【Cell】 First time! Stanford University uses organoids to pinpoint the pathways of the new coronavirus in and out of nasal cells

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Written by Jevin

The nose is an important component of upper respiratory mucosal immunity, and it participates in host-protected immune homeostasis
.
There are two main types of nasal mucosal defenses, physiologically speaking, the barrier composed of tightly bound cilia cells, goblet cells and basal epithelial cells, a bilayer of mucus and a basement membrane is the most important means of defense, but it is unclear which types of cells are the first to be infected
.

On January 5, 2023, the Stanford University research team published a research paper online in the prestigious journal Cell, which for the first time detailed the molecular mechanism of respiratory epithelial cells infected by the new coronavirus infection, and the researchers found that the virus attaches to the cilia of epithelial cells through ACE2 receptors, and uses cilia as a channel
to enter the cell.

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Research process

 01 

The researchers used the ALI airway organoid model, which has many nuances
of airways in the body.
Notably, this culture has a significant kinetic delay (24-48 h) compared to tissue culture models, and they established tissue culture models
associated with airway barrier function.

Using EM and IF microscopy, the researchers tracked the detailed steps
required for the virus to enter the airways.
The virus first attaches to airway polycilia via ACE2 receptors, and binding and cilia transport are critical
for the virus to cross the mucus-mucin protective barrier.
Consumption of cilia blocks SARS-Co-2 infection in ALI cultures, but consumption of mucin with purified mucinase accelerates viral entry
.
At 24-48 h, the number of viruses increases, a large number of rearrangements and amplification of microvilli in epithelial cells and activation of PAK1, PAK4 and SLK
kinases.
Inhibitors of these kinases block late viral transmission but cannot bind to cilia initially
.
PAK kinase inhibitors attenuated viral transmission
in mice.

Study results

 02

Even at high titers, only a low percentage of SARS-CoV-2 infects cilia HNE
at 24 hpi.
As a result, the researchers suspect that the virus cannot penetrate the PCL of most cells and requires a previously unidentified gateway
.
Mucus and PCL in the airway epithelium prevent pathogens and particles from entering more than 40 nm
.
Wasting MUC1 partially increases influenza infection, indicating the importance of
PCL as a physical barrier against viruses.

Using a variety of imaging techniques, the researchers found that SARS-CoV-2 efficiently attaches to the distal end
of cilia early in infection (< 6 hours).
Ciliary consumption in nasal epithelial cells inhibits infection without affecting ACE2 and TMPRSS2 levels
.
Thus, cilia can act as ACE2-mediated high-density binding lattices, forming affinity traps that allow viral particles to find target cells by multivalently binding "landing pads", resulting in hyperselectivity
.

Two non-exclusive entry mechanisms are possible, first, SARS-CoV-2 interacts with ACE2 and TMPRSS2 on the cilia surface, triggering membrane fusion into cilia and releasing a viral RNA genome that can be transported from cilia to the cytoplasm
via ciliary dynein.
Alternatively, cilia-attached SARS-CoV-2 can be transported along the cilia surface and bound to ACE2 on the epithelial cell body surface through a cycle of release and rebinding, guiding the virus for TMPRSS2-mediated membrane fusion
.
In this case, the viral ACE2 complex can be transported along the cilia to the apical surface
of epithelial cells via dynein-dependent retrograde flagellar transport.

Research significance

 03 

In addition, the team explored whether cell-to-cell contact affected the transfer
of the virus.
The respiratory epithelium consists of multiple layers of cells, however, until 48 hpi only the uppermost cells are infected by the virus, and the cells in the lower layer are rarely infected
.
Therefore, cell-to-cell contact may not be the only way
for the virus to spread in the nasal epithelium.
Although the virus may be transmitted through cell-to-cell contact in the nasal epithelium, researchers believe that transmission depends on the flow
of mucus on the apical surface.

Overall, studies have found that the new coronavirus will first infect respiratory ciliate cells, and if cilia are removed, it can prevent infection with the new coronavirus and other respiratory viruses, and at the same time, the invading virus activates the kinase in the cell to promote the formation of the cytoskeleton, and sends the newly formed virus to the mucus layer through a highly extended microvilli structure, thereby improving the transmission ability
of the virus.

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Note: This article is intended to introduce the progress of medical research and cannot be used as a reference
for treatment options.
If you need health guidance, please go to a regular hospital
.