Objective To establish a simple and rapid method for the determination of biological activity of recombinant human tumor necrosis factor receptorⅡ(rhTNFRⅡ) Fc fusion protein(rhTNFRⅡ-Fc) products, and to verify and apply the method, in order to lay a foundation for the process stability evaluation of this kind of products.Methods Using HEK293TNF-κB-Luc transgenic cells, different concentrations of rhTNFRⅡ-Fc fusion protein can inhibit the transcriptional activation of nuclear factor(NF)-kB induced by TNF-α in transgenic cells to different degrees. The biological activity of rhTNFRⅡ-Fc was detected by Bright-Glo~(TM)-Luciferase Assay System, the experimental parameters were optimized, and the specificity,accuracy, linearity and precision of the method were validated. Using the international standard for biological activity assay of rhTNFRⅡ-Fc fusion protein products as the reference, this method was used to determine the specific activity of rhTNFRⅡ-Fc fusion protein products from three different domestic enterprises(A, B and C) and the original drug Etanercept.Results rhTNFRⅡ-Fc fusion protein had a dose-effect relationship in this method and conformed to the four-parameter curve equation with R~2 greater than 0. 99. After optimization, the cell seed plate density was determined to be 4 × 10~5/mL,the action concentration of TNF-α was 10 ng/mL, the initial dilution concentration of rhTNFRⅡ-Fc was 200 ng/mL, 1∶1. 5times the dilution, the action time was 4-5 h, and the sample diluting solution was DMEM + 1% FBS. Only Etanercept inhibited the activation of NF-κB transcription in HEK293T-NF-κB-Luc cells induced by TNF-α, while other fusion proteins had no inhibition on the induction of TNF-α. The recovery samples of five different dilution groups were measured three times, and the average recoveries were(101. 54 ± 4. 63)%,(99. 67 ± 6. 41)%,(101. 20 ± 5. 58)%,(101. 44 ± 6. 80)% and(100. 72 ±6. 15)%, respectively, with the relative standard deviations(RSDs) lower than 7% and the linear fitting R~2 of 0. 999 8. The RSDs of verification for inter-plate and inter-day precision were less than 8. 0%. Using this method for determination, the specific activity of the original drug was 2. 01 × 10~6IU/mg, 2. 59 × 10~6IU/mg for enterprise A, 1. 56 × 10~6IU/mg for enterprise B, and 2. 04 × 10~6IU/mg for enterprise C.Conclusion Using HEK293T-NF-κB-Luc transgenic cells, a method for the determination of biological activity of rhTNFRⅡ-Fc fusion protein was successfully established, which is simple and rapid with strong specificity, high accuracy and good precision, and can be used for the routine detection of biological activity of this kind of products in different enterprises.

Objective To explore the influencing factors of frequent relapse primary nephritic syndrome (PNS) in children according to the statistical analysis of their clinical data and laboratory examinations. Method We collected in-hospital and out-patient medical records of children who were diagnosed PNS from Jan. 2007 to Feb. 2010 and whole clinical data were analyzed. Results In 245 cases of children with steroid sensitine nephrotic syndrome, non-frequent replapse (NFR) accounted for 82.4% and frequent relapse (FR) accounted for 17.6%. In univariate analysis, period of first relapse between initial treatment remission within 3 months , level of serum total protein and cholesterol and IgE at onset had statistical significance in FR children and NFR children, which were significant predictors of FR. The logistic regression analysis revealed that the first relapse after initial treatment remission within 3 months and increasing of serum IgE level (more than 358 IU/ml)were risk factors of FR. Among frequent relapses factors, infection, especially upper respiratory tract infection was the first (65. 1%). The relapse with motivation mainly in January to March, accounted for 30. 4%. Conclusion FR had a proportion of 17. 6% in children with PNS under regular glucocorticoid treatment. It showed seasonality feature that relapse. peak in winter and spring. Level of the serum total protein less than 40g/L and cholesterol more than 10mmol/L at onset can be predictive index of FR. Infections are the main motivation of relapse, but the relapse by irregular treatment and specific constitutions should be pay more attentions too.

Objective To investigate the effects of azithromycin on serum indicators, urine indicators, renal pathology, intercellular adhesion molecule-1 (ICAM-1), nephrin and podocalyxin in adriamycin(ADR)-induced nephropathy(ADN) rats. Methods ADR nephropathy was induced by a tail intravenous injection of ADR. The rats were randomly divided into azithromycin-treated group, prednisone -treated group, integrated treatment group, model group and control group. Serum index and 24 h urine protein were measured serially at 0, 4th, 8th weeks. The values of creatinine clearance (Ccr) were calculated. Kidney tissues were collected for microscopy observation. The expressions of nephrin, podocalyxin and ICAM-1 in renal tissue were detected by immunohistochemistry SP. Results Compared with normal control group, at the 4th week, 24 hours urine protein and albumin reached the level of ADR nephropathy model. Compared with model group, at the 8th week, 24 h urine protein, cholesterol, serum creatinine, renal pathology changes in the three treated group rats were significantly reduced (P<0.05), serum total protein, albumin and Ccr (except C group) significantly raised (P<0.05), ECM/GA and renal pathology score significantly reduced (P<0.01), ICAM-1 significantly decreased (P<0.01), nephrin and podocalyxin significantly increased. Besides, the curative effect of integrated treatment was better than other treatment group. Conclusions Azithromycin induces similar responses in ADR nephropathy as prednisone, but its early protective effect of renal function is worse than prednisone.The integrated treatment of azithromycin and prednisone has a synergistic effect, and the efficacy is superior to each drug alone.

Objective To analyze the consistency of two rapid antigen assays for the diagnosis of influenza A and B. We evaluated the clinical usefulness of silver amplification immunochromatography influenza virus detection kit.Methods Nasopharyngeal swab samples were collected from patients who were suspected of influenza at Huashan Hospital between January and February 2015. Two samples were collected from the same one patient. The samples were tested simultaneously by using IMMUNO AG Cartridge Flu AB kit (AG1) and commercial immunochromatographic assay kit, Clearview exact influenza A&B (CV). PCR method was used as reference.Results A total of 91 samples were tested, of which 7 were positive for influenza A and 53 positive for influenza B by AG1 system; 7 positive for influenza A and 50 positive for influenza B by CV system; and 8 positive for influenza A and 60 positive for influenza B by PCR. The sensitivity and specificity of the AG1 system were 87.5 % and 100 % for influenza A, and 88.3 % and 100 % for influenza B; while the CV system showed sensitivity and specficity of 87.5 % and 100 % for influenza A, 83.3 % and 100 % for influenza B. The AG1 system was 100 % consistent with the CV system in the positive rate of influenza A, and 94.3 % consistent with the CV system in the positive rate of influenza B.Conclusions The AG1 system is well consistent with the conventional immunochromatography-based diagnostic tests in diagnosis of influenza. The AG1 system is useful in earlier diagnosis of influenza due to fewer human error in result interpretation.

Background Malignant osteopetrosis is an extremely rare dense bone disease,and sometimes features ocular disease and cranial nerve palsy.This disease received high attention because of its poor prognosis.And whether the eye manifestation improved after treatment is a problem for concern.Objective This study was to clarify the clinical manifestation,treatment and prognosis of malignant osteopetrosis associated with ocular disease.Methods A retrospective study was adopted.Two children with malignant osteopetrosis associated with eye symptoms were collected from Beijing Children Hospital.The systemic and ocular medical examinations were performed on the patients,including physical examination,hematology laboratory examination,abdominal B ultrasound and bone X ray radiography,external ocular examination,flash visual evoked potential (F-VEP) and CT of orbit.Bone marrow hematopoietic stem cell transplantation was employed and 5-year following-up was cinducted on the chidren.Results The children showed increased bone density,systemic bone sclerosis,basilar thickening,abnormalities of hematology indexes,anemia,hepatosplenomegaly,optical canal stenosis and abnormality of F-VEP P2 wave.In addition,optical disc pale,facial paralysis and paralytic esotropia were seen in a female child and alternating strabismus was found in another boy.After successful treatment,the systemic symptoms remitted in both children,but the eye findings remained unchanged in the female child during the follow-up duration.However,the strabismus diminished in the male child.The optical bone canal widening to 1.9 mm 1 year and 3.2 mm 5 years after treatment in the female child.Conclusions Strabismus and eye disease are the signs of malignant infantile osteopetrosis and reflections of the impairment of the central nervous system.The pathogenetic mechanism of malignant osteopetrosisrelated eye disease is below understanding now.Early bone marrow hematopoietic stem cell transplantation for malignant osteopetrosis can offer the best chance of long-term survival and improve the prognosis of eye diseases.

Objective To explore changes of urine transferrin (TFR),micro albumin (mALb),β2 microspheres protein (MG),α1MG and N-acetylβ-D amino group (NAG) in children with kidney injury induced by cytomegalovirus (CMV) infec?tion, and the outcome after treatment. Methods Fifty children with CMV infection were used as case group, and 35 chil?dren of convalescence stage of upper respiratory tract infection were used as control group. The serum levels of creatinine (Scr), blood urea nitrogen (BUN),β2MG, liver function, CMV-IgM, CMV-PCR and brainstem auditory (BAEP), head CT and urine routine test, urine TFR, mALb,β2MG,α2MG and NAG were detected. The sputum CMV-PCR was detected if childrencombined with CMV pneumonia. Ganciclovir (5mg/kg) was given to two groups, 1/12 h i.v. for 14 d. The urine TFR,mALb,β2MG, α2MG and NAG were detected again after treatment. Results There were no significant differences in the urine mALb and TFR between the two groups. The urine levels ofα1MG, NAG andβ2MG were higher in case group than those of control group. The urine levels ofα1MG, NAG andβ2MG were decreased after 2-week treatment in case group. There were no significant differences in urine mALb and TFR before and after treatment. Conclusion The combined detection ofβ2MG,α1MG and NAG can predict CMV kidney damage in children at a early stage.

Objective:To investigate high mobility group protein 1(HMGB1), tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)levels and their clinical significance in elderly patients with viral pneumonia.Methods:One hundred and sixty elderly patients with viral pneumonia admitted to the Sixth Hospital Affiliated to Anhui Medical University were enrolled as research subjects.In addition, 40 elderly people who underwent regular physical examination were considered as the control group.Patients with viral pneumonia were divided into the low-risk group, middle-risk group and high-risk group according to CURB-65 scores and pneumonia severity index(PSI)scores.HMGB1, TNF-α and IL-6 levels were compared between different groups.The correlations of CURB-65 scores and PSI scores with HMGB1, TNF-α, IL-6 levels were analyzed.Multivariate Logistic regression analysis was used to examine influencing factors for the severity of viral pneumonia in elderly patients.Results:HMGB1, TNF-α and IL-6 levels were higher in research subjects than in the control group.As the severity of viral pneumonia increased, so did HMGB1, TNF-α and IL-6 levels(all P<0.05). HMGB1, TNF-α and IL-6 levels in the severe viral pneumonia group were significantly higher than those in the non-severe viral pneumonia group( P<0.05). HMGB1, TNF-α and IL-6 levels were positively correlated with CURB-65 scores and PSI scores(CURB-65 score: r=0.463, 0.392 and 0.497, P=0.015, 0.003 and 0.025; PSI score: r=0.596, 0.515 and 0.381, P=0.007, 0.011 and 0.009). HMGB1, TNF-α and IL-6 levels were influencing factors for the severity of viral pneumonia in elderly patients( OR=1.344, 1.422 and 1.351, P=0.006, 0.015 and 0.009). Conclusions:HMGB1, TNF-α and IL-6 levels are closely correlated with the severity of viral pneumonia and are helpful for early assessment of viral pneumonia.

Objective To summarize the clinical characteristics of and our experience in managing Klebsiella pneumoniae liver abscess.Methods The clinical data of 323 patients with bacterial liver abscess treated at three hospitals in Shanghai from January 2013 to March 2016 were analyzed retrospectively.Results Bacterial culture Klebsiella pneumoniae was identified in 70 cases.Compared with the patients with K.pneumoniae,the patients without K.pneumoniae had significantly higher prevalence of biliary tract complications (P=0.038),higher neutrophil percentage (P=0.002) and greater abscess diameter (P=0.015).However,the patients with K.pneumoniae showed relatively higher rate of treatment failure.Invasive syndrome was identified in 7 (10％) of the patients with K.pneumoniae,such as endophthalmitis,meningitis.The patients with invasive syndrome showed significantly higher prevalence of biliary tract diseases (P=0.078),more severe thrombocytopenia at early stage (P=0.004) and higher serum bilirubin level (P=0.043).The patients receiving surgical treatment (surgical operation and ultrasound-guided puncture) were associated with significantly shorter hospital stay (15.5± 8.6)d than the patients managed with medical therapy alone (20.1 ± 17.4) d (P=0.029).Conclusions K.pneumoniae is one of the most common pathogens of bacterial liver abscess.K.pneumoniae is relatively susceptible to cephalosporins and fluoroquinolones.Attention should be paid to the incidence of invasive syndrome at early stage.Antimicrobial therapy should be administered timely,especially for the patients complicated with thrombocytopenia or apparent jaundice.Ultrasound-guided percutaneous drainage can shorten hospital stay and reduce mortality.

OBJECTIVE: To detect genetic variant in a sib-pair with Finnish type congenital nephrotic syndrome (CNF). METHODS: Clinical data of the sib-pair was reviewed. Coding regions of the NPHS1 gene was analyzed for the sib-pair and both parents. RESULTS: The sister and brother respectively developed severe proteinuria 1 month and 28 days after birth, in addition with low serum albumin, hypercholesterolemia and severe edema, which were suggestive of CNF. Genetic testing identified that the sib-pair has both carried two heterozygous variants of NPHS1 gene, namely c.2605G>C (p.P869>A) and c.-61G>A, for which their father and mother were heterozygous carriers. CONCLUSION The c.2605G>C (p.869P>A) and c.-61G>A variants of the NHPS1 gene probably underlay the CNF in both sibs. The c.2605G>C(p.869P>A) was unreported previously.
Adult ; Female ; Humans ; Infant, Newborn ; Male ; Membrane Proteins/genetics* ; Mutation ; Nephrotic Syndrome/genetics* ; Siblings

Objective: To characterize the clinical features and outcomes of scimitar syndrome (SS) to aid the understanding of this syndrome. Methods: This retrospective study included 6 children who were diagnosed with SS at the pediatric cardiovascular center of Beijing Anzhen Hospital from January 2012 to September 2018. SS was diagnosed by echocardiography and confirmed by cardiac computed tomography angiography(CTA) or surgery. All data were collected to analyze the clinical and imaging characteristics and prognosis. Results: Among the 6 SS children (aged 2 months to 15 years; 5 males) weighed 5.6-17.1 kg. Three cases were infant type, the clinical manifestations were recurrent respiratory tract infection with growth retardation, including 2 cases with severe pulmonary hypertension, while 3 cases with adult type, were asymptomatic. Cardiac CTA imaging showed that the right single or all pulmonary veins descended through the diaphragm and converged into the inferior vena cava. One case was isolated infracardiac partial anomalous pulmonary venous connection (PAPVC) without other malformations. The remaining 5 cases complicated with atrial septal defect, different vascular and trachea malformations as well as spinal malformations. Vascular malformations included pulmonary veins stenosis, abnormal origin of pulmonary artery branches, collateral branches of systemic artery supplying local lung tissue, and persistent left superior vena cava. The treatment varied according to the specific location of anomalous pulmonary venous connection, the degree of pulmonary hypertension and the severity of clinical symptoms. Four cases underwent one-stage radical surgery, one case accepted intervention to occlude the collateral artery which was supplying the right lower lung and received stage Ⅱ radical surgery half a year later, and the remaining one case died from pulmonary hypertension crisis preoperation. Conclusions Isolated SS can easily miss diagnosis due to mild clinical symptoms. Patients with complicated malformations can benefit from combination therapy. SS associated with severe pulmonary hypertension can lead to early death. Therefore, early diagnosis and appropriate treatment can improve the prognosis of patients.