients with type B influenza. Early application of oseltamivir could signficantly shorten the period of fever. Severe cases should be given active and comprehensive treatments.