Distinct Type 1 Immune Networks Underlie the Severity of Restrictive Lung Disease after COVID-19 (preprint)

The variable etiology of persistent breathlessness after COVID-19 have confounded efforts to decipher the immunopathology of lung sequelae. Here, we analyzed hundreds of cellular and molecular features in the context of discrete pulmonary phenotypes to define the systemic immune landscape of post-COVID lung disease. Cluster analysis of lung physiology measures highlighted two phenotypes of restrictive lung disease that differed by their impaired diffusion and severity of fibrosis. Machine learning revealed marked CCR5+CD95+ CD8+ T-cell perturbations in mild-to-moderate lung disease, but attenuated T-cell responses hallmarked by elevated CXCL13 in more severe disease. Distinct sets of cells, mediators, and autoantibodies distinguished each restrictive phenotype, and differed from those of patients without significant lung involvement. These differences were reflected in divergent T-cell-based type 1 networks according to severity of lung disease. Our findings, which provide an immunological basis for active lung injury versus advanced disease after COVID-19, might offer new targets for treatment.

Comparative single-cell analysis reveals IFN-γ as a driver of respiratory sequelae post COVID-19 (preprint)

Post-acute sequelae of SARS-CoV-2 infection (PASC) represents an urgent public health challenge, with its impact resonating in over 60 million individuals globally. While a growing body of evidence suggests that dysregulated immune reactions may be linked with PASC symptoms, most investigations have primarily centered around blood studies, with few focusing on samples derived from post-COVID affected tissues. Further, clinical studies alone often provide correlative insights rather than causal relationships. Thus, it is essential to compare clinical samples with relevant animal models and conduct functional experiments to truly understand the etiology of PASC. In this study, we have made comprehensive comparisons between bronchoalveolar lavage fluid (BAL) single-cell RNA sequencing (scRNAseq) data derived from clinical PASC samples and relevant PASC mouse models. This revealed a strong pro-fibrotic monocyte-derived macrophage response in respiratory PASC (R-PASC) in both humans and mice, and abnormal interactions between pulmonary macrophages and respiratory resident T cells. IFN-g emerged as a key node mediating the immune anomalies in R-PASC. Strikingly, neutralizing IFN-g post the resolution of acute infection reduced lung inflammation, tissue fibrosis, and improved pulmonary gas-exchange function in two mouse models of R-PASC. Our study underscores the importance of performing comparative analysis to understand the root cause of PASC for developing effective therapies.

Proximal immune-epithelial progenitor interactions drive chronic tissue sequelae post COVID-19 (preprint)

The long-term health effects of SARS-CoV-2, termed Post-Acute Sequelae of COVID-19 (PASC), are quickly evolving into a major public health concern, but the underlying cellular and molecular etiology remain poorly defined. There is growing evidence that PASC is linked to abnormal immune responses and/or poor organ recovery post-infection. However, the exact processes linking non-resolving inflammation, impaired tissue repair, and PASC are still unclear. In this report, we utilized a cohort of respiratory PASC patients with viral infection-mediated pulmonary fibrosis and a clinically relevant mouse model of post-viral lung sequelae to investigate the pathophysiology of respiratory PASC. Using a combination of imaging and spatial transcriptomics, we identified dysregulated proximal interactions between immune cells and epithelial progenitors unique to respiratory PASC but not acute COVID-19 or idiopathic pulmonary fibrosis (IPF). Specifically, we found a central role for lung-resident CD8+ T cell-macrophage interactions in maintaining Krt8hi transitional and ectopic Krt5+ basal cell progenitors, and the development of fibrotic sequelae after acute viral pneumonia. Mechanistically, CD8+ T cell derived IFN-{gamma} and TNF stimulated lung macrophages to chronically release IL-1{beta}, resulting in the abnormal accumulation of dysplastic epithelial progenitors in fibrotic areas. Notably, therapeutic neutralization of IFN-{gamma} and TNF, or IL-1{beta} after the resolution of acute infection resulted in markedly improved alveolar regeneration and restoration of pulmonary function. Together, our findings implicate a dysregulated immune-epithelial progenitor niche in driving respiratory PASC and identify potential therapeutic targets to dampen chronic pulmonary sequelae post respiratory viral infections including SARS-CoV-2.

Microfluidics for nano-drug delivery systems: From fundamentals to industrialization

In recent years, owing to the miniaturization of the fluidic environment, microfluidic technology offers unique opportunities for the implementation of nano drug delivery systems (NDDSs) production processes. Compared with traditional methods, microfluidics improves the controllability and uniformity of NDDSs. The fast mixing and laminar flow properties achieved in the microchannels can tune the physicochemical properties of NDDSs, including particle size, distribution and morphology, resulting in narrow particle size distribution and high drug-loading capacity. The success of lipid nanoparticles encapsulated mRNA vaccines against coronavirus disease 2019 by microfluidics also confirmed its feasibility for scaling up the preparation of NDDSs via parallelization or numbering-up. In this review, we provide a comprehensive summary of microfluidics-based NDDSs, including the fundamentals of microfluidics, microfluidic synthesis of NDDSs, and their industrialization. The challenges of microfluidics-based NDDSs in the current status and the prospects for future development are also discussed. We believe that this review will provide good guidance for microfluidics-based NDDSs.

RSCOEWR: Radical-Based Sentiment Classification of Online Education Website Reviews

Online education is becoming more and more popular with the development of the Internet. In particular, due to the COVID-19 pandemic, many countries around the world are increasing the popularity of online education, which makes the research on sentiment classification of course reviews of online education websites an important research direction in natural language processing tasks. Traditional sentiment classification models are mostly based on English. Unlike English, Chinese characters are based on pictograms. Radicals of Chinese characters can also express certain semantics, and characters with the same radical often have similar meanings. Therefore, RSCOEWR, a word-level and radical-level based sentiment classification model for course reviews of Chinese online education websites is proposed, which solves the problem of data sparsity of reviews by feature extraction of multiple dimensions. In addition, a deep learning model based on CNN, BILSTM, BIGRU and Attention is constructed to solve the problem of high dimension and assigning the same attention to context of traditional sentiment classification model. Extensive comparative experiment results show that RSCOEWR outperforms the state-of-the-art sentiment classification models, and the experimental results on public Chinese sentiment classification datasets prove the generalization ability of RSCOEWR.

Reduction in circulating monocytes correlates with persistent post-COVID pulmonary fibrosis in multi-omic comparison of long-haul COVID and IPF (preprint)

Rationale: Up to 30% of COVID-19 patients experience persistent sequelae, including dyspnea, restrictive physiology, and early radiographic signs of pulmonary fibrosis (PF). The mechanisms that provoke post-COVID progressive PF are poorly understood, and biomarkers to identify at-risk patients are urgently needed. Methods: We evaluated a cohort of 14 symptomatic COVID survivors with impaired respiratory function and imaging worrisome for developing PF, including bilateral reticulation, traction bronchiectasis and/or honeycombing, and compared these to Idiopathic Pulmonary Fibrosis (IPF) patients and age-matched controls without respiratory disease. We performed single-cell RNA-sequencing and multiplex immunostaining on peripheral blood mononuclear cells collected at the COVID-19 patients first visit after ICU discharge. Six months later, symptoms, restriction and PF improved in some (Early-Resolving COVID PF), but persisted in others (Late-Resolving COVID PF). Results: Circulating monocytes were significantly reduced in Late-Resolving COVID PF patients compared to Early-Resolving COVID PF and non-diseased controls. Monocyte abundance correlated with pulmonary function tests FVC and DLCO. Differential expression analysis revealed MHC-II class molecules were upregulated on the CD8 T cells of Late-Resolving COVID PF patients but downregulated in monocytes. IPF patients had a similar decrease in monocyte abundance and marked decrease in monocyte HLA-DR protein expression compared to Late-Resolving COVID PF patients. Conclusion: Circulating monocyte abundance may distinguish between patients whose post-COVID PF resolves or persists. Furthermore, fibrotic progression coincided with decreases in HLA-DR expression on monocytes, a phenotype previously associated with dampened antigen stimulation and severe respiratory failure.

Epidemic Dynamics of a Fractional-Order SIR Weighted Network Model and Its Targeted Immunity Control

With outbreaks of epidemics, an enormous loss of life and property has been caused. Based on the influence of disease transmission and information propagation on the transmission characteristics of infectious diseases, in this paper, a fractional-order SIR epidemic model is put forward on a two-layer weighted network. The local stability of the disease-free equilibrium is investigated. Moreover, a conclusion is obtained that there is no endemic equilibrium. Since the elderly and the children have fewer social tiers, a targeted immunity control that is based on age structure is proposed. Finally, an example is presented to demonstrate the effectiveness of the theoretical results. These studies contribute to a more comprehensive understanding of the epidemic transmission mechanism and play a positive guiding role in the prevention and control of some epidemics.

Alveolar regeneration following viral infection is independent of tuft cells (preprint)

Severe injuries following viral infection cause lung epithelial destruction with the presence of ectopic basal progenitor cells (EBCs), although the exact function of EBCs remains controversial. We and others previously showed the presence of ectopic tuft cells in the disrupted alveolar region following severe influenza infection. Here, we further revealed that the ectopic tuft cells are derived from EBCs. This process is amplified by Wnt signaling inhibition but suppressed by Notch inhibition. Further analysis revealed that p63-CreER labeled population de novo arising during regeneration includes alveolar epithelial cells when Tamoxifen was administrated after viral infection. The generation of the p63-CreER labeled alveolar cells is independent of tuft cells, demonstrating segregated differentiation paths of EBCs in lung repair. EBCs and ectopic tuft cells can also be found in the lung parenchyma post SARS-CoV-2 infection, suggesting a similar response to severe injuries in humans.

Response time analysis of a manufacturing supply chain with performance evaluation process algebra

The supply and demand shocks brought on by the Covid-19 crisis are forcing manufacturers everywhere to reassess or even reorganise their supply chains. This paper proposes a general framework that can achieve fast compositional modelling with stochastic performance evaluation process algebra (PEPA) for supply chain systems. The key feature of such a framework is the PEPA-based quantitative analysis approach which includes a new method for calculating response time based on compositional PEPA models. The framework aims to provide an efficient modelling prototype for supply chain systems while supporting accurate response time-based quantitative analysis. To our knowledge, this is the first work using process algebra-based compositional modelling to quantitatively analyse supply chain systems.

Mucosal immunity against SARS-CoV-2 variants of concern including Omicron following vaccination (preprint)

SARS-CoV-2 mRNA vaccination induces robust humoral and cellular immunity in the circulation; however, it is currently unknown whether it elicits effective immune responses in the respiratory tract, particularly against variants of concern (VOCs), including Omicron. We compared the SARS-CoV-2 S-specific total and neutralizing antibody (Ab) responses, and B and T cell immunity, in the bronchoalveolar lavage fluid (BAL) and blood of COVID-19 vaccinated individuals and hospitalized patients. Vaccinated individuals had significantly lower levels of neutralizing Ab against D614G, Delta and Omicron in the BAL compared to COVID-19 convalescents, despite robust S-specific Ab responses in the blood. Further, mRNA vaccination induced significant circulating S-specific B and T cell immunity, but in contrast to COVID-19 convalescents, these responses were absent in the BAL of vaccinated individuals. Using an animal immunization model, we demonstrate that systemic mRNA vaccination alone induced weak respiratory mucosal neutralizing Ab responses, especially against SARS-CoV-2 Omicron; however, a combination of systemic mRNA vaccination plus mucosal adenovirus-S immunization induced strong neutralizing Ab response, not only against the ancestral virus but also the Omicron variant. Together, our study supports the contention that the current COVID-19 vaccines are highly effective against severe disease development, likely through recruiting circulating B and T cell responses during re-infection, but offer limited protection against breakthrough infection, especially by Omicron. Hence, mucosal booster vaccination is needed to establish robust sterilizing immunity in the respiratory tract against SARS-CoV-2, including infection by Omicron and future variants.

Combinatorial mRNA vaccination enhances protection against SARS-CoV-2 delta variant (preprint)

Emergence of SARS-CoV-2 variants of concern (VOC), including the highly transmissible delta strain, has posed challenges to current COVID-19 vaccines that principally target the viral spike protein (S). Here, we report a nucleoside-modified mRNA vaccine that expresses the more conserved viral nucleoprotein (mRNA-N). We show that mRNA-N alone was able to induce a modest but significant control of SARS-CoV-2 in mice and hamsters. Critically, by combining mRNA-N with the clinically approved S-expressing mRNA vaccine (mRNA-S-2P), we found that combinatorial mRNA vaccination (mRNA-S+N) led to markedly enhanced protection against the SARS-CoV-2 delta variant compared to mRNA-S. In a hamster model, we demonstrated that while mRNA-S alone elicited significant control of the delta strain in the lungs (~45-fold reduction in viral loads compared to un-vaccinated control), its effectiveness in the upper respiratory tract was weak, whereas combinatorial mRNA-S+N vaccination induced markedly more robust control of the delta variant infection in the lungs (~450-fold reduction) as well as in the upper respiratory tract (~20-fold reduction). Immune analyses indicated that induction of N-specific immunity as well as augmented S-specific T-cell response and neutralizing antibody activity were collectively associated the enhanced protection against SARS-CoV-2 delta strain by combinatorial mRNA vaccination. These findings suggest that the combined effects of protection in the lungs and upper respiratory tract could both reduce the risk of severe disease as well as of infection and transmission.

Milk Small Extracellular Vesicles-derived miRNAs Restrict Porcine Epidemic Diarrhea Virus Infectivity (preprint)

Background: Virus-caused diseases are a huge challenge to both animals and human beings, especially coronaviruses. Porcine epidemic diarrhea virus (PEDV), a coronavirus, causes acute diarrhea and up to 100% mortality in piglets less than three weeks of age. Maternal immunity provides protection for piglets in resisting PEDV infection. Small extracellular vesicles (sEV) contain bioactive molecules such as miRNAs to exchange genetic and epigenetic information between cells. Our previous study suggested that milk sEV facilitated intestinal tract development and prevented LPS-induced intestine damage. However, the effects of milk sEV on the inhibition of viral infections remain unclear. Results: In this study, through in vivo experiments, we found that porcine milk sEV protected piglets from PEDV-induced diarrhea and death. In vitro, we clarified that this protective effect was partly generated through the inhibition of the PEDV-N protein and HMGB1 by sEV miR-let-7e and miR-27b, respectively. Conclusions: In conclusion, we report that porcine milk sEVs protected piglets from PEDV-induced diarrhea and death by inhibiting virus replication, and this protective effect was partly generated through the inhibition of the PEDV-N and HMGB1 pathways by exosomal miR-let-7e and miR-27b. This study reveals a new antiviral function of milk sEVs, and the results suggest that milk sEVs may act as a mother-offspring transmission pathway for protecting newborns against PEDV infection.

Tissue-resident CD8+T cells drive age-associated chronic lung sequelae following viral pneumonia (preprint)

Lower respiratory viral infections, such as influenza virus and severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infections, often cause severe viral pneumonia in aged individuals. Here, we report that influenza viral pneumonia leads to chronic non-resolving lung pathology and exaggerated accumulation of CD8 + tissue-resident memory T cells (T RM ) in the respiratory tract of aged hosts. T RM accumulation relies on elevated TGF-β present in aged tissues. Further, we show that T RM isolated from aged lungs lack a subpopulation characterized by expression of molecules involved in TCR signaling and effector function. Consequently, T RM cells from aged lungs were insufficient to provide heterologous protective immunity. Strikingly, the depletion of CD8 + T RM cells dampens persistent chronic lung inflammation and ameliorates tissue fibrosis in aged, but not young, animals. Collectively, our data demonstrate that age-associated T RM cell malfunction supports chronic lung inflammatory and fibrotic sequelae following viral pneumonia in aged hosts.