Leukotriene B4-induced neutrophil extracellular traps impede the clearance of Pneumocystis.

Pneumocystis pneumonia (PCP) is a fungal pulmonary disease with high mortality in immunocompromised patients. Neutrophils are essential in defending against fungal infections; however, their role in PCP is controversial. Here we aim to investigate the effects of neutrophil extracellular traps (NETs) on Pneumocystis clearance and lung injury using a mouse model of PCP. Intriguingly, although neutrophils play a fundamental role in defending against fungal infections, NETs failed to eliminate Pneumocystis, but instead impaired the killing of Pneumocystis. Mechanically, Pneumocystis triggered Leukotriene B4 (LTB4)-dependent neutrophil swarming, leading to agglutinative NET formation. Blocking Leukotriene B4 with its receptor antagonist Etalocib significantly reduced the accumulation and NET release of neutrophils in vitro and in vivo, enhanced the killing ability of neutrophils against Pneumocystis, and alleviated lung injury in PCP mice. This study identifies the deleterious role of agglutinative NETs in Pneumocystis infection and reveals a new way to prevent NET formation, which provides new insights into the pathogenesis of PCP.

Dynamic within-host cefiderocol heteroresistance caused by blaSHV-12 amplification in pandrug-resistant and hypervirulent Klebsiella pneumoniae sequence type 11.

AIMS: Although cefiderocol (FDC) is not prescribed in China, FDC-resistant pandrug-resistant hypervirulent Klebsiella pneumoniae (PDR-hvKp) is emerging. In this study, we performed FDC susceptibility testing of clinical Kp isolates to explore the prevalence of FDC-resistant isolates and the mechanism of FDC-resistance. METHODS: We retrospectively selected 151 carbapenem-resistant Kp isolates to assess FDC susceptibility. Seven isolates harboring blaSHV-12 from two patients were enrolled for whole-genome sequencing. The antimicrobial resistance, virulence, blaSHV-12 expression, and fitness costs in different media were examined. The amplification of blaSHV-12 was further investigated by qPCR and long-read sequencing. RESULTS: The 151 isolates showed a low MIC50/MIC90 (1/4 mg/L) of FDC. The seven isolates were ST11 PDR-hvKp, and two represented FDC-resistance (MIC=32 mg/L). The IncR/IncFII plasmids of two FDC-resistant isolates harbored 6 and 15 copies of blaSHV-12, whereas four FDC-susceptible isolates carried one copy and one harbored three copies. These blaSHV-12 genes concatenated together and were located within the same 7.3 kb fragment flanked by IS26, which contributed to the increased expression and FDC resistance without fitness costs. The amplification of blaSHV-12 and FDC resistance could be induced by FDC in vitro and reversed during continuous passage. CONCLUSIONS: The amplification of blaSHV-12 and the consequent dynamic within-host heteroresistance are important concerns for the rational application of antibiotics. Long-read sequencing might be a superior way to detect resistance gene amplification rapidly and accurately.

Comparative therapeutic efficacy of interferon alfa-2b and combination lopinavir/ritonavir plus interferon alfa-2b against SARS-CoV-2.

BACKGROUND: The outbreak of coronavirus disease 2019 (COVID-19) posed an enormous threat to public health. The use of antiviral drugs in patients with this disease have triggered people's attentions. Whether interferon alfa-2b or lopinavir/ritonavir (LPV/r) plus interferon alfa-2b treatment can against SARS-CoV-2 was unknown. The objectives of this study was to evaluate the efficacy and safety of interferon alfa-2b and LPV/r plus interferon alfa-2b for SARS-CoV-2 infection in adult patients hospitalized with COVID-19. METHODS: This is a retrospective cohort study of 123 patients confirmed SARS-CoV-2 infection by PCR on nasopharyngeal swab and symptoms between Jan. 13 and Apr. 23, 2020. All patients received standard supportive care and regular clinical monitoring. Patients were assigned to standard care group (n = 12), interferon alfa-2b group (n = 44), and combination LPV/r plus interferon alfa-2b group (n = 67). The primary endpoints were duration of required oxygen support and virus clearance time. Associations between therapies and these outcomes were assessed by Cox proportional hazards regression. RESULTS: Baseline clinical characteristics were not significantly different among the three groups (P > 0.05). No significant associations were observed between LPV/r/interferon alfa-2b and faster SARS-CoV-2 RNA clearance (HR, 0.85 [95% confidence interval (CI) 0.45-1.61]; P = 0.61 in interferon alfa-2b group vs HR, 0.59 [95% CI 0.32-1.11]; P = 0.10 in LPV/r plus interferon alfa-2b group). Individual therapy groups also showed no significant association with duration of required oxygen support. There were no significant differences among the three groups in the incidence of adverse events (P > 0.05). CONCLUSIONS: In patients with confirmed SARS-CoV-2 infection, no benefit was observed from interferon alfa-2b or LPV/r plus interferon alfa-2b treatment. The findings may provide references for treatment guidelines of patients with SARS-CoV-2 infection.

Carboxypeptidase B blocks ex vivo activation of the anaphylatoxin-neutrophil extracellular trap axis in neutrophils from COVID-19 patients.

BACKGROUND: Thrombosis and coagulopathy are highly prevalent in critically ill patients with COVID-19 and increase the risk of death. Immunothrombosis has recently been demonstrated to contribute to the thrombotic events in COVID-19 patients with coagulopathy. As the primary components of immunothrombosis, neutrophil extracellular traps (NETs) could be induced by complement cascade components and other proinflammatory mediators. We aimed to explore the clinical roles of NETs and the regulation of complement on the NET formation in COVID-19. METHODS: We recruited 135 COVID-19 patients and measured plasma levels of C5, C3, cell-free DNA and myeloperoxidase (MPO)-DNA. Besides, the formation of NETs was detected by immunofluorescent staining and the cytotoxicity to vascular endothelial HUVEC cells was evaluated by CCK-8 assay. RESULTS: We found that the plasma levels of complements C3 and MPO-DNA were positively related to coagulation indicator fibrin(-ogen) degradation products (C3: r = 0.300, p = 0.005; MPO-DNA: r = 0.316, p = 0.002) in COVID-19 patients. Besides, C3 was positively related to direct bilirubin (r = 0.303, p = 0.004) and total bilirubin (r = 0.304, p = 0.005), MPO-DNA was positively related to lactate dehydrogenase (r = 0.306, p = 0.003) and creatine kinase (r = 0.308, p = 0.004). By using anti-C3a and anti-C5a antibodies, we revealed that the complement component anaphylatoxins in the plasma of COVID-19 patients strongly induced NET formation. The pathological effect of the anaphylatoxin-NET axis on the damage of vascular endothelial cells could be relieved by recombinant carboxypeptidase B (CPB), a stable homolog of enzyme CPB2 which can degrade anaphylatoxins to inactive products. CONCLUSIONS: Over-activation in anaphylatoxin-NET axis plays a pathological role in COVID-19. Early intervention in anaphylatoxins might help prevent thrombosis and disease progression in COVID-19 patients.

Composition and Function of Neutrophil Extracellular Traps.

Neutrophil extracellular traps (NETs) are intricate fibrous structures released by neutrophils in response to specific stimuli. These structures are composed of depolymerized chromatin adorned with histones, granule proteins, and cytosolic proteins. NETs are formed via two distinct pathways known as suicidal NETosis, which involves NADPH oxidase (NOX), and vital NETosis, which is independent of NOX. Certain proteins found within NETs exhibit strong cytotoxic effects against both pathogens and nearby host cells. While NETs play a defensive role against pathogens, they can also contribute to tissue damage and worsen inflammation. Despite extensive research on the pathophysiological role of NETs, less attention has been paid to their components, which form a unique structure containing various proteins that have significant implications in a wide range of diseases. This review aims to elucidate the components of NETs and provide an overview of their impact on host defense against invasive pathogens, autoimmune diseases, and cancer.

Study on Respiratory Function and Hemodynamics of AIDS Patients with Respiratory Failure.

Objective: We performed a comparative analysis of respiratory function and hemodynamics among patients with Acquired Immunodeficiency Syndrome (AIDS)-associated respiratory failure and those with non-AIDS-associated respiratory failure. Methods: Data were collected from critically ill patients diagnosed with Acquired Immunodeficiency Syndrome who were admitted to the Intensive Care Unit (ICU) of Beijing Ditan Hospital, affiliated with Capital Medical University, between January 1, 2019, and December 31, 2019. We simultaneously gathered data from non-AIDS patients admitted to the ICU of Beijing Liangxiang Hospital within the same timeframe. A comparative study was performed to analyze clinical data from these two patient groups, encompassing parameters related to respiratory mechanics and hemodynamic indicators. Results: A total of 12 patients diagnosed with Acquired Immunodeficiency Syndrome (AIDS) and experiencing respiratory failure, along with 23 patients with respiratory failure independent of AIDS, were included in our study. Subsequently, a comparative analysis of clinical information was conducted between the two patient cohorts. Our findings demonstrate non-statistically significant differences between the two patient groups when assessing various indicators, encompassing peak airway pressure, plateau pressure, mean pressure, compliance, oxygenation index, and arterial partial pressure of carbon dioxide (P>0.05). Additionally, the comparison of multiple indicators encompassing mean arterial pressure, central venous pressure, cardiac output index, intrathoracic blood volume index, global end-diastolic volume index, extravascular lung water content, and pulmonary vascular permeability index revealed no statistically significant differences between the two patient groups (P>0.05). Ultimately, the Galileo respiratory system was utilized to assess the pressure-volume (P-V) curve of the experimental cohort, revealing a consistent and seamless trajectory devoid of noticeable points of inflection. Conclusion: No statistically significant differences were found in the respiratory function and hemodynamic profiles between patients diagnosed with AIDS presenting respiratory failure and those experiencing respiratory failure unrelated to AIDS. Additionally, the pressure-volume curve of individuals diagnosed with AIDS presenting respiratory failure displayed a seamless and uninterrupted trajectory devoid of discernible points of inflection. Hence, there might be constraints when utilizing P-V curve-based adjustments for positive end-expiratory pressure (PEEP) during mechanical ventilation in individuals diagnosed with AIDS presenting respiratory failure.

Exogenous glutathione reverses meropenem resistance in carbapenem-resistant Klebsiella pneumoniae.

Background: The rate of carbapenem-resistant Klebsiella pneumoniae (CRKP) infection has been increasing rapidly worldwide and, poses a significant risk to human health. Effective methods are urgently needed to address treatment failures related to antibiotic resistance. Recent research has reported that some drugs in combination with antibiotics have displayed synergistic killing of resistant bacteria. Here, we investigated whether glutathione (GSH) can synergize with meropenem, and enhance its effectiveness against CRKP. Methods: Synergistic activity was assessed by checkerboard and time-killing assays. The mechanism of these combinations was assessed by total ROS and membrane permeability assays. The bacterial metabolites were assessed by LC‒MS/MS. Results: The FICIs of GSH and meropenem were approximately 0.5 and the combined treatment with GSH and meropenem resulted in a more than 2log10 CFU/mL reduction in bacteria compared to the individual treatments. These findings indicated the synergistic effect of the two drugs. Moreover, the meropenem MIC of CRKP was reduced to less than 4 mg/L when combined with 6 mg/mL GSH, indicating that GSH could significantly reverse resistance to meropenem in bacteria. The production of ROS in bacteria was determined by flow cytometry. After adding GSH, the ROS in the GSH group and the combined group was significantly higher than that in the control and meropenem groups, but there was no significant difference between the combined and GSH groups. The metabolic disturbance caused by GSH alone and in combination with meropenem was significant intracellularly and extracellularly, especially in terms of glycerophospholipid metabolism, indicating that the synergistic effect of the combined use of GSH and meropenem was relevant to glycerophospholipid metabolism. In addition, we measured the cell membrane permeability. The cell membrane permeability of the combination group was significantly higher than that of the blank control or monotreatment groups. This confirmed that the GSH can serve as a meropenem enhancers by disturbing glycerophospholipid metabolism and increasing cell membrane permeability. Conclusion: GSH and meropenem display a synergistic effect, wherein GSH increases the sensitivity of CRKP to meropenem. The synergy and susceptibility effects are thought to related to the increased membrane permeability resulting from the perturbations in glycerophospholipid metabolism, presenting a novel avenue for CRKP treatment.

Clinical Course and Risk Factors for Liver Injury of Severe and Critical Patients with COVID-19.

Introduction: Information regarding the clinical course of COVID-19 patients with liver injury is very limited, especially in severe and critical patients. The objective of this study was to describe the characteristics and clinical course of liver function in patients admitted with severe and/or critical SARS-CoV-2 infection, as well as explore the risk factors that affect liver function in the enrolled COVID-19 patients. Methods: Information on clinical characteristics of 63 severe and critical patients with confirmed COVID-19 was collected. Data on patients' demographics, laboratory characteristics, laboratory examination, SARS-CoV-2 RNA results and liver test parameters were acquired and analyzed. Results: The incidence of abnormal aspartate aminotransferase, alanine aminotransferase, and total bilirubin in the critical group was significantly higher than in the severe group (respectively 81.48%, 81.49%, 62.67%, and 45.71%, 63.88%, 22.86%, p < 0.05). The time for liver function parameters to reach their extremes was approximately 2-3 weeks after admission. The independent factors associated with liver injury were patients with invasive ventilators, decreased percentages of neutrophils, lymphocytes and monocytes, and sequential organ failure assessment (SOFA) score ≥2 (p < 0.05). Conclusion: Abnormal liver tests are commonly observed in severe and critical patients with COVID-19. Severe patients infected by SARS-CoV-2 should be closely observed and monitored the liver function parameters, particularly when they present with independent risk factors for liver injury.

Presepsin Predicts Severity and Secondary Bacterial Infection in COVID-19 by Bioinformatics Analysis.

Introduction: Novel coronavirus pneumonia (COVID-19) is an acute respiratory disease caused by the novel coronavirus SARS-CoV-2. Severe and critical illness, especially secondary bacterial infection (SBI) cases, accounts for the vast majority of COVID-19-related deaths. However, the relevant biological indicators of COVID-19 and SBI are still unclear, which significantly limits the timely diagnosis and treatment. Methods: The differentially expressed genes (DEGs) between severe COVID-19 patients with SBI and without SBI were screened through the analysis of GSE168017 and GSE168018 datasets. By performing Gene Ontology (GO) enrichment analysis for significant DEGs, significant biological processes, cellular components, and molecular functions were selected. To understand the high-level functions and utilities of the biological system, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed. By analyzing protein-protein interaction (PPI) and key subnetworks, the core DEGs were found. Results: 85 DEGs were upregulated, and 436 DEGs were downregulated. The CD14 expression was significantly increased in the SBI group of severe COVID-19 patients (P < 0.01). The area under the curve (AUC) of CD14 in the SBI group in severe COVID-19 patients was 0.9429. The presepsin expression was significantly higher in moderate to severe COVID-19 patients (P < 0.05). Presepsin has a diagnostic value for moderate to severe COVID-19 with the AUC of 0.9732. The presepsin expression of COVID-19 patients in the nonsurvivors was significantly higher than that in the survivors (P < 0.05). Conclusion: Presepsin predicts severity and SBI in COVID-19 and may be associated with prognosis in COVID-19.

Multi-State Analysis of the Impact of Childhood Starvation on the Healthy Life Expectancy of the Elderly in China.

Background: Child malnutrition is not only common in developing countries but also an important issue faced by developed countries. This study aimed to explore the influence and degree of childhood starvation on the health of the elderly, which provides a reference for formulating health-related policies under the concept of full lifecycle health. Methods: Based on the Chinese Longitudinal Healthy Longevity Survey (CLHLS) in 2008, 2011, and 2014, this study took a total of 13,185 elderly people aged 65-99 years as the target population. By IMaCH software, with gender and income level as the control variables, the average life expectancy and healthy life expectancy of the elderly were measured. The x2test was used to explore the differences in the socioeconomic status of elderly people with or without starvation in childhood. Statistical differences between average life expectancy and healthy life expectancy were analyzed by rank tests. Results: (1) The results showed that there was a statistically significant difference in age, gender, residency, education level, and income level between the groups with or without starvation (P < 0.05). (2) Transition probabilities in health-disability, health-death, and disability-death all showed an upward trend with age (P < 0.05), where the elderly who experienced starvation in childhood were higher than those without such an experience (P < 0.05). However, the probability of disability-health recovery showed a downward trend with age (P < 0.05), in which the elderly who experienced starvation in childhood were lower than those without starvation (P < 0.05). (3) For the elderly who experienced starvation in childhood, the health indicators of the average life expectancy, healthy life expectancy, and healthy life expectancy proportion accounted for the remaining life were lower than those of the elderly without childhood starvation (P < 0.05). Conclusions: The average life expectancy and healthy life expectancy of the elderly with childhood starvation are lower than those without childhood starvation. It shows that the negative impact of childhood starvation on health through the life course till old age has a persistent negative cumulative effect on the quantity and quality of life. Therefore, it is important to pay attention to the nutritional status of children in poor families from the perspective of social policymaking.

Bacterial and fungal co-infections among COVID-19 patients in intensive care unit.

This study aimed to investigate the frequency and characteristics of respiratory co-infections in COVID-19 patients in the intensive care unit (ICU). In this retrospective observational study, pathogens responsible for potential co-infections were detected by the bacterial culture, real-time polymerase chain reaction (RT-PCR), or serological fungal antigen tests. Demographic and clinical characteristics, as well as microbial results, were analyzed. Bacterial culture identified 56 (58.3%) positive samples for respiratory pathogens, with the most common bacteria being Burkholderia cepacia (18, 18.8%). RT-PCR detected 38 (76.0%) and 58 (87.9%) positive results in the severe and critical groups, respectively. Most common pathogens detected were Stenotrophomonas maltophilia (28.0%) and Pseudomonas aeruginosa (28.0%) in the severe group and S. maltophilia (45.5%) in the critical group. P. aeruginosa was detected more during the early stage after ICU admission. Acinetobacter baumannii and Staphylococcus aureus were more frequently identified during late ICU admission. Fungal serum antigens were more frequently positive in the critical group than in the severe group, and the positive rate of fungal serum antigens frequency increased with prolonged ICU stay. A high frequency of respiratory co-infections presented in ICU COVID-19 patients. Careful examinations and necessary tests should be performed to exclude these co-infections.

Yindan Jiedu Granules, a Traditional Chinese Medicinal Formulation, as a Potential Treatment for Coronavirus Disease 2019.

Background: Yindan Jiedu Granules (YDJDG) have been newly prescribed as a Chinese herbal formula. This study aimed to compare the efficacy of YDJDG and lopinavir-ritonavir in the treatment of coronavirus disease 2019 (COVID-19). Methods: Overall, 131 patients with COVID-19 were included in this study. In addition to standard care, 60 of these patients received YDJDG (YDJDG group) and 71 received lopinavir-ritonavir (lopinavir-ritonavir group). Propensity score matching (PSM) was used to match the characteristics of individuals in the two groups, while the Kaplan-Meier method was used to compare the proportion recovery observed. Results: Cox analysis revealed that YDJDG and CD4 ≥ 660 cells/µL were independent predictive factors of proportion recovery. At baseline, disease types differed between the YDJDG and lopinavir-ritonavir treatment groups. Furthermore, no significant adverse effects or toxicities relevant to YDJDG were observed. The median recovery time was 21 days in the YDJDG group and 27 days in the lopinavir-ritonavir group. After PSM (1:1), 50 patient pairs, YDJDG vs. lopinavir-ritonavir, were analyzed. In the YDJDG group, the proportion of recovered patients was remarkably higher than that observed in the lopinavir-ritonavir group (p = 0.0013), especially for those presenting mild/moderate disease type and CD4 < 660 cells/µL. In the YDJDG group, the mean duration of fever and pulmonary exudative lesions was significantly shorter than that observed in the lopinavir-ritonavir group (p = 0.0180 and p = 0.0028, respectively). Conclusion: YDJDG reveals the potential to hasten the recovery period in COVID-19 patients with mild/moderate disease type or CD4 < 660 cells/µL by shortening the mean duration of fever and pulmonary exudative lesions.

Differences and similarities of high-resolution computed tomography features between pneumocystis pneumonia and cytomegalovirus pneumonia in AIDS patients.

BACKGROUND: Accurately differentiating pneumocystis from cytomegalovirus pneumonia is crucial for correct therapy selection in AIDS patients. Hence, the goal of this study was to compare the computerized tomography (CT) features of pneumocystis pneumonia and cytomegalovirus pneumonia in AIDS patients and identify clinical hallmarks to accurately distinguish these two pathologies. METHODS: A total of 112 AIDS patients (78 with pneumocystis pneumonia and 34 cytomegalovirus pneumonia) at Beijing Ditan Hospital from January 2017 to May 2019 were included in this study. Two experienced chest radiologists retrospectively reviewed CT images for 17 features including ground-glass opacity, consolidation, nodules, and halo sign. Binary logistic regression analyses were conducted to identify the significant parameters that distinguished pneumocystis pneumonia from cytomegalovirus pneumonia. Correlations were analyzed by Pearson or Spearman correlation analyses. Result were considered significant if P < 0.05. RESULTS: The presence of consolidation, halo signs, and nodules (all P < 0.05) were significantly more frequent in patients with cytomegalovirus pneumonia than in those with pneumocystis pneumonia. Small nodules (32.5% in cytomegalovirus pneumonia, 6.41% in pneumocystis pneumonia, P < 0.001) without perilymphatic distribution were particularly common in patients with cytomegalovirus pneumonia. Large nodules were not found in any of patients with cytomegalovirus pneumonia. The presence of ground-glass opacity, reticulation, and bronchial wall thickening (all P > 0.05) were common in both groups. CONCLUSIONS: Analysis of consolidation, nodules, and halo signs may contribute to the differential diagnosis of pneumocystis pneumonia or cytomegalovirus pneumonia. However, some CT features considered typical in one or other diseases appear with similar frequency in both cohorts of AIDS patients. CT features are potentially useful for the differential diagnosis of pneumocystis pneumonia and cytomegalovirus pneumonia in AIDS patients.

GLRB variants regulate nearby gene expression in human brain tissues.

A recent genome-wide association study (GWAS) identified four genetic variants rs78726293, rs191260602, rs17035816 and rs7688285 in GLRB gene to be associated with panic disorder (PD) risk. In fact, GWAS is an important first step to investigate the genetics of human complex diseases. In order to translate into opportunities for new diagnostics and therapies, we must identify the genes perturbed by these four variants, and understand how these variant functionally contributes to the underlying disease pathogenesis. Here, we investigated the effect of these four genetic variants and the expression of three nearby genes including PDGFC, GLRB and GRIA2 in human brain tissues using the GTEx (version 6) and Braineac eQTLs datasets. In GTEx (version 6) dataset, the results showed that both rs17035816 and rs7688285 variants could significantly regulate PDGFC and GLRB gene expression. In Braineac dataset, the results showed that rs17035816 variant could significantly regulate GLRB and GRIA2 gene expression. We believe that these findings further provide important supplementary information about the regulating mechanisms of rs17035816 and rs7688285 variants in PD risk.