Postmortem high-dimensional immune profiling of severe COVID-19 patients reveals distinct patterns of immunosuppression and immunoactivation.

A complete diagnostic autopsy is the gold-standard to gain insight into Coronavirus disease 2019 (COVID-19) pathogenesis. To delineate the in situ immune responses to SARS-CoV-2 viral infection, here we perform comprehensive high-dimensional transcriptional and spatial immune profiling in 22 COVID-19 decedents from Wuhan, China. We find TIM-3-mediated and PD-1-mediated immunosuppression as a hallmark of severe COVID-19, particularly in men, with PD-1+ cells being proximal rather than distal to TIM-3+ cells. Concurrently, lymphocytes are distal, while activated myeloid cells are proximal, to SARS-CoV-2 viral antigens, consistent with prevalent SARS-CoV-2 infection of myeloid cells in multiple organs. Finally, viral load positively correlates with specific immunosuppression and dendritic cell markers. In summary, our data show that SARS-CoV-2 viral infection induces lymphocyte suppression yet myeloid activation in severe COVID-19, so these two cell types likely have distinct functions in severe COVID-19 disease progression, and should be targeted differently for therapy.

A single-cell transcriptomic landscape of the lungs of patients with COVID-19.

The lung is the primary organ targeted by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), making respiratory failure a leading coronavirus disease 2019 (COVID-19)-related mortality. However, our cellular and molecular understanding of how SARS-CoV-2 infection drives lung pathology is limited. Here we constructed multi-omics and single-nucleus transcriptomic atlases of the lungs of patients with COVID-19, which integrate histological, transcriptomic and proteomic analyses. Our work reveals the molecular basis of pathological hallmarks associated with SARS-CoV-2 infection in different lung and infiltrating immune cell populations. We report molecular fingerprints of hyperinflammation, alveolar epithelial cell exhaustion, vascular changes and fibrosis, and identify parenchymal lung senescence as a molecular state of COVID-19 pathology. Moreover, our data suggest that FOXO3A suppression is a potential mechanism underlying the fibroblast-to-myofibroblast transition associated with COVID-19 pulmonary fibrosis. Our work depicts a comprehensive cellular and molecular atlas of the lungs of patients with COVID-19 and provides insights into SARS-CoV-2-related pulmonary injury, facilitating the identification of biomarkers and development of symptomatic treatments.

A cohort autopsy study defines COVID-19 systemic pathogenesis.

Severe COVID-19 disease caused by SARS-CoV-2 is frequently accompanied by dysfunction of the lungs and extrapulmonary organs. However, the organotropism of SARS-CoV-2 and the port of virus entry for systemic dissemination remain largely unknown. We profiled 26 COVID-19 autopsy cases from four cohorts in Wuhan, China, and determined the systemic distribution of SARS-CoV-2. SARS-CoV-2 was detected in the lungs and multiple extrapulmonary organs of critically ill COVID-19 patients up to 67 days after symptom onset. Based on organotropism and pathological features of the patients, COVID-19 was divided into viral intrapulmonary and systemic subtypes. In patients with systemic viral distribution, SARS-CoV-2 was detected in monocytes, macrophages, and vascular endothelia at blood-air barrier, blood-testis barrier, and filtration barrier. Critically ill patients with long disease duration showed decreased pulmonary cell proliferation, reduced viral RNA, and marked fibrosis in the lungs. Permanent SARS-CoV-2 presence and tissue injuries in the lungs and extrapulmonary organs suggest direct viral invasion as a mechanism of pathogenicity in critically ill patients. SARS-CoV-2 may hijack monocytes, macrophages, and vascular endothelia at physiological barriers as the ports of entry for systemic dissemination. Our study thus delineates systemic pathological features of SARS-CoV-2 infection, which sheds light on the development of novel COVID-19 treatment.

Public-Health-Driven Microfluidic Technologies: From Separation to Detection.

Separation and detection are ubiquitous in our daily life and they are two of the most important steps toward practical biomedical diagnostics and industrial applications. A deep understanding of working principles and examples of separation and detection enables a plethora of applications from blood test and air/water quality monitoring to food safety and biosecurity; none of which are irrelevant to public health. Microfluidics can separate and detect various particles/aerosols as well as cells/viruses in a cost-effective and easy-to-operate manner. There are a number of papers reviewing microfluidic separation and detection, but to the best of our knowledge, the two topics are normally reviewed separately. In fact, these two themes are closely related with each other from the perspectives of public health: understanding separation or sorting technique will lead to the development of new detection methods, thereby providing new paths to guide the separation routes. Therefore, the purpose of this review paper is two-fold: reporting the latest developments in the application of microfluidics for separation and outlining the emerging research in microfluidic detection. The dominating microfluidics-based passive separation methods and detection methods are discussed, along with the future perspectives and challenges being discussed. Our work inspires novel development of separation and detection methods for the benefits of public health.

Segmenting lung lesions of COVID-19 from CT images via pyramid pooling improved Unet.

Segmenting lesion regions of Coronavirus Disease 2019 (COVID-19) from computed tomography (CT) images is a challenge owing to COVID-19 lesions characterized by high variation, low contrast between infection lesions and around normal tissues, and blurred boundaries of infections. Moreover, a shortage of available CT dataset hinders deep learning techniques applying to tackling COVID-19. To address these issues, we propose a deep learning-based approach known as PPM-Unet to segmenting COVID-19 lesions from CT images. Our method improves an Unet by adopting pyramid pooling modules instead of the conventional skip connection and then enhances the representation of the neural network by aiding the global attention mechanism. We first pre-train PPM-Unet on COVID-19 dataset of pseudo labels containing1600 samples producing a coarse model. Then we fine-tune the coarse PPM-Unet on the standard COVID-19 dataset consisting of 100 pairs of samples to achieve a fine PPM-Unet. Qualitative and quantitative results demonstrate that our method can accurately segment COVID-19 infection regions from CT images, and achieve higher performance than other state-of-the-art segmentation models in this study. It offers a promising tool to lay a foundation for quantitatively detecting COVID-19 lesions.

Autopsy and Histologic Findings of Patients with New Coronavirus Pneumonia: The Pathologic Associations with Hypoxemia.

BACKGROUND Coronavirus 2 (SARS-CoV-2) was declared a pandemic by the World Health Organization (WHO) in March 2020. To further reveal the pathologic associations between coronavirus and hypoxemia, we report the findings of 4 complete systematic autopsies of severe acute respiratory syndrome coronavirus 2-positive individuals who died of multiple organ failure caused by severe hypoxemia. MATERIAL AND METHODS We examined the donated corpses of 4 deceased patients who had been diagnosed with severe acute respiratory syndrome coronavirus 2. A complete post-mortem examination was carried out on each corpse, and multiple organs were macroscopically examined. RESULTS The 4 corpses were 2 males and 2 females, with an average age of 69 years. Bilateral lungs showed various degrees of atrophy and consolidation, with diffusely tough and solid texture in the sections. A thromboembolism was found in the main pulmonary artery extending into the atrium in 1 corpse, and significant atherosclerotic plaques tagged in the inner wall of the aortic arch were found in 2 corpses. Two corpses were found to have slightly atrophied bilateral renal parenchyma. Atrophic changes in the spleen were found in 2 corpses. Notably, there were significantly expanded alveolar septa and prominent fibroblastic proliferation. CONCLUSIONS The laboratory data of these corpses showed a progressive decrease in blood oxygen saturation, followed by refractory and irreversible hypoxemia. Clinical and laboratory information and autopsy and histologic presentations of multiple organs showed insufficient air exchange due to abnormalities in the respiratory system, and reduced erythropoiesis in bone marrow may play a role.

Retracted: Autopsy and Histologic Findings of Patients with New Coronavirus Pneumonia: The Pathologic Associations with Hypoxemia.

This manuscript has been retracted due to the identification of undeclared duplication of content, including Figure images, from a -previous publication by some of the authors: Wang C, Xie J, Zhao L, Fei X, Zhang H, Tan Y, Nie X, Zhou L, Liu Z, Ren Y, Yuan L, Zhang Y, Zhang J, Liang L, Chen X, Liu X, Wang P, Han X, Weng X, Chen Y, Yu T, Zhang X, Cai J, Chen R, Shi ZL, Bian XW. Alveolar macrophage dysfunction and cytokine storm in the pathogenesis of two severe COVID-19 patients. EBioMedicine. 2020; 57: 102833. All authors are requested to declare that manuscripts submitted to this journal are original. This journal makes clear that research fraud of any kind will not be tolerated and will result in immediate retraction.

Pathological changes in the lungs and lymphatic organs of 12 COVID-19 autopsy cases.

Systematic autopsy and comprehensive pathological analyses of COVID-19 decedents should provide insights into the disease characteristics and facilitate the development of novel therapeutics. In this study, we report the autopsy findings from the lungs and lymphatic organs of 12 COVID-19 decedents-findings that evaluated histopathological changes, immune cell signature and inflammatory factor expression in the lungs, spleen and lymph nodes. Here we show that the major pulmonary alterations included diffuse alveolar damage, interstitial fibrosis and exudative inflammation featured with extensive serous and fibrin exudates, macrophage infiltration and abundant production of inflammatory factors (IL-6, IP-10, TNFα and IL-1ß). The spleen and hilar lymph nodes contained lesions with tissue structure disruption and immune cell dysregulation, including lymphopenia and macrophage accumulation. These findings provide pathological evidence that links injuries of the lungs and lymphatic organs with the fatal systematic respiratory and immune malfunction in critically ill COVID-19 patients.

Alveolar macrophage dysfunction and cytokine storm in the pathogenesis of two severe COVID-19 patients.

BACKGROUND: The novel coronavirus pneumonia COVID-19 caused by SARS-CoV-2 infection could lead to a series of clinical symptoms and severe illnesses, including acute respiratory distress syndrome (ARDS) and fatal organ failure. We report the fundamental pathological investigation in the lungs and other organs of fatal cases for the mechanistic understanding of severe COVID-19 and the development of specific therapy in these cases. METHODS: The autopsy and pathological investigations of specimens were performed on bodies of two deceased cases with COVID-19. Gross anatomy and histological investigation by Hematoxylin and eosin (HE) stained were reviewed on each patient. Alcian blue/periodic acid-Schiff (AB-PAS) staining and Masson staining were performed for the examinations of mucus, fibrin and collagen fiber in lung tissues. Immunohistochemical staining was performed on the slides of lung tissues from two patients. Real-time PCR was performed to detect the infection of SARS-CoV-2. Flow cytometry analyses were performed to detect the direct binding of S protein and the expression of ACE2 on the cell surface of macrophages. FINDINGS: The main pathological features in lungs included extensive impairment of type I alveolar epithelial cells and atypical hyperplasia of type II alveolar cells, with formation of hyaline membrane, focal hemorrhage, exudation and pulmonary edema, and pulmonary consolidation. The mucous plug with fibrinous exudate in the alveoli and the dysfunction of alveolar macrophages were characteristic abnormalities. The type II alveolar epithelial cells and macrophages in alveoli and pulmonary hilum lymphoid tissue were infected by SARS-CoV-2. S protein of SARS-CoV-2 directly bound to the macrophage via the S-protein-ACE2 interaction. INTERPRETATION: Infection of alveolar macrophage by SARS-CoV-2 might be drivers of the "cytokine storm", which might result in damages in pulmonary tissues, heart and lung, and lead to the failure of multiple organs . FUNDING: Shanghai Guangci Translational Medical Research Development Foundation, Shanghai, China.