Objective To compare the two kinds of purification method for purifying recombinant human cardiac troponin I(cT-nI)to obtain the stable cTnI and promote the study of cTnI diagnosis standardization.Methods The cTnI inclusion body was ob-tained by the ultrasonic broken engineering,after washing by 2% Tritonx-100,2M urea,dissolved in 8M urea,then purified by the column refolding on CM-FF and the dilution refolding respectively.The cTnI yields were compared between the two kinds of meth-od and the stability at 4 ℃,20 ℃,-80 ℃ and on the freeze-dried condition was compared.Then the purification method to effi-ciently obtain the stable cTnI was established.Results The protein about 2 mg and 1.4 mg could be obtained by CM-FF on the col-umn refolding and the dilution refolding from 0.1 g of wet inclusion body,respectively.The former method had the short cycle and high efficiency.The cTnI purified by the column refolding on CM-FF was more stable at 4 ℃,20 ℃,-80 ℃ and on the freeze-dried condition.Conclusion The column refolding on CM-FF is more stable and highly efficient in purification of cTnI than the dilution refolding.

Objective:To investigate the etiology of pleural effusion in hospitalized children in Beijing Children′s Hospital.Methods:Clinical information of children with pleural effusion admitted to Beijing Children′s Hospital Affiliated to Capital Medical University from January 2016 to December 2018 was retrospectively analyzed.According to the etiology, the children were divided into infection group (parapneumonic pleural effusion, tuberculous pleurisy and empyema) and non infection group.According to the age, the children were further divided into ≤ 3 years old, >3-7 years old and > 7 years old groups.Classification of statistics was performed, and the etiology of pleural effusion were retrospectively analyzed.Results:Among the 1 165 children with pleural effusion, 746 cases(64.0%) were infected with pleural effusion, 697 cases (697/746, 93.4%) of who were parapneumonic effusion.In patients with parapneumonic effusion, 457 cases (61.3%) had Mycoplasma pneumonia (MP) infection.Infectious pleural effusion was more common in children >7 years old(339/479 cases, 70.8%), while non-infectious pleural effusion was prevalent in children under 3 years old(188/324 cases, 58.0%). The difference was statistically significant ( χ2=96.33, P<0.05). Among the patients with non-infectious pleural effusion, 239 cases (239/419 cases, 57.0%) had multi-system diseases and 97 cases (97/419 cases, 23.2%) had malignant pleural effusion.All the 18 deaths were non-infectious pleural effusion. Conclusions:The leading reason for pleural effusion in children is infection.The most prevalent symptom is parapneumonic effusion, which is mainly caused by MP.

Objective To investigate the value of strong ion gap (SIG) in predicting the severity of acute pancreatitis (AP) based on the revised Atlanta classification. Methods A total of 133 patients with AP admitted from January, 2015 to December, 2016 were enrolled. Of them, there were 55 with mild AP (MAP), 52 with moderately severe AP (MSAP) and 26 with severe AP (SAP). All patients with AP conformed to the diagnostic criteria of Guidelines or Diagnosis and Treatment of Acute Pancreatitis set in 2014 in China. Patients with other underlying diseases that might influence the clinical outcome were excluded, including those with diabetic ketoacidosis, chronic renal failure and other disorders. The changes in blood SIG levels in each group were observed. The correlations between SIG and acute physiology, chronic health evaluation (APACHE) Ⅱ score, Ranson score and length of hospital stay were analyzed. The receiver operating characteristic curves (ROC) were plotted to determine the efficiency of SIG, Scr, APACHE Ⅱ score, and Ranson score for predicting the severity of acute pancreatitis. Results The level of SIG in the SAP group was the highest, followed by the MSAP group and the lowest in the MAP group.There were significant differences in pairwise comparisons (P<0.01). The correlations between SIG and APACHE Ⅱ score (r=0.567, P<0.01), Ranson score (r=0.502, P<0.01), and length of hospital stay were positive (r=0.589, P<0.01). There was no statistical difference in the area under curve (AUC) between SIG and APACHE Ⅱ score (0.874±0.029 vs.0.895±0.025, P>0.05) and as well as Ranson score (0.874±0.029 vs. 0.890±0.027, P>0.05) for predicting moderately-severe acute pancreatitis, but SIG was superior to Scr (0.874±0.029 vs. 0.735±0.043, P<0.01). There was a significant difference in AUC between SIG and Scr (0.910±0.030 vs. 0.755±0.054, P<0.01), but no statistical differences between SIG and APACHE Ⅱ score (0.910±0.030 vs. 0.867±0.034, P>0.05) and Ranson score (0.910±0.030 vs. 0.871±0.032, P>0.05) for predicting severe acute pancreatitis. Conclusion SIG has important clinical significance for predicting the severity of acute pancreatitis.

OBJECTIVETo investigate the effects of bufalin in reversing hepatocyte growth factor (HGF)-induced resistance to afatinib in H1975 lung cancer cells, and explore its possible mechanism.

METHODSThe afatinib-resistant H1975 lung cancer cells (H1975AR) were induced by exogenous HGF and transfected with recombinant adenoviral vector Ad-HGF-GFP. The cytostatic effects of bufalin, afatinib and bufalin plus afatinib on H1975AR cells were evaluated by MTT assay. The impact of combined therapy with bufalin and afatinib on invasion of H1975AR cells was determined by transwell migration assay. The concentrations of HGF in the culture supernatants of H1975/Vec and H1975/HGF cells were determined by ELISA. The expression of EGFR, cMET and EMT signal pathway-related proteins in H1975AR cells treated with bufalin, afatinib and bufalin plus afatinib were detected by Western blot.

RESULTSThe results of MTT assay showed that afatinib did not inhibit the growth of H1975 cells, but after 72 h of the combined treatment with bufalin and afatinib and in the presence of HGF, the growth rate of H1975 cells was (38.67 ± 8.76)%, significantly lower than the growth rate of (63.45 ± 12.65)% in the H1975 cells treated with HGF alone (P < 0.05). The results of transwell migration assay showed that in the presence of HGF, afatinib plus bufalin combination therapy markedly decreased the number of invaded H1975 cells through the Matrigel chamber (48.98 ± 11.43), significantly lower than the 118.92 ± 37.29 of afatinib-treated or the 88.84 ± 19.53 of bufalin-treated cells (P < 0.05 for all). The result of ELISA showed that H1975/HGF cells secreted high levels of HGF, and afatinib and bufalin had no effect on the HGF secretion in H1975/HGF cells. The results of Western blot analysis showed that the expression of p-EGFR, p-cMet, p-AKT, p-ERK, vimentin and snail in H1975AR cells treated with bufalin puls afatinb was down-regulated markedly, and the expression of E-cadherin was up-regulated markedly.

CONCLUSIONSCombination of bufalin and afatinib strongly inhibits the growth of H1975AR lung cancer cells and decreases their invasion ability. The possible mechanism of combined treatment with bufalin and afatinib may be related to the blocking of cMet/PI3K/AKT and cMet/MAPK/ERK pathways and inhibiting of epithelial-mesenchymal transition.

Antineoplastic Agents ; pharmacology ; Antineoplastic Combined Chemotherapy Protocols ; pharmacology ; Bufanolides ; pharmacology ; Cadherins ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Coloring Agents ; Drug Resistance, Neoplasm ; drug effects ; Epithelial-Mesenchymal Transition ; drug effects ; Hepatocyte Growth Factor ; pharmacology ; Humans ; Lung Neoplasms ; drug therapy ; metabolism ; pathology ; MAP Kinase Signaling System ; Neoplasm Proteins ; metabolism ; Phosphatidylinositol 3-Kinases ; Quinazolines ; pharmacology ; Receptor, Epidermal Growth Factor ; Signal Transduction ; Tetrazolium Salts ; Thiazoles

Objective:To explore how electrical stimulation of the sciatic nerve affects learning and memory in cases of chronic cerebral hypoperfusion and its mechanism.Methods:Thirty-two Sprague-Dawley rats were randomly divided into a normal group, a sham-operation group, a model group and a stimulation group, each of 8. In the model and stimulation groups a chronic cerebral hypoperfusion model was established using the modified 2-VO method. The stimulation group then received electrical stimulation of the sciatic nerve for 4 weeks. Afterward all of the rats′ learning and memory were tested using the Morris water maze. HE staining and immunohistochemistry were used to observe any morphological change in the hippocampus and the expression of neuron-specific enolase (NSE) and vascular endothelial growth factor (VEGF).Results:The model and stimulation groups demonstrated significantly longer escape latency, fewer platform crossing times and shorter target quadrant times than the normal and sham-ope-ration groups before the intervention. All of these indicators had improved after the intervention. According to the HE staining, neuronal damage in the hippocampus was aggravated significantly in the model group compared with the normal and sham-operation groups, while the degree of damage was reduced in the stimulation group. Moreover, the immunohistochemistry results suggested that the expressions of NSE and VEGF were reduced significantly in the model group compared with the normal and sham-operation groups, while in the stimulation group they had increased significantly compared with the model group.Conclusions:Electrical stimulation of the sciatic nerve can improve learning and memory in cases of chronic cerebral hypoperfusion, at least in rats. It is probably related to the increased expression of VEGF in the hippocampus.

Objective:To investigate the clinical features of Kawasaki disease complicated with multiple pulmonary nodules and to summarize the experience of diagnosis and treatment.Methods:Clinical data of 2 cases of Kawasaki disease complicated with multiple pulmonary nodules admitted to the Department of Respiratory, Beijing Children′s Hospital in 2019 were retrospectively analyzed, and relevant literature was reviewed.Results:The 2 cases were both 3-month-old infants, with fever as the main manifestation.Both were consistent with the diagnosis of Kawasaki disease, and the respiratory symptoms were mild or not obvious.Radiological examination revealed multiple pulmonary nodules, and the results of etiology, immunology and tumor-related exams were all negative.In the follow up stage after treatment, pulmonary nodules gradually disappeared.Among 10 cases of Kawasaki disease with pulmonary nodules (8 cases from previous studies and 2 cases from this report), 90% were 3-6-month-old babies, 80% were diagnosed with incomplete Kawasaki disease, and all patients presented more than 2 nodules in the lungs, with irregular distribution and different sizes.Besides, 80% of the 10 cases had coronary artery involvement of varying degrees.One patient died in the acute disease phase, while pulmonary nodules of the remaining patients disappeared in 12 days to 1 year.Conclusions:Kawasaki disease may be complicated with multiple pulmonary nodules, especially in young infants.Most of the patients present with incomplete Kawasaki disease and are prone to coronary artery lesions, which may be related to pulmonary vascular involvement caused by Kawasaki disease.The prognosis is mostly good, and pulmonary nodules can disappear naturally with the treatment and course of Kawasaki disease.

Primary immunodeficiency diseases (PIDs) refer to immune function decrease and deficiency or immune regulation function imbalance resulted from immune cell or immunomolecular defects caused by single gene mutation.Children with PIDs are prone to develop infectious diseases.Because the immune mechanisms of hosts infected by different pathogens infecting vary, different types of PIDs are relatively susceptible to different pathogens.A number of studies have indicated that some PIDs children are more likely to develop Bacilles Calmétte-Guerin infection, severe tuberculosis and non- Tuberculous mycobacteria severe infection.In this review, several major primary immunodeficiency diseases closely related to the susceptibility of mycobacteria were summarized, including chronic granulomatous disease, severe combined immunodeficiency disease, mendelian susceptibility to mycobacterial disease, and high IgM syndrome, in order to provide a theoretical basis for early identification of such diseases.

【Objective】 To analyze the detection characteristics of a novel serum marker, hepatitis B core-associated antigen (HBcrAg), in the HBsAg-/HBV DNA+ blood donors in Wuxi. 【Methods】 A total of 37 previous HBsAg-/HBV DNA+ blood donors were followed up by telephone and their serum was obtained, and the serum of 22 HBsAg-/HBV DNA+ blood donors was detected by electrochemiluminescence and real-time PCR nucleic acid screening as the OBI group for HBcrAg enzyme-linked immunosorbent assay(ELISA). The serum of 20 healthy blood donors who underwent dual ELISA and one nucleic acid testing(NAT) was selected as the healthy control group, and the serum of 20 patients with chronic hepatitis B who were clinically diagnosed by Wuxi Fifth People's Hospital was selected as the experimental CHB group, and HBcrAg ELISA was detected respectively. The correlation analysis between HBcrAg and HBeAb, HBcAb, ALT and HBV DNA in the OBI group was performed. 【Results】 Thirty-seven blood samples were detected by chemiluminescence for HBsAg and NAT, and 22 HBsAg-/HBV DNA+ samples were detected in the OBI group, with a detection rate of 59.46%. The serum HBcrAg expression content (ng/mL) between the OBI group, the healthy control group and the CHB group were (0.92±0.13), (0.47±0.09) and (1.14±0.23), respectively, and the differences were statistically significant (P<0.05), the expression of HBcrAg in the OBI group and CHB group was higher than that in the healthy control group (P<0.05). There was no correlation between HBcrAg and HBeAb, HBcAb, ALT and HBV DNA indexes in the OBI group (P>0.05). 【Conclusion】 The expression of HBcrAg in the OBI group and CHB group was higher than that in the healthy control group, and the serum HBcrAg was not correlated with HBeAb, HBcAb, ALT and HBV DNA to a certain extent. HBcrAg has a good application prospect in screening HBsAg-/HBV DNA+ blood donors.

Objective:To investigate epidemiological characteristics and macrolide-resistance of hospitalized children with Mycoplasma pneumoniae (MP) infections in Beijing from 2016 to 2019, so as to provide basis for the prevention and treatment of pediatric Mycoplasma pneumoniae pneumonia (MPP).Methods:The clinical data were analyzed retrospectively from 8 691 children hospitalized with community acquired pneumonia in Beijing Children′s Hospital between January 2016 and September 2019.MP RNA was detected by simultaneous amplification and testing (SAT), and macrolide resistance of MP was examined by MP and macrolide-resistant isolate diagnostic kit (PCR with fluorescence probes). Chi- square test was used for categorical analysis. Results:Among 8 691 cases detected by SAT, the overall detection rate of MP was 28.10% (2 442/8 691 cases). The detection rates of MP from 2016 to 2019 were 26.23%, 31.36%, 27.84 % and 26.57%, respectively.The detection rate of MP in 2017 was significantly higher than that in other years ( χ2=16.11, P<0.05). The detection rate of MP in females was 29.65%(1 107/3 733 cases), which was evidently higher than that in males 26.93%(1 335/4 958 cases) ( χ2=7.85, P<0.05). The positive rates of MP in summer[32.21% (726/2 254 cases)] and autumn[39.76%(852/2 143 cases)] were significantly higher than those in spring[17.00% (327/1 924 cases)] and winter[22.66%(537/2 370 cases)] ( χ2=315.15, P<0.001). The percentages of MP were 35.06%(732/2 088 cases) in preschoolers and 37.71%(1 160/3 076 cases) in school-age children, which were significantly higher than 11.20%(232/2 072 cases) in infants and 22.01% (318/1 445 cases) in toddlers ( χ2=509.89, P<0.001). Macrolide resistance detection was conducted in 1 524 patients by fluorescent PCR.Among them, 1 386 patients were positive for drug resistance, and the positive rate was 90.94%.The prevalence of macrolide-resistant MP from 2016 to 2019 were 88.19%, 90.93%, 90.56% and 92.90%, respectively.Macrolide-resistant rates were not related with gender, age and season. Conclusions:MP can be detected in all seasons, but most prevalently in summer and autumn.Girls are more prone to MP infections than boys.The detection rate of MP increases with age, and the positive rate is higher in preschoolers and school-age children.During the 4-year study period, the drug resistant rate of MP remain high.