Etiological Diagnosis and Treatment Value of mNGS in Alveolar Lavage Fluid of Children with Severe Pneumonia.

Objective: Metagenomic sequencing (mNGS) is a promising technique for pathogen detection. However, the use of mNGS in pediatric lung infections is still rarely reported. Methods: A total of 59 cases were included between January 2019 and December 2021. To compare the performance of mNGS and routine detection in diagnosing pulmonary infection and identifying pathogenic bacteria. Results: 59 children (33.90%) were infected with Mycoplasma pneumoniae, 15 were infected with adenovirus type 7 (25.42%), 11 were infected with Streptococcus pneumoniae (18.64%), 6 were infected with Staphylococcus aureus (10.17%), 5 were infected with Klebsiella pneumoniae, Acinetobacter baumannii, pertussis, and oral streptococcus (8.47%), 3 were infected with Haemophilus influenzae (5.08%), and 3 were infected with Pseudomonas aeruginosa (5.08%). Among 59 patients, 41 were co-infected with multiple pathogens and 18 were infected with independent pathogens. Mycoplasma pneumoniae infection was most common in 6 out of 18 independent pathogen patients (33%). Among them were 3 cases of type 7 adenovirus, 53 cases of human herpesvirus (16.7%), and 2 cases of pertussis bacillus (11.1%). One case (5.6%) was infected with Streptococcus pneumoniae, Proteus albicans, Mycobacterium tuberculosis, Staphylococcus aureus, and Klebsiella pneumoniae. In cases where routine testing results are negative, mNGS improves the diagnostic efficiency of mixed pulmonary infections. 17 cases (17/59=28.81%) were diagnosed as single infection through routine testing, and the mNGS results of 4 cases were consistent with routine testing, all of which were Mycoplasma pneumoniae infections. Three cases were tested for partial mNGS matching, of which one case tested positive for Mycoplasma pneumoniae antibody and tested positive for Mycoplasma pneumoniae infection and human infection γ Herpes virus type 4, one case tested positive for antibodies and tested for infection with Mycoplasma pneumoniae and Streptococcus pneumoniae. The routine examination results of the remaining 11 patients were inconsistent with the mNGS examination results. Two patients tested positive for Mycoplasma antibodies. Patient 17 presented with mNGS infection of Haemophilus, Neisseria, and adenovirus type 7, while another patient 18 presented with herpes virus type 5 infection. Conclusion: mNGS is a promising technique for detecting co-pathogens in mixed pediatric lung infections, with potential benefits in terms of speed and sensitivity.

Cytokine Storm Related to CD4+ T Cells in Influenza Virus-Associated Acute Necrotizing Encephalopathy.

Acute necrotizing encephalopathy (ANE) is a rare but deadly complication with an unclear pathogenesis. We aimed to elucidate the immune characteristics of H1N1 influenza virus-associated ANE (IANE) and provide a potential therapeutic approach for IANE. Seven pediatric cases from a concentrated outbreak of H1N1 influenza were included in this study. The patients' CD4+ T cells from peripheral blood decreased sharply in number but highly expressed Eomesodermin (Eomes), CD69 and PD-1, companied with extremely high levels of IL-6, IL-8 in the cerebrospinal fluid and plasma. Patient 2, who showed high fever and seizures and was admitted to the hospital very early in the disease course, received intravenous tocilizumab and subsequently showed a reduction in temperature and a stable conscious state 24 h later. In conclusion, a proinflammatory cytokine storm associated with activated CD4+ T cells may cause severe brain pathology in IANE. Tocilizumab may be helpful in treating IANE.

UHPLC/MS-based metabolomics of asthmatic mice reveals metabolic changes in group 2 innate lymphoid cells.

Helper Th2-type immune responses are essential in allergic airway diseases, including asthma and allergic rhinitis. Recent studies have indicated that group 2 innate lymphoid cells (ILC2s) play a crucial role in the occurrence and development of asthma. However, the metabolic profile of ILC2s and their regulatory mechanisms in asthma remain unclear. Therefore, we established two asthma mouse models: an ovalbumin (OVA)-induced asthma model and an IL-33-induced asthma model. We then used ultra-high-performance liquid chromatography/mass spectrometry (UHPLC/MS) to conduct high-throughput untargeted metabolic analysis of ILC2s in the lung tissues of the asthma models. The identified metabolites primarily consisted of lipids, lipid-like molecules, benzene, organic acids, derivatives, and organic oxidation compounds. Specifically, 34 differentially accumulated metabolites influenced the metabolic profiles of the control and OVA-induced asthma model groups. Moreover, the accumulation of 39 metabolites significantly differed between the Interleukin 33 (IL-33) and control groups. These differentially accumulated metabolites were mainly involved in pathways such as sphingolipid, oxidative phosphorylation, and fatty acid metabolism. This metabolomic study revealed, for the first time, the key metabolites and metabolic pathways of ILC2s, revealing new aspects of cellular metabolism in the context of airway inflammation. These findings not only contribute to unraveling the pathogenesis of asthma but also provide a crucial theoretical foundation for the future development of therapeutic strategies targeting ILC2s.

Combined transcriptomics and TMT-proteomics reveal abnormal complement and coagulation cascades in cow's milk protein allergy.

Cow's milk protein allergy (CMPA) is primarily due to the inability of the intestinal mucosa to establish typical immunological tolerance to proteins found in cow's milk, and the specific molecular mechanism is still unclear. In order to investigate molecular alterations in intestinal tissues during CMPA occurrence, this study analyzed the jejunal tissue of ß-lactoglobulin (BLG)-sensitized mice through transcriptomics and quantitative tandem mass tag (TMT)-labeled proteomics. A total of 475 differentially expressed genes (256 up-regulated, 219 down-regulated) and 94 differentially expressed proteins (65 up-regulated, 29 down-regulated) were identified. Comparing the KEGG pathways of the two groups, it was found that both were markedly enriched in the signaling pathways of complement and coagulation cascade. Among these, kallikrein B1 (KLKB1) in this pathway is speculated to be pivotal in CMPA. It may potentially enhance the release of bradykinin by activating the kallikrein-kinin system, leading to pro-inflammatory effects and exacerbating intestinal mucosal damage. This study suggests that the pathways of complement and coagulation cascades could be significant in the context of intestinal immunity in CMPA, and KLKB1 may be its potential therapeutic target.

Clinical report and genetic analysis of a novel variant in ZMIZ1 causing neurodevelopmental disorder with dysmorphic factors and distal skeletal anomalies in a Chinese family.

BACKGROUND: Neurodevelopmental disorder with dysmorphic factors and distal skeletal anomalies (NEDDFSA) is a rare and phenotypically variable disorder. The zinc finger MIZ-type containing 1 gene (ZMIZ1) is a causative gene of NEDDFSA that encodes a protein inhibitor of the activated STAT-like family transcriptional regulator. Given the rarity of reported NEDDFSA cases, new phenotypes and genotypes of this disorder are still being discovered. OBJECTIVE: This study describes the phenotype characteristics of a Chinese NEDDFSA family caused by a novel ZMIZ1 variant. METHODS: We reviewed the clinical phenotype of a Chinese patient with NEDDFSA and performed whole-exome sequencing (WES) of the patient's family. We simulated the potential biological harmfulness of the mutant protein. Plasmids were constructed and used for western blot and immunofluorescence assays to analyze protein expression levels. RESULTS: The patient was a 6-month-old male infant who exhibited dysmorphic facial features, neurodevelopmental abnormalities, congenital heart disease, and previously unreported genitourinary system anomalies. WES revealed a non-frameshift deletion variant in ZMIZ1 (NM_020338.4: c.858_875del, p.Val288_Ala293del), resulting in a structural alteration in the protein's alanine-rich domain. Western blot and immunofluorescence assays indicated a significant decrease in the expression level of the mutant ZMIZ1 protein compared to the wild-type protein. CONCLUSION: The clinical manifestations of this patient may be associated with the ZMIZ1 variant, and the structural alteration in the alanine-rich domain of the ZMIZ1 protein may contribute to a more complex disease phenotype. These results expand the genotype-phenotype correlation of ZMIZ1.

ATG2B upregulated in LPS-stimulated BMSCs-derived exosomes attenuates septic liver injury by inhibiting macrophage STING signaling.

Pretreated mesenchymal stem cells (MSCs)-derived exosomes have shown great potential in the treatment of various inflammatory diseases. Recent evidence suggests that macrophage stimulator of interferon genes (STING) signal activation plays a critical role in sepsis and septic liver injury. Here, we aimed to investigate the role and effects of lipopolysaccharide (LPS)-pretreated bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (L-Exo) on macrophage STING signaling in septic liver injury. Exosomes were collected from the BMSCs medium via ultracentrifugation. Liver injury, intrahepatic inflammation, and the activation of macrophage STING signaling were analyzed. Mitophagy and the release of mitochondrial DNA (mtDNA) into the cytosol were investigated. Through in vivo and in vitro experiments, L-Exo could markedly attenuate cecal ligation and puncture-induced septic liver injury and inhibit macrophage STING signaling. Mechanistically, L-Exo inhibited macrophage STING signaling by enhancing mitophagy and inhibiting the release of mtDNA into the cytosol. Furthermore, autophagy-related protein 2 homolog B (ATG2B) may be a major factor involved in this effect of L-Exo. These findings reveal that macrophage STING signaling plays an important role in septic liver injury and may be a therapeutic target. In addition, LPS pretreatment is an effective and promising approach for optimizing the therapeutic efficacy of MSCs-derived exosomes in septic liver injury, providing new strategies for treatment.

[Analysis of clinical features and ADNP variant in a child with Helsmoortel-Van der Aa syndrome].

OBJECTIVE: To analyze the clinical characteristics and genetic etiology of a child with Helsmoortel-Van der Aa syndrome (HVDAS). METHODS: Genetic testing was carried out for the child and his parents, and the clinical phenotypes and genetic variants of reported cases were summarized through literature review. RESULTS: The child has featured peculiar facies, accompanied by autism spectrum disorder, intellectual disability and motor retardation, and curving of the second toes, which was unreported previously. Genetic testing revealed that the child has harbored a heterozygous c.2157C>G (p.Tyr719*) variant of the ADNP gene, which was not found in either parent. Based on the guidelines of the American College of Medical Genetics and Genomics, this variant was rated as pathogenic. Among 80 HVDAS cases described in the literature, most had various degrees of behavioral abnormalities, intellectual disability, language retardation and motor retardation, with common features involving the nervous system, gastrointestinal system and eye. Variants of the ADNP gene mainly included frameshift variants and nonsense variants, with the hotspot variants including p.Tyr719*, p.Asn832lysfs*81 and p.Arg730*. CONCLUSION: The clinical phenotype of the child is closely correlated with the heterozygous variant of the ADNP gene, which expanded the phenotypic spectrum of HVDAS. As HVDAS may involve multiple systems and have high phenotypic heterogeneity, genetic testing technology can facilitate accurately diagnose.

Surfactant Therapy for Respiratory Distress Syndrome in High- and Ultra-High-Altitude Settings.

Objective: The objective of this study is to investigate the therapeutic effect of surfactant replacement therapy (SRT) on respiratory distress syndrome (RDS) in premature infants in the Qinghai-Tibet Plateau. Materials and Methods: This multi-center retrospective cohort study collected and screened reasonable clinical data of 337 premature infants with RDS from 10 hospitals in the Qinghai-Tibet Plateau from 2015 to 2017. We grouped the cases by rationally analyzing their baseline characteristics, using logistic analysis to evaluate each factor's effect on the prognosis of the infants, and comparing the short-term improvement in blood gas and mortality after SRT treatment at different altitudes, in high-altitude (1,500-3,500 m) and ultra-high-altitude (3,500-5,500 m) groups. Results: Independent of altitude, the mortality rate of children with RDS in the SRT group was significantly lower than that of children in the non-SRT group (both P < 0.05). The effect of SRT on preterm infants with RDS in the high-altitude group [odds ratio (OR) = 0.44, 95% confidence interval (CI) = 0.22-0.87, P = 0.02] was better than that in the infants in the ultra-high-altitude group (OR = 0.26, 95% CI = 0.13-0.58, P < 0.01), with death rates of 34.34 and 49.71%, respectively. Similarly, after SRT, the improvement of PaO2/FiO2 and pH of children at high altitude was significantly better than those of children at ultra-high altitude (all P < 0.01). Conclusions: SRT plays a prominent role in curing infants with RDS in both high- and ultra-high-altitude regions, although with better effects at high rather than ultra-high altitude. This study provides a basis for further large-scale studies on SRT for RDS treatment at high altitudes.

[Treatment and genetic analysis of a child with Kabuki syndrome type 2 and secondary pulmonary infection due to a de novo variant of KDM6A gene].

OBJECTIVE: To explore the genetic basis of a child with recurrent infection, multiple malformation and dysmorphism. METHODS: The child and his parents were subjected to trio whole exome sequencing. RESULTS: The child had a complaint of fever and cough, with long and thin eye fissures and long eyelashes. Genetic testing revealed that the child has carried a non-triplet deletion of the KDM6A gene, which was unreported previously. The variant resulted in frameshift and premature termination of the translation. His parents were both of the wild type for the locus. After antibiotic and immunoglobulin treatment, the severe secondary pneumonia caused by immunodeficiency has improved. CONCLUSION: With combined laboratory test, imaging examination and genetic testing, the child was ultimately diagnosed with Kabuki syndrome type 2. The characteristics of immunodeficiency of Kabuki syndrome may render conventional antibiotic treatment ineffective, which deserves clinical attention.

[Value of sTREM-1 in serum and bronchoalveolar lavage fluid, APACHE II score, and SOFA score in evaluating the conditions and prognosis of children with severe pneumonia].

OBJECTIVE: To study the significance of the level of soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) in serum and bronchoalveolar lavage fluid (BALF), Acute Physiology and Chronic Health Evaluation II (APACHE II) score, and Sequential Organ Failure Assessment (SOFA) score in evaluating the conditions and prognosis of children with severe pneumonia. METHODS: A total of 76 children with severe pneumonia who were admitted from August 2017 to October 2019 were enrolled as the severe pneumonia group. According to the treatment outcome, they were divided into a non-response group with 34 children and a response group with 42 children. Ninety-four children with common pneumonia who were admitted during the same period of time were enrolled as the common pneumonia group. One hundred healthy children who underwent physical examination in the outpatient service during the same period of time were enrolled as the control group. The serum level of sTREM-1, APACHE II score, and SOFA score were measured for each group, and the level of sTREM-1 in BALF was measured for children with severe pneumonia. The correlation of the above indices with the severity and prognosis of severe pneumonia in children was analyzed. RESULTS: The severe pneumonia group had significantly higher serum sTREM-1 level, APACHEII score, and SOFA score than the common pneumonia group and the control group (P<0.05). For the children with severe pneumonia, the non-response group had significant increases in the levels of sTREM-1 in serum and BALF and SOFA score on day 7 after admission, while the response group had significant reductions in these indices, and there were significant differences between the two groups (P<0.05). Positive correlation was found between any two of serum sTREM-1, BALF sTREM-1, and SOFA score (P<0.05). APACHE II score was not correlated with serum sTREM-1, BALF sTREM-1, and SOFA score (P>0.05). CONCLUSIONS: The level of sTREM-1 in serum and BALF and SOFA score can be used to evaluate the severity and prognosis of severe pneumonia in children.

[Intervention measures for maintenance of clinical control in the remission stage of childhood asthma].

OBJECTIVE: To explore the intervention measures to maintain clinical control in children with asthma in the remission stage when concomitant with acute upper respiratory infection (AURI). METHODS: A total of 100 asthmatic children who had achieved clinical control were randomly divided into observation group and control group. The two groups were both treated with a combination of inhaled corticosteroids and long-acting ß2 receptor agonist (ICS/LABA) at the lowest dose every night. Conventional therapies were used for the two groups when suffering from AURI. In addition to conventional therapies, the observation group was given early short-term upgrade therapy, i.e., on the basis of maintenance therapy, the same amount of ICS/LABA complex preparation was inhaled every morning, which lasted for 7-10 days. Both groups were treated following asthma guidelines according to the severity of the disease at the time of acute attacks. The control rate of asthma, severity of acute attacks, changes in pulmonary function indices, and occurrence of adverse events were evaluated after 3, 6, 9, and 12 months of treatment. RESULTS: At each time point of follow-up, the rate of asthma control in the observation group was significantly higher than that in the control group (90% vs 80%; P<0.05). The severity of acute attacks in the observation group was significantly lower than that in the control group at all follow-up time points (P<0.05). Compared with the control group, the observation group had significantly improved pulmonary function indices of large and small airways (P<0.05) and significantly reduced mean amount of inhaled glucocorticoids and impact on family life (P<0.01). CONCLUSIONS: Early short-term upgrade therapy for children with asthma in the remission stage when concomitant with AURI can prevent acute attacks of asthma, raise the rate of asthma control and improve pulmonary function.

Pediatric Asthma and Food Allergy.

OBJECTIVE: To quantitatively summarize the evidence from observational studies on the relation between pediatric asthma and food allergy. METHODS: A literature search was conducted in Medline and EMBASE (August 2016). Two independent reviewers appraised the studies and extracted the estimates of interest. Methodological quality of the included studies was assessed using National Heart Lung and Blood Institute (NHLBI) Quality Assessment Tools. Data were pooled using random-effects meta-analysis. RESULTS: A total of 32 relevant studies were identified but only 8 studies met the inclusion criteria. Using random-effect model, food allergy showed strong association with asthma in children (OR = 2.87 [95% CI: 2.05-4.00]; P < 0.0001). CONCLUSIONS: This study suggested that food allergy is associated with an increased risk of asthma in children.

[Causes of chronic cough in children].

OBJECTIVE: To study the causes of chronic cough in children. METHODS: Retrospective analysis was carried out on the clinical data of 132 children with chronic cough from August 2010 to September 2011. RESULTS: Several conditions were found to contribute to chronic cough in children, including cough variant asthma (CVA, n=56), upper airway cough syndrome (UACS, n=44), infections/postinfectious cough (IC/PIC, n=22), allergic cough (AC, n=8), gastroesophageal reflux cough (GERC, n=5), and others (n=3). There was significant difference in the distribution of IC/IPC among an infant group (<1 year), a group of young children (>1 year), a group of preschool aged children (>3 years) and a group of school-age children (6-14 years) (χ2=11.638, P=0.001), and the infant group showed a significantly higher prevalence of IC/PIC than the other three age groups (P<0.05). IC/PIC was the main cause of chronic cough in the infant group, while CVA and UACS were the main causes in each of the other groups. A relatively large proportion of AC, CVA and UACS cases had a personal history of allergy, a family history of allergy/asthma and a history of exposure to harmful environments. CONCLUSIONS: CVA, UACS, and IC/PIC are main causes of chronic cough in children, varying among different age groups. Children with a personal history of allergy, family history of allergy/asthma and a history of exposure to harmful environment are more vulnerable to AC, CVA and UACS.

[Inoculation of bacille calmette guerin combined with intraperitoneal adenosine injection enhances antibacterial effect of macrophage against Mycobacterium tuberculosis in mice].

OBJECTIVE: The protection rate of inoculation with BCG vaccine is only 50 percent, and most of patients with tuberculosis had a history of BCG vaccine inoculation. Adenosine (ADO) has an immunomodulating effect; it promotes immune reaction by increasing number of macrophage and enhancing phagocytosis. The present study was designed to investigate if combined use of adenosine with BCG enhances the anti-Mycobacterium tuberculosis effect of macrophage in mice. METHODS: Fifty BALB/C mice were divided randomly into 3 groups: BCG group (n = 21), BCG plus ADO group (n = 21) and control group (n = 8). The mice in BCG and BCG plus ADO groups were inoculated with 0.1 ml BCG intradermally and the mice in BCG plus ADO group were injected intraperitoneally with ADO 30 mg/(kg.d) for 5 days. The mice in BCG group and control group were injected with NS 0.1 ml/d for 5 days. Six weeks after the last injection, all mice were challenged with intravenous 1 x 10(6) CFU human Mycobacterium tuberculosis virulent strain. After challenging, lung and spleen specimens were taken at the 10th, 20th and 30th days from the mice of BCG and BCG plus ADO groups and at the 30th day from mice in control group. The pathological examinations of lung and spleen sections were performed after HE staining and acid-fast staining, and detection of cell apoptosis was also performed. RESULTS: Consolidation with neutrophil infiltration was found in most of the lung tissue taken at the day 30; there were a lot of tuberculous granulomas and Mycobacterium tuberculosis in the lungs of control group. The alveolar septum in BCG gradually became wide and in interstitium lymphocyte infiltration dominated, and there were less tuberculous granulomas but there were large number of Mycobacterium tuberculosis in the lungs from 10th to 30th days after challenging. The widening of alveolar septum and consolidation of lung tissue in BCG plus ADO group became milder with monocytes infiltration, and there were few tuberculosis granulomas and Mycobacterium tuberculosis in the lungs from 10th to 30th days after challenging. CONCLUSION: ADO could increase the number of monocyte-macrophages and promoted anti-bacterial effects of these cells.