Objective: To investigate the efficacy of bronchoalveolar lavage (BAL) and its influence factors in the treatment of Mycoplasma pneumoniae pneumonia (MPP) with atelectasis. Methods: A retrospective case control study was performed on hospitalized MPP patients with atelectasis and received BAL in the Department of Pulmonology, Children's Hospital Zhejiang University School of Medicine from January 1, 2015 to July 31, 2017. Fever relieved in 48 hours and chest imaging improved in one week after BAL were considered effective. Clinical data, including age, sex, blood routine tests, lactate dehydrogenase (LDH), cytokines, complications, fever duration before BAL, course of disease before BAL, sputum plug, atelectasis area and its CT values of atelectasis site were collected. Student's t test, Mann-Whitney U test, or chi square test were used. Results: (1) A total of 163 patients were enrolled, including 69 boys and 94 girls, with the ratio of 1∶1.36. Their ages ranged from 6 months to 12.6 years. (2) On the day of bronchoscope, 113 patients still had fever. They were divided into effective group (n=66) and ineffective group (n=47) according to whether fever was relieved in 48 hours after BAL. The effective group were found to have less sputum plug compared with the ineffective group (33% (22/66) vs. 57% (27/47), χ²=6.499, P=0.011). The other factors such as sex, age, fever duration before BAL, course of disease before BAL, C reactive protein (CRP), LDH, IL-2, IL-4, IL-6, IL-10, TNF, IFN-γ, atelectasis area and CT value showed no significant difference between the two groups (all P>0.05). (3)A total of 122 cases had chest imaging after BAL. According to chest imaging improvement, they were divided into effective group (n=81) and ineffective group (n=41). The effective group showed lower CT value ((58±9) vs. (63±8) HU, t=-2.436, P=0.017), IL-6 and IL-10 (M(Q₁, Q₃)) (21.0 (1.9, 48.4) vs. 36.4(21.8, 93.6), 4.9 (3.7, 9.6) vs. 7.7 (4.4, 12.0) ng/L, Z=-2.387,-2.009, P=0.017, 0.045). Sex, age, fever duration before BAL, course of disease before BAL, CRP, LDH, IL-2, IL-4, TNF, IFN-γ, atelectasis area showed no significant differences between the two groups (all P>0.05). (4) Patients were divided into sputum plug group (57 cases) and non sputum plug group (106 cases) according to bronchoscopic findings. The sputum plug group showed higher LDH, CRP, IL-6, IFN-γ, incidence of pleural effusion and extrapulmonary complications (585(433, 833) vs. 369 (312, 588) U/L, 42 (19, 103) vs. 25 (12, 45) mg/L, 38.8 (22.1, 71.3) vs. 20.7 (9.2, 48.3) ng/L, 33.1 (13.5, 89.3) vs. 12.7 (6.5, 33.6) ng/L, 73.7% (42/57) vs. 52.8% (56/106), 40.4% (23/57) vs. 17.0% (18/106)), with statistically significant differences (Z=-4.865,-3.435,-3.098,-3.704, χ²= 0.010, 0.001, all P<0.01) . Additionally, fewer patients showed fever relief within 48 hours after BAL in the cases with sputum plug cases compared those without sputum plug (44.9% (22/49) vs. 68.8% (44/64), χ²= 0.011, P=0.009). Fewer patients showed chest imaging improvement within one week after BAL in the cases with sputum plug compared with those without sputum plug, but did not show significant difference (56.5% (26/46) vs. 72.4% (55/76), χ²=0.073, P=0.056). Conclusions BAL has some therapeutic effect on fever or atelectasis in MPP children complicated with atelectasis. Chest imaging improvement or fever relief may be hampered by sputum plug, increased IL-6 or IL-10.

Objective:To explore the value of nucleic acid detection methods in pharyngeal swabs in the etiological diagnosis of Mycoplasma pneumoniae (MP) infection in children. Methods:Four hundred and fifty-four (male 210, female 244) children with pneumonia in Department of Pulmonology, Children′s Hospital of Zhejiang University School of Medicine were enrolled from July, 2018 to October, 2018. Pharyngeal swabs and venous blood were obtained on the first or the second day after hospitalization. Fluorescence detection quantitative amplification of DNA, thermostatic amplification of RNA, MP culture and MP-IgM were used to detect MP simultaneously. MP infection is diagnosed if MP culture is positive or the two of the other three methods are positive. Pharyngeal swabs were acquired and detected using fluorescence quantitative amplification of DNA, thermostatic amplification of RNA and MP culture again for children with confirmed MP infection before discharge. The detection rates and quantitative changes of the three methods were compared, and χ 2 test was used for comparison among groups. Results:A total of 454 hospitalized children with pneumonia were included in this study. The detection rates of fluorescence quantitative amplification of DNA, thermostatic amplification of RNA, MP culture and MP-IgM IgM were 43.6% (198/454), 43.2% (196/454), 40.0% (180/454) and 30.6% (139/454) respectively. The difference of detection rates of the four methods was statistically significant (χ 2=20.8, P<0.05),but no significant difference between the detection rates of fluorescence quantitative amplification of DNA and thermostatic amplification of RNA was found (χ 2=0.018, P=0.900). They both had higher detection rates than MP-IgM or MP culture. MP infection is diagnosed if MP culture is positive or the two of the other three methods are positive, and two hundred and nine children were diagnosed as MP infection. In the second test of MP infection in 209 children before discharge, the positive rate of MP culture was 67.5% (141/209), with 39.4% (13/33) changed from negative to positive, and 27.3% (48/176) changed from positive to negative. The positive rate of thermostatic amplification of RNA was 82.3% (172/209), with 16.2% (31/191) turned from positive to negative, and 66.7% (12/18) turned from negative to positive. The positive rate of fluorescence quantitative amplification of DNA was 67.0% (140/209), with 52.9% (18/34) cases changed from negative to positive, and 30.3% (53/175) cases changed from positive to negative. MP-DNA load decreased in 141 cases (67.5%) and increased in 68 cases (32.5%) in the second test among the positive samples tested by fluorescence quantitative amplification of DNA. The detection rates of the four methods in the non-severe group and the severe group were similar, and the differences among the groups were not statistically significant (all P>0.05). In the second test, the proportion of changing from negative to positive in the severe group was higher than that in the non-severe group, but only the difference in the thermostatic amplification of RNA was statistically significant ( P=0.038) and the cases of changing from negative to positive of thermostatic amplification of RNA in the severe group and non-severe group are 7 and 5 respectively. Conclusions:The methods of pharyngeal swab nucleic acid detection have high sensitivity and application value in the etiological diagnosis of acute MP infection in children. The results of fluorescence quantitative amplification of DNA and thermostatic amplification of RNA are highly consistent, and they are both more advantageous than MP-IgM. Repeated testing in the acute phase is helpful to find MP infection children whose first test is negative. The load of MP-DNA did not decrease in some children in the acute stage after antibiotic treatment.

Tumor-derived exosomes play an important role in the tumor micro-environment. The exosome-derived non-coding RNAs are transmitted in the tumor microenvironment in three ways, communication between tumor cells, normal cells affecting tumor cells, and tumor cells affecting normal cells. Through these three ways, exosomal non-coding RNAs are involved in the regulation of tumor progression, affecting tumor angiogenesis, tumor invasiveness, drug resistance, stemness, tumor metabolic repro-gramming and immune escape, resulting in dual roles in promoting or inhibiting tumor development. Exosomes have a membranous structure and their contents are resistant to degradation by extracellular proteases and remain highly stable in body fluids, thus exosome-derived non-coding RNAs are expected to serve as diagnostic and prognostic indicators for a variety of cancers. In addition, exosomes can be used to deliver non-coding RNAs for targeted therapy, or to knock down or modify tumor-promoting non-coding RNAs for tumor therapy. This article reviews the function and communication mechanism of exosomal non-coding RNAs in the tumor microenvironment, including their pathways of action, effects, potential values for tumor biomarkers and treatment targets. This article also points out the issues that need to be further studied in order to promote the progress of extracellular non-coding RNAs in cancer research and their application in tumor diagnosis and treatment.
Humans ; Exosomes ; Neoplasms/genetics* ; Biomarkers, Tumor ; Body Fluids ; RNA, Untranslated/genetics* ; Tumor Microenvironment

Objective:To explore potential predictors of refractory Mycoplasma pneumoniae pneumonia (RMPP) in early stage. Methods:The prospective multicenter study was conducted in Zhejiang, China from May 1 st, 2019 to January 31 st, 2020. A total of 1 428 patients with fever >48 hours to <120 hours were studied. Their clinical data and oral pharyngeal swab samples were collected; Mycoplasma pneumoniae DNA in pharyngeal swab specimens was detected. Patients with positive Mycoplasma pneumoniae DNA results underwent a series of tests, including chest X-ray, complete blood count, C-reactive protein, lactate dehydrogenase (LDH), and procalcitonin. According to the occurrence of RMPP, the patients were divided into two groups, RMPP group and general Mycoplasma pneumoniae pneumonia (GMPP) group. Measurement data between the 2 groups were compared using Mann-Whitney U test. Logistic regression analyses were used to examine the associations between clinical data and RMPP. Receiver operating characteristic (ROC) curves were used to analyse the power of the markers for predicting RMPP. Results:A total of 1 428 patients finished the study, with 801 boys and 627 girls, aged 4.3 (2.7, 6.3) years. Mycoplasma pneumoniae DNA was positive in 534 cases (37.4%), of whom 446 cases (83.5%) were diagnosed with Mycoplasma pneumoniae pneumonia, including 251 boys and 195 girls, aged 5.2 (3.3, 6.9) years. Macrolides-resistant variation was positive in 410 cases (91.9%). Fifty-five cases were with RMPP, 391 cases with GMPP. The peak body temperature before the first visit and LDH levels in RMPP patients were higher than that in GMPP patients (39.6 (39.1, 40.0) vs. 39.2 (38.9, 39.7) ℃, 333 (279, 392) vs. 311 (259, 359) U/L, both P<0.05). Logistic regression showed the prediction probability π=exp (-29.7+0.667×Peak body temperature (℃)+0.004×LDH (U/L))/(1+exp (-29.7+0.667×Peak body temperature (℃)+0.004 × LDH (U/L))), the cut-off value to predict RMPP was 0.12, with a consensus of probability forecast of 0.89, sensitivity of 0.89, and specificity of 0.67; and the area under ROC curve was 0.682 (95% CI 0.593-0.771, P<0.01). Conclusion:In MPP patients with fever over 48 to <120 hours, a prediction probability π of RMPP can be calculated based on the peak body temperature and LDH level before the first visit, which can facilitate early identification of RMPP.

Objective:To investigate the etiology of necrotizing pneumonia (NP) in children and the clinical characteristics of NP caused by different pathogens in China.Methods:A retrospective, case-control study was performed in children with NP who were admitted to 13 hospitals in China from January 2008 to December 2019. The demographic and clinical information, laboratory data, etiological and radiological findings were analyzed. The data were divided into three groups based on the following years: 2008-2011, 2012-2015 and 2016-2019, and the distribution characteristics of the pathogens in different period were compared. Meanwhile, the pathogens of pediatric NP in the southern and northern China were compared. And the clinical characteristics of the Mycoplasma pneumoniae (MP) NP and the bacterial NP were also compared. T-test or Mann-Whitney nonparametric test was used for comparison of numerical variables, and χ 2 test was used for categorical variables. Results:A total of 494 children with NP were enrolled, the median ages were 4.7 (0.1-15.3) years, including 272 boys and 222 girls. Among these patients, pathogens were identified in 347 cases and the pathogen was unclear in the remaining 147 cases. The main pathogens were MP (238 cases), Streptococcus pneumoniae (SP) (61 cases), Staphylococcus aureus (SA) (51 cases), Pseudomonas aeruginosa (13 cases), Haemophilus influenzae (10 cases), adenovirus (10 cases), and influenza virus A (7 cases), respectively. MP was the most common pathogen in all three periods and the proportion increased yearly. The proportion of MP in 2016-2019 was significantly higher than that in 2012-2015 (52.1% (197/378) vs. 36.8% (32/87), χ 2= 6.654, P=0.010), while there was no significant difference in the proportion of MP in 2012-2015 and that in 2008-2011 (36.8% (32/87) vs. 31.0% (9/29), χ2=0.314, P=0.575).Regarding the regional distribution, 342 cases were in the southern China and 152 in the northern China. Also, MP was the most common pathogen in both regions, but the proportion of MP was higher and the proportion of SP was lower in the north than those in the south (60.5% (92/152) vs. 42.7% (146/342), χ 2=13.409, P<0.010; 7.9% (12/152) vs. 14.3% (49/342), χ 2= 4.023, P=0.045). Comparing the clinical characteristics of different pathogens, we found that fever and cough were the common symptoms in both single MP and single bacterial groups, but chest pain was more common (17.0% (34/200) vs. 6.1% (6/98), χ 2=6.697, P=0.010) while shortness of breath and wheezing were less common in MP group (16.0% (32/200) vs. 60.2% (59/98), χ 2=60.688, P<0.01; 4.5% (9/200) vs. 21.4% (21/98), χ 2=20.819, P<0.01, respectively). The white blood cell count, C-reactive protein and procalcitonin in the bacterial group were significantly higher than those in the MP group (14.7 (1.0-67.1)×10 9/L vs. 10.5 (2.5-32.2)×10 9/L, 122.5 (0.5-277.3) mg/L vs. 51.4 (0.5-200.0) g/L, 2.13 (0.05-100.00) μg/L vs. 0.24 (0.01-18.85) μg/L, Z=-3.719, -5.901 and -7.765, all P<0.01). Conclusions:The prevalence of pediatric NP in China shows an increasing trend during the past years. MP, SP and SA are the main pathogens of NP, and the most common clinical symptoms are fever and cough. The WBC count, C-reactive protein and procalcitonin in bacterial NP are significantly higher than those caused by MP.

ObjectiveTo explore the process and guidelines for ethical review in decentralized drug clinical trials， promote clinical trial progress， and ensure drug development progress. MethodsThe key points of the ethical review were summarized by studying the relevant laws and regulations on decentralized drug clinical trials， analyzing the advantages and challenges of decentralized drug clinical trials， and combining the experience of the ethics committee of the institution in reviewing decentralized drug clinical trials. ResultsRelevant laws and regulations were the basis for the ethical review， and the ethics committee should adopt appropriate review methods based on regulations and hospital ethical standard operating procedures. The ethics committee should focus on the feasibility， applicability， and rationality， the adequacy of informed consent， the protection of rights and interests and privacy of subjects， as well as the qualification and standard operating procedures of electronic platforms for conducting decentralized drug clinical trials. ConclusionDecentralized drug clinical trials are in their early stages and urgently require guidance from relevant laws and regulations. Ethical review is also constantly being refined through exploration. It is necessary to supervise the implementation of responsibilities by all parties， pay attention to the rights and interests of subjects， and gradually promote the implementation of decentralized drug clinical trials.

ObjectiveTo explore the mechanism of Qigesan （QGS） in intervening in the migration and invasion of esophageal carcinoma TE-1 cells. MethodMicroarray technology was used to screen differentially expressed genes （DEGs） in the normal group and the QGS group， and the ontological functions and signaling pathways of DEGs were analyzed. The thiazolyl tetrazolium （MTT） assay was used to detect the effect of QGS on the viability of TE-1 cells. In the subsequent experiments for verification， a blank group， a transforming growth factor-β₁ （TGF-β₁） group， a TGF-β₁ + QGS group， and a TGF-β₁ + SB431542 group were set up. The cell morphology in each experimental group was observed by microscopy. The migration and invasion abilities of cells were detected by wound healing assay， and the mRNA expression levels of E-Cadherin， vimentin， Smad2， and Smad7 were detected by Real-time quantitative polymerase chain reaction （Real-time PCR）. The protein expression of E-Cadherin， vimentin， p-Smad2/3， Smad2/3， and Smad7 was detected by Western blot. ResultThere were 1 487 DEGs between the QGS group and the blank group， including 1 080 down-regulated ones （accounting for 72.63%） and 407 up-regulated ones. The down-regulated genes were mainly involved in biological processes such as cytoskeletal protein binding， ATP binding， adenylate nucleotide binding， and adenylate ribonucleotide binding， and the involved Kyoto Encyclopedia of Genes and Genomes （KEGG） pathways included TGF-β signaling pathway， cell cycle， extracellular matrix-receptor interaction protein， tumor pathways， and oocyte meiosis. The up-regulated genes were mainly involved in RNA binding， DNA binding， transcriptional regulator activity， transcriptional activator activity， and nucleotide binding， and the KEGG pathways involved mainly included mitogen-activated protein kinase （MAPK） signaling pathway， bladder cancer， renal cell carcinoma， cancer pathways， and p53 signaling pathway. Compared with the blank group， the inhibition rate of cell viability of TE-1 cells increased after QGS （20， 30， 40， 60， 80 mg·L^-1） intervention for 12， 24， 36， 48， 60 h （P<0.05）， and the inhibition rate was time- and dose-dependent. Compared with the blank group， the TGF-β₁ group showed lengthened cells with fibroblast phenotype. Compared with the TGF-β₁ group， the TGF-β₁ + QGS group showed shortened cells with normal morphology and epithelial phenotype. The cell morphology in the TGF-β₁ + SB431542 group was similar to that of the TGF-β₁ + QGS group. Compared with the blank group， the TGF-β₁ group showed potentiated ability of cell migration and invasion （P<0.05）. Compared with the TGF-β₁ group， the TGF-β₁ + QGS group and the TGF-β₁ + SB431542 group showed inhibited and weakened migration and invasion abilities of cells （P<0.05）. However， there was no significant difference in migration and invasion abilities between the TGF-β₁ + QGS group and the TGF-β₁ + SB431542 group. The mRNA expression levels of vimentin and Smad2 in the TGF-β₁ group were higher （P<0.05）， and the mRNA expression levels of E-Cadherin and Smad7 were lower （P<0.05） than those in the blank group. Compared with the TGF-β₁ group， the TGF-β₁ + QGS group and the TGF-β₁+ SB431542 group exhibited decreased expression levels of vimentin and Smad2 mRNA （P<0.05）， and elevated expression levels of E-Cadherin and Smad7 mRNA （P<0.05）. Compared with the blank group， the TGF-β₁ group showed up-regulated protein expression levels of vimentin， p-Smad2/3， and Smad2/3 （P<0.05）， and reduced protein expression levels of E-Cadherin and Smad7 （P<0.05）. Compared with the TGF-β₁ group， the TGF-β₁ + QGS group and the TGF-β₁ + SB431542 group displayed decreased protein expression levels of vimentin， p-Smad2/3， and Smad2/3 （P<0.05）， and increased protein expression levels of E-Cadherin and Smad7 （P<0.05）. ConclusionThe ethyl acetate extract of QGS inhibits the epithelial-mesenchymal transition （EMT） of TE-1 cells through the TGF-β₁ pathway to reduce the migration and invasion of TE-1 cells.

ObjectiveTo investigate the mechanism of icariin in ameliorating efferocytosis dysfunction and inflammatory response of alveolar macrophages induced by cigarette smoke extract via the peroxisome proliferator-activated receptor gamma （PPARγ） signaling pathway. MethodThe untreated rat alveolar macrophages （NR8383） were taken as the blank group. The NR8383 cells treated with 10% cigarette smoke extract were divided into model， low-， medium-， and high-dose （10， 20， 40 μmol·L^-1） icariin， PPARγ inhibitor， and PPARγ inhibitor + low-， medium-， and high-dose icariin groups. Alamar blue colorimetry was employed to examine the proliferation and toxicity of icariin on NR8383 cells. The efferocytosis rate of NR8383 cells was detected by flow cytometry. Enzyme-linked immunosorbent assay was employed to measure the levels of tumor necrosis factor-alpha （TNF-α）， transforming growth factor-β₁ （TGF-β₁）， and milk fat globule-epidermal growth factor 8 （MFG-E8）. Western blot and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） were employed to determine the protein and mRNA levels， respectively， of PPARγ， CD36， and RAS-related C3 botulinum toxin substrate 1 （Rac1）. ResultThe efferocytosis dysfunction model of NR8383 was established with the cigarette smoke extract. Compared with the blank control group， the model group showed decreased efferocytosis rate （P<0.05）， elevated TNF-α level （P<0.05）， lowered TGF-β₁ and MFG-E8 levels （P<0.01）， and down-regulated mRNA and protein levels of PPARγ， CD36， and Rac1 （P<0.05， P<0.01）. Compared with the model group， the treatment with icariin increased the efferocytosis rate （P<0.05， P<0.01）， lowered the TNF-α level （P<0.01）， elevated TGF-β₁ and MFG-E8 levels （P<0.05）， and up-regulated the protein and mRNA levels of PPARγ， CD36， and Rac1 （P<0.05， P<0.01）. Compared with icariin alone， PPARγ inhibitor + icariin decreased the efferocytosis rate （P<0.05） and down-regulated the protein and mRNA levels of PPARγ （P<0.05， P<0.01）. In addition， PPARγ inhibitor + low-dose icariin down-regulated the protein level of CD36 （P<0.01） and PPARγ inhibitor + low-/medium-dose icariin up-regulated the protein level of Rac1 （P<0.05）. ConclusionIcariin ameliorates the cigarette smoke extract-induced efferocytosis dysfunction of alveolar macrophage by regulating the PPARγ signaling pathway and cytoskeletal structure rearrangement.

The coronavirus disease 2019 (COVID-19) has caused a global pandemic. All people including children are generally susceptible to COVID-19, but the condition is relatively mild for children. The diagnosis of COVID-19 is largely based on the epidemiological evidence and clinical manifestations, and confirmed by positive detection of virus nucleic acid in respiratory samples. The main symptoms of COVID-19 in children are fever and cough; the total number of white blood cell count is usually normal or decreased; the chest imaging is characterized by interstitial pneumonia, which is similar to other respiratory virus infections and infections. Early identification, early isolation, early diagnosis and early treatment are important for clinical management. The treatment of mild or moderate type of child COVID-19 is mainly symptomatic. For severe and critical ill cases, the oxygen therapy, antiviral drugs, antibacterial drugs, glucocorticoids, mechanical ventilation or even extracorporeal membrane oxygenation (ECMO) may be adopted, and the treatment plan should be adjusted timely through multi-disciplinary cooperation.
Betacoronavirus ; isolation & purification ; Child ; Coronavirus Infections ; diagnosis ; pathology ; therapy ; Humans ; Pandemics ; Pneumonia, Viral ; diagnosis ; diagnostic imaging ; etiology ; pathology ; therapy

Betacoronavirus ; isolation & purification ; Child ; Clinical Laboratory Techniques ; Coronavirus Infections ; diagnosis ; drug therapy ; therapy ; Humans ; Pneumonia, Viral ; diagnosis ; therapy