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Background Methicillin-resistant Staphylococcus aureus (MRSA) can cause invasive infections with significant mortality in neonates. This study aimed to analyze the clinical characteristics and antibiotic resistance profiles of invasive MRSA infections and determine risk factors associated with invasive MRSA infections in newborn inpatients. Methods This multicenter retrospective study of inpatients from eleven hospitals in the Infectious Diseases Surveillance of Pediatrics (ISPED) group of China was performed over a two-year period (2018–2019). Statistical significance was calculated by applying the χ2 test or by Fisher’s exact test in the case of small sample sizes. Results A total 220 patients were included. Among included cases, 67 (30.45%) were invasive MRSA infections, including two deaths (2.99%), while 153 (69.55%) were noninvasive infections. The invasive infections of MRSA occurred at a median age of 8 days on admission, which was significantly younger compared to 19 days in noninvasive cases. Sepsis (86.6%) was the most common invasive infection, followed by pneumonia (7.4%), bone and joint infections (3.0%), central nervous system infection (1.5%), and peritonitis (1.5%). Congenital heart disease, low birth weight infant ( Conclusion Low age at admission (≤8 days), congenital heart disease, and low birth weight were associated with invasive MRSA infections in neonates, and no isolates resistant to vancomycin and linezolid were found. Determining these risks in suspected neonates may help identify patients with imminent invasive infections who may require intensive monitoring and therapy.

ObjectivesTo develop a rapid and low-cost method for 16S rDNA nanopore sequencing.MethodsThis was a prospective study on a 16S rDNA nanopore sequencing method. We developed this nanopore barcoding 16S sequencing method by adding barcodes to the 16S primer to reduce the reagent cost and simplify the experimental procedure. Twenty-one common pulmonary bacteria (7 reference strains, 14 clinical isolates) and 94 samples of bronchoalveolar lavage fluid from children with severe pneumonia were tested. Results indicating low-abundance pathogenic bacteria were verified with the polymerase chain reaction (PCR). Further, the results were compared with those of culture or PCR.ResultsThe turnaround time was shortened to 6~8 hours and the reagent cost of DNA preparation was reduced by employing a single reaction adding barcodes to the 16S primer in advance. The accuracy rate for the 21 common pulmonary pathogens with an abundance ≥ 99% was 100%. Applying the culture or PCR results as the gold standard, 71 (75.5%) of the 94 patients were positive, including 25 positive cultures (26.6%) and 52 positive quantitative PCRs (55.3%). The median abundance in the positive culture and qPCR samples were 29.9% and 6.7%, respectively. With an abundance threshold increase of 1%, 5%, 10%, 15% and 20%, the test sensitivity decreased gradually to 98.6%, 84.9%, 72.6%, 67.1% and 64.4%, respectively, and the test specificity increased gradually to 33.3%, 71.4%, 81.0%, 90.5% and 100.0%, respectively.ConclusionsThe nanopore barcoding 16S sequencing method can rapidly identify the pathogens causing bacterial pneumonia in children.

AIMS: To explore the contagiousness and new SARS-CoV-2 mutations in pediatric COVID-19. METHODS: This cohort study enrolled all pediatric patients admitted to 8 hospitals in Zhejiang Province of China between 21 January and 29 February 2020, their family members and close-contact classmates. Epidemiological, demographic, clinical and laboratory data were collected. Bioinformatics was used to analyze the features of SARS-CoV-2. Individuals were divided into 3 groups by the first-generation case: Groups 1 (unclear), 2 (adult), and 3 (child). The secondary attack rate (SAR) and R0 were compared among the groups. RESULTS: The infection rate among 211 individuals was 64% (135/211). The SAR in Groups 2 and 3 was 71% (73/103) and 3% (1/30), respectively; the median R0 in Groups 2 and 3 was 2 (range: 1-8) and 0 (range: 0-1), respectively. Compared with adult cases, the SAR and R0 of pediatric cases were significantly lower (p

Tumors escape immune attack by upregulating the surface expression of PD-L1, which interacts with PD-1 on T cells to activate immune inhibitory signaling. Anti-PD-1 treatments can effectively block this inhibitory signaling and activate antitumor immune responses. However, anti-PD-1 treatment also has a tumor suppressive effect in patients whose tumor cells do not express PD-L1. The underlying mechanisms are poorly defined. Here, we report that exosomes from bone marrow-derived cells (BMDCs) in tumor-bearing mice, but not in healthy mice, carry PD-L1. PD-L1 on these exosomes is biofunctional and can inhibit CD8(+) T cell proliferation and activation in vitro and in vivo. The transfer of exogenous exosomes from BMDCs and the inhibition of the production of endogenous exosomes by BMDCs promote and suppress tumor growth, respectively. PD-L1(+) exosomes from BMDCs can be found in tumor tissues. In addition, exosomes from BMDCs promote tumor metastasis in a PD-L1-dependent manner. Therefore, our results indicate that exosomes from BMDCs play important roles in tumor immunosuppression via PD-L1.

Chronic active Epstein-Barr virus (CAEBV) infection is a very rare and potentially life-threatening illness caused by long-term EBV infection. Globally, the prevalence is the highest among young children and adolescents with increased frequency in Asians and Native Americans, whereas it is sporadically encountered in European countries. Typically, patients present with nonspecific symptoms such as fever, lymphadenopathy, hepatosplenomegaly, and liver dysfunction. However, the complications of CAEBV and its treatment are quite complex and require great care. We report a case of a 3-year-old boy with CAEBV infection who was later diagnosed with pancreatic mass and recurrent pancreatitis. A multidisciplinary board was consulted for correct diagnosis and treatment plan making. The treatment included pharmaceutical and surgical (duodenum-preserving pancreatic head resection) approaches. The patient showed tremendous improvement following the third discharge from the hospital and is still free of any symptoms. In this case report, we discuss differential diagnosis and comprehensively examined the possibility of the development of pancreatitis caused by EBV infection, the possibility of autoimmune pancreatitis, and the possibility of hereditary pancreatitis. To confirm or rule out the first two etiologies, laboratory and pathology results were studied. We also performed exon sequencing using Agilent exome capture kit to rule out hereditary pancreatitis. The clinical course of this disease and the way it was handled deserves attention so that similar cases receive prompt treatment.

As antibiotics are always toxic to the antibiotic-producing strains themselves, most Streptomyces strains have evolved several self-resistance mechanisms, among which the antibiotic efflux system is understood best and is commonly found. Among the efflux systems, the ATP-binding cassette (ABC) transporter superfamily and the major facilitator superfamily (MFS) are two important transporter families. In this work, the ABC transporters and the MFS transporters from the four reported natamycin-producing Streptomyces strains have been investigated in order to clarify whether these Streptomyces strains share similar efflux strategies for natamycin metabolism. Fifty-one groups of homologous exporter genes were identified as shared by four strains. Differential transcriptional analysis between the natamycin-producing strain Streptomyces chattanoogensis L10 and its ΔscnS0 mutant, which produces no natamycin, reveals that the expression levels of 25 of the above groups of genes were observably changed. The production of natamycin declined over 30% after solely knocking out several of these 25 groups of genes in S. chattanoogensis L10. This indicates that these transporters participate in the efflux of molecules related to natamycin biosynthesis. Our study is the first to demonstrate that the exporters participating in a particular antibiotic metabolism can be excavated and identified quickly by the strategy of genome mining and homologous comparison in the antibiotic-producing strains, leading to deeper understanding of the complex self-resistance mechanisms in Streptomycetes.

Fas induces apoptosis in activated T cell to maintain immune homeostasis, but the effects of non-apoptotic Fas signaling on T cells remain unclear. Here we show that Fas promotes TH9 cell differentiation by activating NF-κB via Ca2+-dependent PKC-β activation. In addition, PKC-β also phosphorylates p38 to inactivate NFAT1 and reduce NFAT1-NF-κB synergy to promote the Fas-induced TH9 transcription program. Fas ligation exacerbates inflammatory bowel disease by increasing TH9 cell differentiation, and promotes antitumor activity in p38 inhibitor-treated TH9 cells. Furthermore, low-dose p38 inhibitor suppresses tumor growth without inducing systemic adverse effects. In patients with tumor, relatively high TH9 cell numbers are associated with good prognosis. Our study thus implicates Fas in CD4+ T cells as a target for inflammatory bowel disease therapy. Furthermore, simultaneous Fas ligation and low-dose p38 inhibition may be an effective approach for TH9 cell induction and cancer therapy.

Background: HAdV is one of the common pathogens in hospitalized children with respiratory infection (RI). we aim to describe the epidemiological characteristics, clinical features, and genotype of inpatients with HAdV positive with RI.Methods: Respiratory samples were gathered from inpatients diagnosed RI in Children’s Hospital, Zhejiang University School of Medicine and were detected by using Direct Immunofluorescence Assay (DFA) from 2018 to 2019. PCR amplification and sequencing of the hypervariable zone of hexon gene were used for genotyping. The clinical and laboratory features, and typing of HAdV, and epidemiological characteristic analysis were retrospectively performed. Results: Of 7072 samples collected, 488 cases were identified as HAdV-positive RI. The overall detection rate was 6.9%. The peaked detection rate was 14.1% in January 2019. HAdV-positive cases with RI mainly appeared in winter and summer. The detection rate was highest among children between 6 months and 2 years (8.7%, 123/1408). Clinical diagnosis included pneumonia (70.3%, 343/488), bronchitis (7.0%, 34/488) and acute upper respiratory tract infection (22.7%, 111/488). The common clinical manifestations were fever (93.4%, 456/488), cough (94.7%, 462/488), wheezing (26.2%,128/48), and shortness of breath (14.8%, 72/488). 213 (43.6%) cases had co-infection and 138 (28.3%) cases had extrapulmonary symptoms. 96(19.7%) cases had intrapulmonary and intrathoracic complications.78 (16.0%) had an underlying condition, the most of them were congenital heart diseases (20.5%,17/78). The proportions of hyperpyrexia, duration of fever more than 10 days, severe pneumonia, and proportion of wheezing in the co-infection group were remarkably higher than those in HAdV single-infection group (pp=0.000), while, HAdV-B55 was detected only in the severe group. HAdV-B7’s detection rate in the severe group was significantly higher than the non-severe infected group. Conclusion: HAdV detection rate is related to age and season. Bronchopneumonia accounts for about 70% HAdV -positive inpatients. The common manifestations include hyperpyrexia, cough, wheezing, and shortness of breath. HAdV-B3 and HAdV-B7 are the most common types in children diagnosed with RI.

Background HAdV is one of the common pathogens in hospitalized children with acute respiratory infections (ARIs). We aim to describe the clinical and laboratory features, epidemiological characteristics, and HAdV species and/or types of inpatients with HAdV respiratory infections. Methods Respiratory samples were gathered from inpatients diagnosed ARIs in Children’s Hospital, Zhejiang University School of Medicine, and were detected by using Direct Immunofluorescence Assay from 2018 to 2019. PCR amplification and sequencing of the hypervariable zone of hexon gene were used for genotyping. The clinical and laboratory features, and HAdV genotyping, and epidemiological characteristic analysis were retrospectively performed. Results Of 7072 samples collected, 488 were identified as HAdV-positive. The overall detection rate was 6.9%. The peaked detection rate was 14.1% in January 2019. HAdV-positive cases with ARIs mainly appeared in winter. The detection rate was highest among children between 6 months and 2 years (8.7%, 123/1408). Clinical diagnosis included pneumonia (70.3%, 343/488), bronchitis (7.0%, 34/488) and acute upper respiratory tract infection (22.7%, 111/488). The common clinical manifestations were fever (93.4%, 456/488), cough (94.7%, 462/488), wheezing (26.2%, 128/488), and shortness of breath (14.8%, 72/488). 213 (43.6%) cases had co-infection and 138 (28.3%) cases had extrapulmonary symptoms. 96(19.7%) cases had intrapulmonary and intrathoracic complications.78 (16.0%) had an underlying condition, most of which were congenital heart diseases (20.5%, 16/78). The proportions of hyperpyrexia, duration of fever > 10 days, severe pneumonia, and wheezing in the co-infection group were remarkably higher than those in HAdV single-infection group (all p 10 days, wheezing, shortness of breath, change in level of consciousness, serosal fluids, extrapulmonary symptoms, co-infections and underlying diseases were significantly higher in severe pneumonia group than those in the mild pneumonia group (all p p = 0.000), while, HAdV-B55 was detected only in the severe group. HAdV-B7’s detection rate in the severe pneumonia group was significantly higher than the non-severe pneumonia group. Conclusion HAdV detection rate is related to age and season. Bronchopneumonia accounts for about 70% HAdV-positive inpatients. The common clinical manifestations include hyperpyrexia, cough, wheezing, and shortness of breath. HAdV-B3 and HAdV-B7 are the most common types in children diagnosed with respiration infections.

Streptomyces is well known for biosynthesis of secondary metabolites with diverse bioactivities. Although oils have been employed as carbon sources to produce polyketide antibiotics for several industrial Streptomyces strains, the intrinsic correlation between oil utilization and high production of antibiotics still remains unclear. In this study, we investigated the correlation between oil metabolism and salinomycin biosynthesis in Streptomyces albus ZD11, which employs soybean oil as the main carbon source. Comparative genomic analysis revealed the enrichment of genes related to triacylglycerol (TAG) metabolism in S. albus ZD11. Transcriptomic profiling further confirmed the enhancement of TAG metabolism and acyl coenzyme A biosynthesis in S. albus ZD11. Multiple secreted lipases, which catalyze TAG hydrolysis, were seen to be working in a synergistic and complementary manner in aiding the efficient and stable hydrolyzation of TAGs. Together, our results suggest that enhanced TAG hydrolysis and fatty acid degradation contribute to the high efficiency of oil utilization in S. albus ZD11 in order to provide abundant carbon precursors for cell growth and salinomycin biosynthesis. IMPORTANCE In order to obtain high-level production of antibiotics, oils have been used as the main carbon source for some Streptomyces strains. Based on multiomics analysis, this study provides insight into the relationship between triacylglycerol (TAG) metabolism and antibiotic biosynthesis in S. albus ZD11, an oil-preferring industrial Streptomyces strain. Our investigation into TAG hydrolysis yielded further evidence that this strain utilizes complicated strategies enabling an efficient TAG metabolism. In addition, a novel secreted lipase was identified that exhibited highly hydrolytic activity for medium- and long-chain TAGs. Our findings represent a good start toward clarifying the complicated relationship between TAG catabolism and high-level antibiotic production in the industrial strains.

Exosomes derived from heat‐stressed tumour cells (HS‐TEXs), which contain abundant heat shock protein (HSP) 70, strongly induce antitumour immune responses. HSP70‐induced interleukin (IL)‐6 promotes IL‐17 expression and causes rejection of established prostate tumours. However, it remains unclear whether HS‐TEXs exhibit antitumour effects by converting regulatory T cells (T(regs)) into T helper type 17 (Th17) cells. In this study, we found that compared with TEXs, HS‐TEXs were more potent in stimulating secretion of IL‐6 from dendritic cells. In vitro, IL‐6 blocked tumour cell‐derived transforming growth factor beta 1‐induced T(reg) differentiation and promoted Th17 cell differentiation. HS‐TEXs exerted strong antitumour effects, converting T(regs) into Th17 cells with high efficiency, a process that was entirely dependent upon IL‐6. Neutralization of IL‐17 completely abolished the antitumour effect of TEXs, but only partially inhibited that of HS‐TEXs. In addition, we found higher levels of IL‐6 and IL‐17 in serum from tumour patients treated with hyperthermia, and an increase in Th17 cells and a decrease in T(regs) was detected in peripheral blood mononuclear cells isolated from these patients after hyperthermia. Therefore, our results demonstrate that HS‐TEXs possess a powerful capacity to convert immunosuppressive T(regs) into Th17 cells via IL‐6, which contributes to their potent antitumour effect.

The pandemic influenza A (H1N1)pdm09 virus continues to be a threat to human health. Low doses of mannan-binding lectin (MBL) (

Herpangina is a common infectious disease in childhood caused by an enterovirus. This consensus is aiming to standardize and improve herpangina prevention and clinical diagnosis. The Subspecialty Group of Infectious Diseases, the Society of Pediatric, Chinese Medical Association and Nation Medical Quality Control Center for Infectious Diseases gathered 20 experts to develop the consensus, who are specialized in diagnosis and treatment of herpangina. The main pathogenic serotypes of herpangina include Coxsackievirus-A, Enterovirus-A and Echovirus. Its diagnosis can be rendered on the basis of history of epidemiology, typical symptoms, characteristic pharyngeal damage and virological tests. The treatment is mainly symptomatic, and incorporates topical oral spray with antiviral drugs. The course of herpangina generally lasts 4–6 days with a good prognosis. The consensus could provide advices and references for the diagnosis, treatment and management of herpangina in children.

Predominant T helper (Th)2 and impaired Th1 cell polarization has a crucial role in the development of asthma. Cluster of differentiation (CD)38 is associated with the increased release of interleukin (IL)‑12 from dendritic cells (DCs) and DC‑induced Th1 cell polarization. However, whether CD38 expression affects DC function in asthma development remains unknown. In the current study, adenoviruses were constructed containing the murine CD38 gene. Overexpression of CD38 protein level in DCs induced from bone‑marrow derived DCs (BMDCs) by recombinant mouse granulocyte macrophage colony‑stimulating factor and IL‑4 was achieved through 24 h adenovirus infection. The results demonstrated that BMDCs with CD38 overexpression exhibited no phenotypic change; however, following stimulation with lipopolysaccharide (LPS), maturation and IL‑12 secretion were increased. In addition, CD38‑overexpressing BMDCs stimulated with LPS exhibited more effective Th1 cell differentiation. Mice that were administered CD38‑overexpressing BMDCs exhibited milder symptoms of asthma. Furthermore, decreased IL‑4, IL‑5 and IL‑13 levels were detected in bronchoalveolar lavage fluid (BALF), reduced immunoglobulin E levels were measured in the sera, and increased interferon‑γ was detected in BALF from the recipients of CD38‑overexpressing BMDCs. Increased phosphorylated‑p38 expression was also detected in LPS-stimulated CD38-overexpressing BMDCs, whereas pretreatment with a p38‑specific inhibitor was able to abolish the effects of LPS stimulation and CD38 overexpression on IL‑12 release and Th1 cell differentiation in BMDCs. These results suggested that CD38 may be involved in the DC function of alleviating asthma via restoration of the Th1/Th2 balance, thus providing a novel strategy for asthma therapy.

Background: Prognosis of pneumococcal meningitis (PM) remains very poor, especially in less developed countries. Currently, few multi-centric studies on pediatric PM have been reported in mainland China. Objectives: This study aimed to explore the correlation of clinical and laboratory findings with complications and prognosis in pediatric PM. Methods: The pediatric PM patients were retrospectively recruited from ten pediatric tertiary hospitals across China between January 2013 and June 2018. Clinical, biochemical, and microbiological data and follow-up information were collected. Predictive factors for complications and prognostic factors for overall survival (OS) and sequelae-free survival (SFS) were analyzed. Results: A total of 132 pediatric PM patients were included. Seventy-one patients had complications, 25 patients died, and 39 patients had neurological sequelae. Multivariate logistic regression suggested that age less than 28 months (adjusted OR=2.654, 95%CI=1.067–6.600, P=0.036) and lower white blood cells in blood (aOR=3.169, 95%CI=1.395–7.202, P=0.006) were associated with high risk of complications. Multivariate Cox’s proportional hazard regression suggested that age less than 28 months (aHR=6.479, 95%CI=1.153–36.404, P=0.034), coma (aHR=9.808, 95%CI=2.802–34.323, P=0.000), and non-adjuvant steroid therapy (aHR=4.768 95%CI=1.946–11.678, P=0.001) were independent prognostic factors for poor OS; coma (aHR=5.841, 95%CI=2.652–12.864, P=0.000), septic shock on admission (aHR=2.949, 95%CI=1.049–8.290, P=0.040), and lower glucose level in cerebrospinal fluid (CSF) (aHR=2.523, 95%CI=1.336–4.765, P=0.004) were independent prognostic factors for poor SFS. Conclusion: Age, coma, and adjuvant steroid therapy were independent factors for OS, while coma, septic shock on admission, and lower glucose level in CSF were independent factors for SFS in pediatric PM patients. These factors might be used to identify PM patients with poor prognosis and guide individual treatment.

Fas induces apoptosis in activated T cell to maintain immune homeostasis, but the effects of non-apoptotic Fas signaling on T cells remain unclear. Here we show that Fas promotes TH9 cell differentiation by activating NF-κB via Ca2+-dependent PKC-β activation. In addition, PKC-β also phosphorylates p38 to inactivate NFAT1 and reduce NFAT1-NF-κB synergy to promote the Fas-induced TH9 transcription program. Fas ligation exacerbates inflammatory bowel disease by increasing TH9 cell differentiation, and promotes antitumor activity in p38 inhibitor-treated TH9 cells. Furthermore, low-dose p38 inhibitor suppresses tumor growth without inducing systemic adverse effects. In patients with tumor, relatively high TH9 cell numbers are associated with good prognosis. Our study thus implicates Fas in CD4+ T cells as a target for inflammatory bowel disease therapy. Furthermore, simultaneous Fas ligation and low-dose p38 inhibition may be an effective approach for TH9 cell induction and cancer therapy., Fas signalling induces apoptosis of activated T cells to maintain immune homeostasis. Here the authors show that Fas also induces PKC-β activation to promote NF-κB-mediated TH9 cell differentiation, while p38 activation by PKC-β antagonizes this effect, thereby supporting a synergy between p38 inhibitor and Fas for TH9 differentiation.

Background: Human cytomegalovirus (HCMV) infection is very common and latency can be reactivated in the future. And it can alter the intracellular environment, similar to other herpesviruses, for viral replication and survival. The aim of this study was to investigate the influence of HCMV infection on autophagy in human acute monocytic leukemia cell line (THP-1 cells). Methods: Reverse transcription polymerase chain reaction (RT-PCR), western blot, and transmission electron microscope (TEM) were used to examine autophagy level. The concentrations of autophagy-related proteins Beclin 1, Atg5, and the light chain three (LC3) were counted when compared with actin level. Results: The expression levels of Beclin1, Atg5, and LC3II mRNAs increased gradually between 1 and 5 days(d) postinfection (p.i.) and subsequently decreased little by little when compared with the control THP-1 cells. However, results of western blot analysis displayed that the level of LC3II increased gradually after 1 day p.i. and decreased at 7 days after infection. But the levels of Atg5 and Beclin1 increased gradually after 2 days p.i. and began to decrease at 5 days after infection, respectively. Conclusion: These results suggested that HCMV infection can facilitate the autophagy and autophagy level may decrease in latent phase. More studies on the relationship between HCMV latency and autophagy are needed to determine the role of autophagy in HCMV latent infection that may help find out a therapeutic approach for clinical treatment.

Summary Systemic immunosuppression greatly affects the chemotherapeutic antitumor effect. Here, we showed that CD19+ extracellular vesicles (EVs) from B cells through CD39 and CD73 vesicle-incorporated proteins hydrolyzed ATP from chemotherapy-treated tumor cells into adenosine, thus impairing CD8+ T cell responses. Serum CD19+ EVs were increased in tumor-bearing mice and patients. Patients with fewer serum CD19+ EVs had a better prognosis after chemotherapy. Upregulated hypoxia-inducible factor-1α (HIF-1α) promoted B cells to release CD19+ EVs by inducing Rab27a mRNA transcription. Rab27a or HIF-1α deficiency in B cells inhibited CD19+ EV production and improved the chemotherapeutic antitumor effect. Silencing of Rab27a in B cells by inactivated Epstein-Barr viruses carrying Rab27a siRNA greatly improved chemotherapeutic efficacy in humanized immunocompromised NOD Prkdcscid Il2rg−/− mice. Thus, decreasing CD19+ EVs holds high potential to improve the chemotherapeutic antitumor effect.

We have previously demonstrated that exosomes from dendritic cells (DCs) secreting TGF-β1 (sTGF-β1-EXOs) delay the development of murine inflammatory bowel disease (IBD). In this study, we isolated exosomes from DCs expressing membrane-associated TGF-β1 (mTGF-β1-EXOs) and found mTGF-β1-EXOs had more potent immunosuppressive activity than sTGF-β1-EXOs in vitro. Treatment of mice with mTGF-β1-EXOs inhibited the development and progression of myelin oligodendrocyte glycoprotein (MOG) peptide-induced EAE even after disease onset. Treatment of mice with mTGF-β1-EXOs also impaired Ag-specific Th1 and IL-17 responses, but promoted IL-10 responses ex vivo. Treatment with mTGF-β1-EXOs decreased the frequency of Th17 cells in EAE mice, which might be associated with the down-regulation of the p38, ERK, Stat3, and NF-κB activation and IL-6 expression in DCs. Treatment with mTGF-β1-EXOs maintained the regulatory capacity of Treg cells, and adoptive transfer of CD4(+)Foxp3(+)Treg cells from mTGF-β1-EXO-treated EAE mice dramatically prevented the development of EAE in the recipients. Moreover, treatment with mTGF-β1-EXOs from C57BL/6 mice effectively prevented and inhibited proteolipid protein (PLP) peptide-induced EAE in BALB/c mice. These results indicate that mTGF-β1-EXOs possess powerful immunosuppressive ability and can effectively inhibit the development and progression of EAE in different strains of mice.

Autophagy can mediate antiviral immunity. However, it remains unknown whether autophagy regulates the immune response of dendritic cells (DCs) to influenza A (H1N1) pdm09 infection. In this study, we found that infection with the H1N1 virus induced DC autophagy in an endocytosis-dependent manner. Compared with autophagy-deficient Beclin-1(+/-) mice, we found that bone-marrow-derived DCs from wild-type mice (WT BMDCs) presented a more mature phenotype on H1N1 infection. Wild-type BMDCs secreted higher levels of interleukin-6 (IL-6), tumour necrosis factor- α (TNF-α), interferon-β (IFN-β), IL-12p70 and IFN-γ than did Beclin-1(+/-) BMDCs. In contrast to Beclin-1(+/-) BMDCs, H1N1-infected WT BMDCs exhibited increased activation of extracellular signal-regulated kinase, Jun N-terminal kinase, p38, and nuclear factor-κB as well as IFN regulatory factor 7 nuclear translocation. Blockade of autophagosomal and lysosomal fusion by bafilomycin A1 decreased the co-localization of H1N1 viruses, autophagosomes and lysosomes as well as the secretion of IL-6, TNF-α and IFN-β in H1N1-infected BMDCs. In contrast to Beclin-1(+/-) BMDCs, H1N1-infected WT BMDCs were more efficient in inducing allogeneic CD4(+) T-cell proliferation and driving T helper type 1, 2 and 17 cell differentiation while inhibiting CD4(+) Foxp3(+) regulatory T-cell differentiation. Moreover, WT BMDCs were more efficient at cross-presenting the ovalbumin antigen to CD8(+) T cells. We consistently found that Beclin-1(+/-) BMDCs were inferior in their inhibition of H1N1 virus replication and their induction of H1N1-specific CD4(+) and CD8(+) T-cell responses, which produced lower levels of IL-6, TNF-α and IFN-β in vivo. Our data indicate that autophagy is important in the regulation of the DC immune response to H1N1 infection, thereby extending our understanding of host immune responses to the virus.

Wi-Fi based indoor positioning, which is based on attenuation of Received Signal Strength Indicator (RSSI) is an emerging Location Based Service (LBS) technology. As positioning accuracy is sensitive to environmental factors, most of the existing algorithms based on experimental test perform badly without adaptation to dynamics of environment. In this paper, we propose an indoor positioning method by locating the representation of a cluster within similar environments. The K-Means algorithm is used to extract the similarities of the objects within the nearby area. To overcome the problem of parameter determination under the circumstances of lack of fingerprint and extra hardware, we proposed a Log-normal shadowing model (LNSM) with Artificial Neural Networks to estimate distance enabling the parameters to be dynamically adjusted according to the change of the environment. The experimental results of one day auto fair data demonstrate the performance of our method with a higher degree of accuracy than other methods.

Tumour-cell-derived exosomes (Exo) have been proposed as a new kind of drug carrier, and heat stress can promote release of exosomes from tumour cells. This study investigated the impact of heat stress on the quantity of doxorubicin in exosomes from the same number of doxorubicin-treated MFC-7 tumour cells and their anti-tumour effects.Exosomes were isolated from phosphate-buffered saline (Exo), doxorubicin (Exo-Dox) or doxorubicin combined with heat-stress-treated (Exo-Dox-HS) MCF-7 cells. The content of doxorubicin in the exosomes was determined by flow cytometry. The effects of individual types of exosomes on the MCF-7 cell proliferation and apoptosis as well as the tumour growth were determined by MTT assay, flow cytometry and murine xenograft tumour modelling.We found that the amount of Exo-Dox-HS was higher than that of Exo-Dox from the same number of MCF-7 cells, and Exo-Dox-HS contained higher levels of doxorubicin than Exo-Dox from the same number of cells. Exo-Dox and Exo-Dox-HS, but not Exo or 10 µg/mL doxorubicin, significantly inhibited the MCF-7 cell proliferation and triggered MCF-7 cell apoptosis, associated with increased levels of cleaved caspase-3 and -8 and morphological changes in MCF-7 cells. Treatment with Exo-Dox and Exo-Dox-HS inhibited the growth of implanted breast tumours in mice.Our study indicated that heat stress increased the quantity of doxorubicin-containing exosomes from tumour cells, and enhanced the anti-tumour effect of exosomes from the doxorubicin-treated tumour cells. Our findings may aid in designing new strategies for cancer therapy by combination of chemotherapy and hyperthermia.

We have previously demonstrated that exosomes from dendritic cells (DCs) secreting TGF-β1 (sTGF-β1-EXOs) delay the development of murine inflammatory bowel disease (IBD). In this study, we isolated exosomes from DCs expressing membrane-associated TGF-β1 (mTGF-β1-EXOs) and found mTGF-β1-EXOs had more potent immunosuppressive activity than sTGF-β1-EXOs in vitro. Treatment of mice with mTGF-β1-EXOs inhibited the development and progression of myelin oligodendrocyte glycoprotein (MOG) peptide-induced EAE even after disease onset. Treatment of mice with mTGF-β1-EXOs also impaired Ag-specific Th1 and IL-17 responses, but promoted IL-10 responses ex vivo. Treatment with mTGF-β1-EXOs decreased the frequency of Th17 cells in EAE mice, which might be associated with the down-regulation of the p38, ERK, Stat3, and NF-κB activation and IL-6 expression in DCs. Treatment with mTGF-β1-EXOs maintained the regulatory capacity of Treg cells, and adoptive transfer of CD4(+)Foxp3(+)Treg cells from mTGF-β1-EXO-treated EAE mice dramatically prevented the development of EAE in the recipients. Moreover, treatment with mTGF-β1-EXOs from C57BL/6 mice effectively prevented and inhibited proteolipid protein (PLP) peptide-induced EAE in BALB/c mice. These results indicate that mTGF-β1-EXOs possess powerful immunosuppressive ability and can effectively inhibit the development and progression of EAE in different strains of mice.

Mannose-binding lectin (MBL) has broad range of activity against viruses through the mechanisms of neutralization, opsonization, and complement activation. Prior studies have demonstrated that MBL inactivated the season's influenza virus. Due to the fact that children have no neutralizing antibody against H1N1 2009 virus, innate immunity may be crucial in the defense against influenza. Therefore, we studied whether MBL levels played a role in H1N1 2009 infection in children. In a prospective survey, we revealed that MBL levels in ICU influenza cases were significantly lower than in children with influenza from infection disease ward. MBL may be involved in innate immune responses to H1N1 2009 infection in children.

Objective To highlight the clinical presentations of influenza A (H1N1) infection, for early diagnosis and recognition by the pediatricians. Methods In this retrospective study, the medical records of inpatients with influenza A (H1N1) infection between November 1, 2009 and May 31, 2011were reviewed. Results Eighty pediatric in-patients with median age 41.9 mo were studied. ARDS (11/80), pneumothorax (8/80), pleural effusion (7/80) and encephalopathy (7/80) were the most frequent complications. Six of 11 ARDS patients died;all of them were under 5 y. The median days of viral shedding was 11.4 d. Slight increase of Il-6, Il-10 and TNF-γ were revealed in some cases. Conclusions During late stage of pandemic wave, the majority of patients were young children. Children with severe Influenza A (H1N1) are prone to develop complications, and die from ARDS. If influenza-like illness is accompanied by neurologic signs, influenza A (H1N1) virus infection should be considered. The viral shedding in children is longer than in adults.

A limited number of drugs have been used for treatment of human cytomegalovirus (HCMV), all sharing the similar antiviral mechanism of inhibiting virus replication. This study investigates the anti-HCMV activities of mannose-binding lectin (MBL) from blocking virus entry and inhibiting virus spread. Recombinant human MBL was produced in CHO cells and native human MBL was isolated from human serum. A HCMV neutralization test was performed by pre-treating HCMV with each diluted MBL solution. Then the treated HCMV was inoculated onto the human embryonic pulmonary fibroblasts (HELF), which was followed by HCMV-DNA detection, PP65 positivity examination and confocal imaging of the infected cells. To test the activity of MBL in inhibiting viral spreading after viral invasion, HCMV growth inhibition test was performed. The infected cells were incubated with each diluted MBL, every 24 h, the supernatant was tested for HCMV-DNA. After 72 h, cells were collected for HCMV-DNA and PP65 examination. Then the cytopathic effect was observed and cell viability was measured at the 5 days after infection. HCMV neutralization test revealed 10 μg/mL MBL significantly decreased the HCMV invasion in HELF and the anti-HCMV activity can be blocked by 20 mg/mL mannan. HCMV growth inhibition test indicated that at 48 h after HCMV invasion, the HCMV-DNA level in the culture supernatant with 10 μg/mL MBL was lower than the control. After 72 h, both the HCMV-DNA levels and PP65 positivity in cells incubated with MBL were reduced. This is the first to report on the anti-HCMV activities of MBL by in vitro studies.

This retrospective study was conducted to estimate the shedding of 2009 H1N1 virus and the risk analysis by review of medical charts, laboratory and radiological findings of all inpatients with confirmed pandemic influenza A (H1N1) at a provincial pediatric hospital. A total of 41 cases attending the inpatient department between 15 November, 2009 to 14 December, 2009 were included. Prolonged and discontinuous shedding of 2009 H1N1 virus (median, 10 days; range, 2 to 24 days) were detected by real-time RT-PCR. The interval from onset of symptom to the start of oseltamivir therapy was an independent risk factor for prolonged virus shedding.

The authors report a case of hemophagocytic lymphohistiocytosis that became apparent and was confirmed by a specific Th1/Th2 cytokine pattern at the initiation of Kawasaki disease in an 18-month-old child. His condition deteriorated fast and produced no response to intravenous immunoglobulin and dexamethasone. A standard HLH-2004 regimen was started. But he developed respiratory failure, seizure, and cardiac arrest, and died. This case is unique for developing hemophagocytic lymphohistiocytosis at the initiation of Kawasaki disease, with fatal outcome. This case may indicate a yet unknown mechanism triggers these 2 diseases; a specific Th1/Th2 cytokine pattern helps rapid differential diagnosis between these 2 diseases.

Introduction and objectives Hemophogocytic syndrome (HPS) is an infrequent disease in children with a high mortality rate. Currently, the diagnosis of HPS largely depends on non-specific clinical manifestations, but the clinical features allowing to meet the criteria of diagnosis of HPS usually do not appear simultaneously which make the early diagnosis very difficult. According to the literature, hypercytokinemia has been noticed for this disease. However, the pattern of hypercytokinemia in HPS has not been proposed. In this study, the quick determination of Th1/Th2 cytokines in the sera from patients with HPS was performed and the patterns of the cytokines were analyzed to seek for the possibility of using the cytokine pattern for the early diagnosis of HPS and its prognostic significance. Patients and Methods A total of 16 children with HPS based on the 2004 HPS criteria from June 2005 ∼ December 2006 were enrolled into this study. Peripheral blood samples were collected at the time of diagnosis, during the induction therapy and at the time of relapse. Sera from 21 healthy children, 17 ALL at remission and 17 with sepsis were used as controls. A cytometric bead assay (CBA) was used to quantitatively determine the Th1/Th2 cytokines including IL-2, IL-4, IL-6, IL-10, TNFα and IFNγ. Results Levels of IL-6 (22.20 (10.60–139.25) pg/ml), IL-10 (195.60(63.85–918.70)pg/ml) and IFNγ (4740.20(1054.72– >5000.00)pg/ml) from patients with HPS were significantly higher than those of normal control (3.35 (2.50–5.40) pg/ml, P < 0.05; 2.90(2.20–4.80)pg/ml, P5000.00)pg/ml) of IFNγ in HPS patients were much higher than those (52.80(37.50∼66.60)pg/ml, P5000.00)pg/ml, P=0.002) of prior to treatment. While those in patients insensitive to chemotherapy were not significantly reduced (P > 0.05). No significant differences in the IL-2, IL-4 and TNFα levels among patients at acute phase, remission stage and the healthy controls were observed. Univariate analysis showed that the initial IL-10 level >100pg/ml and IFNγ concentration > 4000pg/ml were both unfavorable prognostic factors for HPS (IFNγ, P =0.034; IL-10, P=0.034). Positive correlations between the levels of IFNγ and IL-10(r=0.893, P Conclusion s The quick determination of Th1/Th2 cytokines is a useful approach for the early and quick diagnosis in HPS patients based on the pattern of significant elevations of IFNγ and IL-10 with slight elevation of IL-6 and it can be used for the monitoring of the progress and the prognosis of the disease.

Owing to the intensification of chemotherapy, the complete remission (CR), overall survival (OS), disease free survival (DFS) and event free survival (EFS) rates in childhood AML have been significantly improved during the past two decades in western countries. However, reports on the long-term outcome of this disease in Chinese children remain to be very limited. Objectives The aims of this study were to investigate the outcome and prognostic factors in Chinese childhood AML treated in our hospital during the past 7 years and to provide the basis for further improvement of outcome in children with AML. Methods From April 1998 to August 2005, 96 children (51 boys and 45 girls, aged 1.5 ~ 16.1 yrs with a median age of 9) with de novo AML diagnosed in our hospital based on MICM criteria were enrolled into this study. In total, 57 patients with non-APL AML were treated with the induction therapy of daunorubicin (maximum total cumulative dose 360 mg/M2) + Ara-C (DA) or homoharringtonine + Ara-C (HA), post-remission therapy with high-dose (1~2 g/M2 q12h for 6 doses) Ara-C containing regimens for three-month courses, and low-dose therapy with daunorubicin, homoharringtonine, etoposide (maximum total cumulative dose 1800 mg/M2) and Ara-C for three-month courses, followed by maintenance therapy for a total of 2~2.5 yrs. The 39 patients with APL were treated with all-trans retinoic acid (ATRA) and drugs above. Results Out of 96 patients, 86 children (89.6%) achieved CR. The estimated OS rates at three, five and seven years were (67.2±5.5)%, (58.1±6.4)% and (49.6±8.0)%, respectively, while the EFS rates were (52.3±5.8)%, (47.6±6.2)% and (47.6±6.2)%, and the DFS rates were (55.9±6.0)%, (50.7±6.4)% and (50.7±6.4)%, respectively. Univariant analysis showed that the variables predicted higher CR rates included the WBC counts (93.2% for patients < 100 × 109/L vs 50.0% for those ≥ 100 × 109/L, P = 0.004) at diagnosis, and the blasts (100% for patients without blasts vs 85.2% for those with blasts, P = 0.023) in the peripheral blood. The variables predicted longer survival duration included WBC counts ((49.1±6.5)% of 5yr EFS for patients < 100 × 109/L vs (33.3±18.0)% for those ≥ 100 × 109/L, P = 0.022) at diagnosis, FAB subtypes ((55.9±9.7)% for M3 vs (40.4±7.7)% for non-M3, P =0.048), course numbers [(76.0±12.2)% of 5-yr ESF for 1 course vs (31.1±9.0)% for more than 2 courses, P = 0.026) for achieving CR, CD38+ ((57.9± 7.3)% for CD38+ vs (16.7%± 10.8)% for CD38−, P = 0.005), CD14− ((49.9± 7.3)% for CD14− vs (21.5%± 14.2)% for CD14+, P = 0.001), CD34− ((54.7± 9.7)% for CD34− vs (32.6%± 8.8)% for CD34+, P = 0.038) and HLA-DR- ((57.7± 17.7)% for HLA-DR- vs (39.9%± 7.4)% for HLA-DR+, P = 0.024). 17 patients lost follow-up. The total cumulative relapse rate was 31.6% (25/79) with all BM relapse. The total treatment related modality rate was 5.3%. No secondary malignancy case was identified. Conclusions DA-based induction and high dose Ara-C-based post-remission treatment are effective to improve the CR rate and long-term survival in Chinese children with AML. The results are comparable to those reported in western countries. Initial WBC counts, early response to chemotherapy, FAB subtypes and some immunological markers are proved to be the significant prognostic factors for childhood AML under our current protocol.

Acute promyelocytic leukemia (APL) is a specific type of hematopoietic malignancy, accounting for ~ 10% of the de novo acute myeloid leukemia (AML). During the old days, severe complications as disseminated intravascular coagulation (DIC) and intracranial hemorrhage were the most common causes of treatment failure after conventional chemotherapy without all-trans retinoic acid (RA). Owing to the application of RA for the induction treatment, the overall survival (OS), the disease free survival (DFS) and the event free survival (EFS) rates have been dramatically improved in adult patients with APL. However, data on long-term outcome of APL in children, especially in Chinese children, have been very limited. Objective The aim of this study was to investigate the clinical biological features, diagnosis, prognosis and long-term survival in childhood APL. Methods 46 children (26 boys and 20 girls, aged 1.5 ~13.8 yrs with a median of 9.3 yrs) with APL from April 1998 to October 2005 were enrolled into this study. Immunophenotyping analysis was carried out in 43 patients using multi-parameter flow cytometry. 32 patients went through PML/RARα fusion gene detection using RT-PCR. Induction treatment consisted of ATRA and daunorubicin(DNR) or pirarubicin (THP) followed by 6 courses of multi-drug chemotherapy consolidation and a long-term maintenance therapy including ATRA, high dose Ara-C (HD-Ara-C), DNR+Ara-C (DA), homoharringtonine+Ara-C (HA) and etoposide+Ara-C (EA). Results Pale, hemorrhage and fever were the most common symptoms in APL patients at the time of presentation. 19 patients (41.3%) were found to have WBC count more than 10.0×109/L at diagnosis. Immunophenotyping analysis showed that CD13, CD33, CD117 and MPO were the most commonly expressed antigens while HLA-DR, CD14 and CD34 were mostly the negative markers on APL cells. 71.9%(23/32) of the patients analyzed were PML/RARα fusion gene positive. Of the 39 patients receiving treatment, 36 children (92.3%) achieved complete remission. 7 children replased during therapy, and 3 relapsed after finishing the entire courses. The 5-year cumulative incidence of relapse (CIR) was 28.6%. The estimated overall survival (OS) rates at one, three and five years were 86.1%, 76.1%and 70.2%, respectively while the event free survival (EFS) rates were 78.4%, 63.6%and 53.1%, respectively. The probability of remaining alive after 5 years for patients with WBC≤10.0×109/L group was 81.4%, significantly higher than those with WBC>10.0×109/L group (51.6%, P=0.026). 5 children with positive PML/RARα S (short) subtype died eventually although all of them achieved CR, which was significantly different from the group with the L (long) subtype (13/13, P=0.001). Conclusion Induction with ATRA + DNR or THP is an effective and safe therapy for newly diagnosed childhood APL with very high long-term survival rates after 2.5 years of alternative multi-drug chemotherapy and maintenance. High WBC count and S subtype of PML-RARα are the two poor prognostic factors for children with APL.