Lower respiratory tract infections are one of the leading causes of death worldwide. Most such infections, including COVID-19, begin harmlessly in the upper respiratory tract and only become dangerous when they reach the alveoli, where gas exchange occurs. Until recently, it was difficult to determine when and where the critical transition to life-threatening pneumonia and acute respiratory distress syndrome (ARDS) occurs. However, the widespread use of multi-omics profiling has provided a detailed picture of COVID-19 pathogenesis and how specific cells and molecules contribute to disease progression.
Multi-omics Profiling Sheds Light on COVID-19 Pathogenesis
Single-cell multi-omics profiling is a method that allows the simultaneous study of multiple parameters of a single cell, including gene activity, protein synthesis, DNA changes, and more. Using this technique, scientists have been able to create models of COVID-19 pathogenesis that involve not only alveolar epithelial infection but also the engagement of alveolar capillaries, macrophages, and other myeloid cells, along with the production of various inflammatory cytokines and chemokines.
Stanford University researchers discovered that two groups of pulmonary macrophages are the main targets of the coronavirus. In one of these groups, the virus alters gene activity. It triggers an antiviral program, including an interferon response and the production of chemokines and cytokines, which can impact different types of immune and structural lung cells, leading to inflammation and fibrosis. This group of lung macrophages contributes to the transition to lethal and systemic forms of COVID-19.
Specific Lung Cells Express Thousands of Viral RNA Molecules
The researchers used lung samples from surgeries or donors and exposed them to SARS-CoV-2 in laboratory conditions. Over 24-72 hours, the virus replicated actively.
Most cells in the infected samples contained few viral RNA molecules. However, 1% of the cells expressed dozens to hundreds of viral RNA molecules per cell, with rare cells (0.01%) accumulating thousands of viral RNAs.
The highest viral load was detected in six types of cells. AT2 cells — type 2 alveolar epithelial cells, myofibroblasts, lipofibroblasts, T-cells, NK cells, and macrophages. Macrophages were the most prominent targets of the coronavirus in the lungs, accounting for 75% of the cells with 50 or more viral RNA molecules per cell. However, these macrophages represented only 0.5% of all macrophages, indicating the existence of a specific, coronavirus-sensitive macrophage subtype.
Activated Interstitial Macrophages Are the Most Susceptible to the Coronavirus
The researchers identified three groups of macrophages:
- Alveolar Macrophages (AM): These maintain normal functions related to lipid homeostasis and immune defense. AMs are located in the alveolar airspace.
- Interstitial Macrophages (IM): These cells express lower levels of typical AM markers and are located in the interstitium, the supportive tissue surrounding the alveoli.
- Activated Interstitial Macrophages (a-IM): The most susceptible to the coronavirus, these a-IM cells express genes related to inflammation and hypoxia. Only a-IMs contain over 300 viral RNA molecules per cell, leading to the hijacking of cellular resources to produce new viral particles. In a-IMs, viral RNA comprised 60% of the total genetic material. By contrast, viral RNA in AMs did not exceed 2%.
a-IM Macrophages: The Focal Point of Inflammation and Fibrosis in COVID-19
In a-IM macrophages, the accumulation of viral RNA triggers the activation of antiviral genes, including those involved in the type I interferon response. Additionally, viral RNA accumulation in a-IMs was associated with the production of five chemokines (CCL2, CCL7, CCL8, CCL13, and CXCL10) and the cytokines IL10 and IL6, which play vital roles in the cytokine storm observed in severe COVID-19 cases. TGFB1, a protein whose excessive activity can lead to fibrosis, was also overproduced, and fibrosis-promoting genes were activated.
The production of the chemokine CXCL10 in a-IMs may attract CD4 and CD8 T-cells, while the chemokine CCL8 can attract neutrophils and other macrophages. CCL2 attracts dendritic cells and monocytes. The production of the cytokine IL6 suggests that infected a-IMs may send strong pro-inflammatory signals to most other lung cell types, while the production of the anti-inflammatory cytokine IL10 may limit adaptive immune activation while enhancing innate inflammation.
Coronavirus Uses DC-SIGN/CD209 Protein to Enter IMs, Not ACE2
DC-SIGN/CD209 is a protein expressed only in IMs. Blocking antibodies against DC-SIGN/CD209 suppressed viral entry into IMs, while blocking antibodies against ACE2 did not affect viral entry into these cells.
Conclusion
Alveolar and interstitial macrophages respond differently to the coronavirus. Interstitial macrophages are more vulnerable, with the virus actively replicating in these cells, hijacking their resources for reproduction and causing severe inflammation and tissue damage. Up to 60% of the genetic material in infected interstitial macrophages consists of viral RNA, leading to cell destruction and the formation of new viral particles. Infected interstitial macrophages drive the inflammation and fibrosis characteristic of severe COVID-19. Treatments targeting interstitial macrophages and the DC-SIGN/CD209 protein, which the coronavirus uses to enter these cells, may prevent severe complications.
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