Pathology of lung-specific thrombosis and inflammation in COVID-19

Abstract

Background: Infection by SARS-CoV-2 produces significant pulmonary pathology including endothelial damage with resultant thrombotic events. While pathologic features were described, there are limited data on the relationship of these changes to the inflammatory response and the production of thromboses.

Objective: To investigate pathology of COVID-19-related immunothrombosis.

Patients/methods: Tissue samples from lung, kidney, brain and heart that were collected from 45 patients who died of COVID-19. Histopathological examination was performed after H&E and Picro-Mallory staining in combination with (immuno)fluorescence to visualize neutrophil extracellular traps. Ultrastructural alterations in lungs were studied with scanning and transmission electron microscopy.

Results: Inflammatory changes and thrombosis were substantially more pronounced in the lung than in the kidney, heart, and brain. The most common pathologic finding was diffuse alveolar damage. In addition, most lung samples showed thrombi in vessels. The cause of death in single cases was massive pulmonary embolism. Ultrastructural examination revealed neutrophils attached to endothelium, perhaps as a step towards transendothelial migration. In addition, platelets were identified in the midst of fibrin as individual procoagulant balloon-like cells. Ultrastructural examination demonstrated numerous virion-like particles.

Conclusions: Studying (ultra)structural features of the autopsy lung samples from patients with COVID-19 has provided evidence for a pathogenic link between inflammation and thrombosis. The major features in the lungs of COVID-19 patients comprised primary inflammatory thrombosis associated with diffuse alveolar damage. The lungs had pronounced circulatory changes with inflammation-dependent intravascular blood clotting, whereas heart, brain, and kidneys had predominantly degenerative changes that were distinct from the lung pathology.

Keywords: COVID-19; blood coagulation; inflammation; lungs; thrombosis.

FIGURE 1

Representative histological images showing characteristic structural alterations in the autopsy lung tissue samples from patients who died of COVID‐19. (A) Multiple microthrombi (M) consisting mainly of erythrocytes; focal pneumofibrosis and perivascular fibrosis (P); and leukocyte infiltration (L). Picro‐Mallory stain, magnification bar =100 μm. (B) Thrombi (T) attached to the vessel walls and containing predominantly fibrin and leukocytes. H&E, magnification bar =200 μm. (C) Internal structure of a pulmonary thrombotic embolus with alternation of the blue (old) fibrin (F) and erythrocyte (Er) layers. Picro‐Mallory stain, magnification bar =100 μm. (D) Alveoli with microthrombi (M), diapedesis of single erythrocytes (Er), and hyaline membranes (H). H&E, magnification bar =50 μm. (E) Infiltration of lung tissue with neutrophils (L), focal edema (E) and hemorrhage (H). H&E, magnification bar =100 μm. (F) Blood vessels containing microthrombi stained for histones CitH3 (green) and DNA (blue) to visualize neutrophils and NETs. The surrounding extravascular cells are DAPI‐positive (blue) due to the presence of DNA. Fluorescence, magnification bar =100 μm

FIGURE 2

Representative electron microscopy images showing characteristic structural alterations in the lung tissue samples from patients who died of COVID‐19. (A) A scanning electron micrograph of compacted non‐porous depositions of fibrin, dead cells and surfactant (dotted ovals) attached to the alveolar septa. Magnification bar =25 μm. (B) A scanning electron microscopy image of a spongy fibrous fibrin‐like structure (dotted oval) in the lumen of alveoli, not attached or tightly adherent to the alveolar septa. Magnification bar =5 μm. (C) A scanning electron micrograph showing the lumen of a dilated blood vessel (within the dotted circle) containing sparse and compacted blood cells, mainly erythrocytes (Er). Magnification bar =50 μm. (D) Transmission electron microscopy of capillary lumens with irregularly shaped (compressed) erythrocytes (Er) and leukocytes (L) inside. Destroyed vessel walls as well as the surrounding cells and tissues are shown (dotted oval). Collagen fibers and edematous tissue are seen behind the basement membrane. Magnification bar =5 μm. (E) Scanning electron microscopy showing individual balloon‐like platelets (within the dotted oval) among fibrin fibers. Magnification bar =5 μm. (F) Echinocytes (within the dotted ovals) in the extravascular space. Magnification bar =15 μm

FIGURE 3

Representative high‐resolution transmission electron microscopy images of coronavirus‐like particles in the lung tissue of a patient who died of COVID‐19. (A) Small (lined oval) and large (below) membrane‐bound vacuoles containing particles within the cytoplasm of a type II pneumocyte. (B) A cluster of extracellular virion‐like particles (lined oval) in the alveolar space. (C) An extracellular particle with distinctive spike‐like projections (shown by arrows). Magnification bars =200 nm

FIGURE 4

Representative scanning electron microscopy images of a pulmonary blood clot from a patient who died of COVID‐19. (A) Compacted polyhedrocytes and a small amount of free barbed fibrin fiber ends (E). Magnification bar =5 μm. (B) Fibrin bundles (dotted ovals) and fibers in the clot. Magnification bar =10 μm. (C) The peripheral part of the clot containing a large amount of fibrin. Magnification bar =10 μm. (D) Fibrin bundles coated with sparse spherical microparticles. Magnification bar =1 μm

FIGURE 5

Representative histological images, showing characteristic structural alterations of the autopsy samples of heart (A, B), brain (C) and kidney (D) from patients who died of COVID‐19. (A) A heart cavity with a thrombus composed mainly of fibrin (F) and neutrophilic leukocytes (L). H&E, magnification bar =100 μm. (B) Microthrombi (M) in a vascular lumen; extensive sclerosis of the coronary arteries and cardiosclerosis (S), degeneration of cardiomyocytes. H&E, magnification bar =50 μm. (C) A microscopic thrombus (M) composed mainly of erythrocytes; pericellular and perivascular edema of brain tissue (E). H&E, magnification bar =100 μm. (D) Acute tubular necrosis appearing as the absence of nuclei in the cells of tubules (N); parenchymatous focal degeneration of tubular epithelium (D); glomerulosclerosis (S); and fibrin deposits (F) in a Bowman's capsule. Picro‐Mallory stain, magnification bar =100 μm