Single-cell transcriptome reveals highly complement activated microglia cells in association with pediatric tuberculous meningitis.

Background: Tuberculous meningitis (TBM) is a devastating form of tuberculosis (TB) causing high mortality and disability. TBM arises due to immune dysregulation, but the underlying immune mechanisms are unclear. Methods: We performed single-cell RNA sequencing on peripheral blood mononuclear cells (PBMCs) and cerebrospinal fluid (CSF) cells isolated from children (n=6) with TBM using 10 xGenomics platform. We used unsupervised clustering of cells and cluster visualization based on the gene expression profiles, and validated the protein and cytokines by ELISA analysis. Results: We revealed for the first time 33 monocyte populations across the CSF cells and PBMCs of children with TBM. Within these populations, we saw that CD4_C04 cells with Th17 and Th1 phenotypes and Macro_C01 cells with a microglia phenotype, were enriched in the CSF. Lineage tracking analysis of monocyte populations revealed myeloid cell populations, as well as subsets of CD4 and CD8 T-cell populations with distinct effector functions. Importantly, we discovered that complement-activated microglial Macro_C01 cells are associated with a neuroinflammatory response that leads to persistent meningitis. Consistently, we saw an increase in complement protein (C1Q), inflammatory markers (CRP) and inflammatory factor (TNF-α and IL-6) in CSF cells but not blood. Finally, we inferred that Macro_C01 cells recruit CD4_C04 cells through CXCL16/CXCR6. Discussion: We proposed that the microglial Macro_C01 subset activates complement and interacts with the CD4_C04 cell subset to amplify inflammatory signals, which could potentially contribute to augment inflammatory signals, resulting in hyperinflammation and an immune response elicited by Mtb-infected tissues.

p53 suppresses the inflammatory response following respiratory syncytial virus infection by inhibiting TLR2.

-Respiratory syncytial virus (RSV) is a pivotal virus leading to acute lower respiratory tract infections in children under 5 years old. This study aimed to explore the correlation between p53 and Toll-like receptors (TLRs) post RSV infection. p53 levels exhibited a substantial decrease in nasopharyngeal aspirates (NPAs) from infants with RSV infection compared to control group. Manipulating p53 expression had no significant impact on RSV replication or interferon signaling pathway. Suppression of p53 expression led to heightened inflammation following RSV infection in A549 cells or airways of BALB/c mice. while stabilizing p53 expression using Nutlin-3a mitigated the inflammatory response in A549 cells. Additionally, Inhibiting p53 expression significantly increased Toll-like receptor 2 (TLR2) expression in RSV-infected epithelial cells and BALB/c mice. Furthermore, the TLR2 inhibitor, C29, effectively reduced inflammation mediated by p53 in A549 cells. Collectively, our results indicate that p53 modulates the inflammatory response after RSV infection through TLR2.

Polystyrenenanoplastics lead to ferroptosis in the lungs.

INTRODUCTION: It has been shown that polystyrenenanoplastic (PS-NP) exposure induces toxicity in the lungs. OBJECTIVES: This study aims to provide foundational evidence to corroborate that ferroptosis and abnormal HIF-1α activity are the main factors contributing to pulmonary dysfunction induced by PS-NP exposure. METHODS: Fifty male and female C57BL/6 mice were exposed to distilled water or 100 nm or 200 nm PS-NPs via intratracheal instillation for 7 consecutive days. Hematoxylin and eosin (H&E) and Masson trichrome staining were performed to observe the histomorphological changes in the lungs. To clarify the mechanisms of PS-NP-induced lung injury, we used 100 µg/ml, 200 µg/ml and 400 µg/ml 100 or 200 nm PS-NPs to treat the human lung bronchial epithelial cell line BEAS-2B for 24 h. RNA sequencing (RNA-seq) of BEAS-2B cells was performed following exposure. The levels of glutathione, malondialdehyde, ferrous iron (Fe2+), and reactive oxygen species (ROS) were measured. The expression levels of ferroptotic proteins were detected in BEAS-2B cells and lung tissues by Western blotting. Western blotting, immunohistochemistry, and immunofluorescence were used to evaluate the HIF-1α/HO-1 signaling pathway activity. RESULTS: H&E staining revealed substantial perivascular lymphocytic inflammation in a bronchiolocentric pattern, and Masson trichrome staining demonstrated critical collagen deposits in the lungs after PS-NP exposure. RNA-seq revealed that the differentially expressed genes in PS-NP-exposed BEAS-2B cells were enriched in lipid metabolism and iron ion binding processes. After PS-NP exposure, the levels of malondialdehyde, Fe2+, and ROS were increased, but glutathione level was decreased. The expression levels of ferroptotic proteins were altered significantly. These results verified that PS-NP exposure led to pulmonary injury through ferroptosis. Finally, we discovered that the HIF-1α/HO-1 signaling pathway played an important role in regulating ferroptosis in the PS-NP-exposed lung injury. CONCLUSION: PS-NP exposure caused ferroptosis in bronchial epithelial cells by activating the HIF-1α/HO-1 signaling pathway, and eventually led to lung injury.

Andrographolide Attenuates RSV-induced Inflammation by Suppressing Apoptosis and Promoting Pyroptosis after Respiratory Syncytial Virus Infection In Vitro.

BACKGROUND: Respiratory syncytial virus (RSV), which is the predominant viral pathogen responsible for causing acute lower respiratory tract infections in children, currently lacks specific therapeutic drugs. Despite andrographolide's demonstrated effectiveness against various viral infections, its effects on RSV infection remain unclear. METHODS: In this study, RSV infection and andrographolide-intervened A549 cell lines were used. The virus load of RSV and the levels of IL-6 and IL-8 in the cell supernatant were quantified. The potential targets of andrographolide in the treatment of RSV-infected airway epithelial cells were analyzed using the Gene Expression Omnibus (GEO) database and the PharmMapper Database, and the changes in mRNA expression of these target genes were measured. To further illustrate the effect of andrographolide on the death pattern of RSV-infected airway epithelial cells, Annexin V-FITC/PI apoptosis assays and Western blotting were conducted. RESULTS: Andrographolide decreased the viral load and attenuated IL-6 and IL-8 levels in cell supernatant post-RSV infection. A total of 25 potential targets of andrographolide in the treatment of RSV-infected airway epithelial cells were discovered, and CASP1, CCL5, JAK2, and STAT1 were identified as significant players. Andrographolide noticeably suppressed the increased mRNA expressions of these genes post-RSV infection as well as IL-1ß. The flow cytometry analysis demonstrated that andrographolide alleviated apoptosis in RSV-infected cells. Additionally, RSV infection decreased the protein levels of caspase-1, cleaved caspase-1, cleaved IL-1ß, N-terminal of GSDMD, and Bcl-2. Conversely, andrographolide increased their levels. CONCLUSION: These results suggest that andrographolide may reduce RSV-induced inflammation by suppressing apoptosis and promoting pyroptosis in epithelial cells, leading to effective viral clearance.

Andrographolide exerts anti-respiratory syncytial virus activity by up-regulating heme oxygenase-1 independent of interferon responses in human airway epithelial cells.

BACKGROUND: Respiratory syncytial virus (RSV) is the leading cause of mortality and morbidity in children under the age of five. Despite this, there is still a lack of safe and effective vaccines and antiviral agents for clinical use. Andrographolide exerts antiviral functions against a variety of viruses, but whether (and how) it exerts antiviral effects on RSV remains unclear. METHODS AND RESULTS: In vitro RSV infection models using A549 and 16HBE cell lines were established, and the effects of andrographolide on RSV were analyzed via RSV N gene load and proinflammatory cytokine levels. The RNA transcriptome was sequenced, and data were analyzed by R software. Andrographolide-related target genes were extracted via network pharmacology using online databases. Lentiviral transfection was applied to knockdown the heme oxygenase-1 gene (Hmox1, HO-1). Results showed that andrographolide suppressed RSV replication and attenuated subsequent inflammation. Network pharmacology and RNA sequencing analysis indicated that the hub gene HO-1 may play a pivotal role in the anti-RSV effects of andrographolide. Furthermore, andrographolide exerted antiviral effects against RSV partially by inducing HO-1 but did not activate the antiviral interferon response. CONCLUSION: Our findings demonstrated that andrographolide exerted anti-RSV activity by up-regulating HO-1 expression in human airway epithelial cells, providing novel insights into potential therapeutic targets and drug repurposing in RSV infection.

Breastfeeding of infants born to mothers with COVID-19: a rapid review.

BACKGROUND: Existing recommendations on whether mothers with COVID-19 should continue breastfeeding are still conflicting. We aimed to conduct a rapid review of mother-to-child transmission of COVID-19 during breastfeeding. METHODS: We systematically searched Medline, Embase, Web of Science, Cochrane Library, China Biology Medicine disc, China National Knowledge Infrastructure, Wanfang, and preprint articles up to March 2020. We included studies relevant to transmission through milk and respiratory droplets during breastfeeding of mothers with COVID-19, SARS, MERS and influenza. Two reviewers independently screened studies for eligibility, extracted data, assessed risk of bias and used GRADE to assess certainty of evidence. RESULTS: A total of 4,481 records were identified in our literature search. Six studies (five case reports and one case series) involving 58 mothers (16 mothers with COVID-19, 42 mothers with influenza) and their infants proved eligible. Five case reports showed that the viral nucleic acid tests for all thirteen collected samples of breast milk from mothers with COVID-19 were negative. A case series of 42 influenza infected postpartum mothers taking precautions (hand hygiene and wearing masks) before breastfeeding showed that no neonates were infected with influenza during one-month of follow-up. CONCLUSIONS: The current evidence indicates that SARS-CoV-2 viral nucleic acid has not been detected in breast milk. The benefits of breastfeeding may outweigh the risk of SARS-CoV-2 infection in infants. Mothers with COVID-19 should take appropriate precautions to reduce the risk of transmission via droplets and close contact during breastfeeding.

[Clinical effect of azithromycin adjuvant therapy in children with bronchiolitis: a systematic review and Meta analysis].

OBJECTIVE: To systematically evaluate the clinical effect of azithromycin (AZM) adjuvant therapy in children with bronchiolitis. METHODS: Related databases were searched for randomized controlled trials (RCTs) on AZM adjuvant therapy in children with bronchiolitis published up to February 17, 2019. RevMan 5.3 was used to perform the Meta analysis. RESULTS: A total of 14 RCTs were included, with 667 children in the intervention group and 651 in the control group. The pooled effect size showed that in the children with bronchiolitis, AZM adjuvant therapy did not shorten the length of hospital stay (MD=-0.29, 95%CI: -0.62 to 0.04, P=0.08) or oxygen supply time (MD=-0.33, 95%CI: -0.73 to 0.07, P=0.10), while it significantly shortened the time to the relief of wheezing (MD=-1.00, 95%CI: -1.72 to -0.28, P=0.007) and cough (MD=-0.48, 95%CI: -0.67 to -0.29, P<0.00001). The analysis of bacterial colonization revealed that AZM therapy significantly reduced the detection rates of Streptococcus pneumoniae (OR=0.24, 95%CI: 0.11-0.54, P=0.0006), Haemophilus (OR=0.28, 95%CI: 0.14-0.55, P=0.0002), and Moraxella catarrhalis (OR=0.21, 95%CI: 0.11-0.40, P<0.00001) in the nasopharyngeal region. CONCLUSIONS: AZM adjuvant therapy can reduce the time to the relief of wheezing and cough in children with bronchiolitis, but it has no marked effect on the length of hospital stay and oxygen supply time.