Increased TMPRSS2 expression in tongue tissue of females and alcohol drinkers identified as a potential risk factor for SARS-CoV-2 infection

By Dr. Chinta SidharthanOct 17 2022Reviewed by Danielle Ellis, B.Sc.

In a recent study published in the Journal of Clinical Biochemistry and Nutrition, researchers explored the association between the expressions of angiotensin converting enzyme-2 (ACE-2) receptor and transmembrane serine protease 2 (TMPRSS2) in the tongue tissue and dysgeusia during severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections.

Background

Studies on coronavirus disease 2019 (COVID-19) have revealed that ACE-2 receptors and serine proteases play an important role in the entry of SARS-CoV-2 into host cells. The SARS-CoV-2 spike protein binds to the ACE-2 receptors, and further cleavage of the spike protein by serine proteases enables the entry of viral ribonucleic acid (RNA) into the host cell cytosol. 

One commonly reported symptom of COVID-19 is dysgeusia or the loss of taste. Recent research on oral mucosa has identified increased expression of ACE-2 receptors and TMPRSS2 in the tongue epithelia and saliva. However, the association between ACE-2 and serine protease expression and loss of taste during SARS-CoV-2 infections remains to be examined.

About the study

In the present study, the researchers used immunoblot analysis and immunohistochemistry techniques to determine ACE-2 and TMPRSS2 expression in the human and murine tongue tissue.

Tongue, submandibular glands, and kidneys were acquired from euthanized five-week-old male mice. Half the tissues were fixed in phosphate-buffered formalin for immunohistochemical and histological analysis, while the other half was frozen at -80 °C for the immunoblot assay.

With informed consent, human tongue samples were acquired from patients with primary tongue cancer who underwent surgery. The non-tumorous regions of the tongue were fixed in formalin and embedded in paraffin for the immunohistochemical tests. Three fresh human tongue samples were used for the immunoblot analysis.

Paraffin-fixed tissues were sectioned and mounted on glass slides. Immunostaining was performed using ACE-2 and TMPRSS2 antibodies. Tissues were homogenized and lysed using a lysis buffer after sonication, for the immunoblot analysis. Protein concentration was determined using a protein assay kit and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The immunoblots were incubated with ACE-2 and TMPRSS2 secondary antibodies conjugated with horseradish-peroxidase.

Results

The results reported a higher expression of ACE-2 receptors in the stratified squamous epithelia, perineurium, and arterial wall of murine tongue tissue and luminal side of the salivary glands compared to similar tissues from human tongue samples. However, the expression of TMPRSS2 was higher in the stratified squamous epithelia of human tongue tissue, salivary glands, taste buds, and perineurium than in murine tissue.

The immunoblot results and immunohistochemistry results were similar. Logistic regression analysis was used to analyze the immunohistochemistry data according to sex, age, smoking behavior, and alcohol intake. The findings showed that while there was no significant difference in ACE-2 receptor expression in human tongue tissue according to age, sex, alcohol intake, or smoking, TMPRSS2 expression in the tongue was significantly upregulated in females and alcohol drinkers. Smoking habit was associated with a potential increase in TMPRSS2 expression.

Additionally, the evaluation of ACE-2 and TMPRSS2 expression in other human organs revealed that ACE-2 expression is high in the kidneys, testes, and small intestine, whereas TMPRSS2 expression is upregulated in the breast tissue, liver, kidneys, and prostate gland.

According to the authors, the lower expression of ACE-2 and higher TMPRSS2 expression in human tongue tissue as compared to murine tongue tissue indicated that susceptibility to SARS-CoV-2 infections is associated more with the balance between ACE-2 and TMPRSS2 expression and not directly with the level of ACE-2 expression. Studies have reported that mice are less susceptible to COVID-19 than other animals, which could be linked to the low TMPRSS2 expression in their tongue tissue.

Furthermore, the study found that the perineurium and taste buds of the human tongue expressed higher levels of ACE-2 and TMPRSS2 than other regions, which could provide a link to the dysgeusia experienced by many COVID-19 patients.

Conclusions

To summarize, the study examined the levels of ACE-2 receptor and TMPRSS2 in the tongue tissue of humans and mice and found that murine tongue tissue has higher levels of ACE-2 receptors and lower TMPRSS2 expression than human tongue tissue.

Alcohol intake and female sex were linked to higher TMPRSS2 expression in the tongue, as was smoking behavior, albeit only potentially. The lower susceptibility of mice to SARS-CoV-2 infections suggested that ACE-2 levels alone do not determine the susceptibility, and TMPRSS2 levels play a role in increased COVID-19 risk. The authors believe that the possibility of SARS-CoV-2 infections could be low if TMPRSS2 expressions were low, even with high ACE-2 receptor levels.

Lastly, the study also indicated an association between SARS-CoV-2 entry sites in the tongue tissue and the loss of taste in COVID-19 patients, with the taste buds and tongue perineurium exhibiting higher TMPRSS2 levels than other tissues.