William Haseltine

There has been a growing body of work investigating the role of innate lymphoid cells (ILCs) in immunity and host defense. The more we learn about them, the more we recognize that innate lymphoid cells comprise a crucial part of the innate immune response. This includes our ability to contain and suppress SARS-CoV-2 infection.

In our gut, ILCs help prevent SARS-CoV-2 from entering the endothelial lining. They do this by stimulating the production of human defensin-5 (HD-5), which effectively “hides” our endothelial cells’ ACE2 receptors— the primary point of entry for SARS-CoV-2. Beyond the gut, ILCs also impact Covid-19 prognosis. Patients with lower levels of blood ILCs are at a higher risk of progressing to severe disease and requiring hospitalization. This may partly explain why Covid-19 mortality increases with age and is more common in men, both groups displaying naturally decreased levels of blood ILCs.

What are innate lymphoid cells?

Innate lymphoid cells are the latest class of innate immune cells to be discovered. Hints of their existence emerged during the 1970s —by way of research on natural killer (NK) cells and lymphoid tissue inducer (LTi) cells— but a more detailed characterization of the family did not arrive until the early 2010s.1 They are generally found in “barrier environments” —areas of our body that are constantly exposed to the outside world, like our lungs and gastrointestinal tract— where they play an active role in regulating homeostasis. This includes responding to invasive and potentially harmful microorganisms, and helping to repair tissue damaged during infection or injury.

ILCs closely mirror T cells, which are central to the adaptive immune response. They are often considered the innate immune system’s counterpart to T cells. The major difference between the two is that ILCs lack antigen-specific receptors, instead depending on cytokine receptors that allow them to pick up changes in their micro-environment in response to tissue damage. They also have an array of other receptors sensitive to microbial products, neuronal transmitters, and nutrient components.2

The ILC family is made up of three groups: ILC1s, ILC2s and ILC3s (Figure 1). They are classified according to their functions.3 Broadly speaking, ILC1s are in charge of intracellular clearance of pathogens. They are defined by their ability to produce interferon gamma (IFNγ), a small signaling protein that calls macrophages into action and plays a crucial part in both the innate and adaptive immune responses. NK Cells are the prototypical member of this group.

Nomenclature figure 1
Figure 1. Schematic representation of the three groups of innate lymphoid cells (ILCs), including the transcription factors in charge of differentiating the three groups (marked in purple) and the interleukins they help activate (marked in yellow). FROM: “Innate lymphoid cells — a proposal for uniform nomenclature” HERGEN SPITS ET AL. 2013

Group 2 ILCs, on the other hand, are able to produce interleukin-5 (IL-5) and interleukin-13 (IL-13). Interleukins are another kind of signaling molecule, primarily responsible for the growth and differentiation of different immune cells. IL-5 helps regulate the growth of B cells, which go on to produce antibodies. IL-13 contributes to the regulation of the inflammatory response, inhibiting the production of inflammatory cytokines when and where needed.4

Finally, we have group 3 ILCs. These are predominantly involved in gut immunity and are made up of all of the innate lymphoid cells able to produce interleukin-17 and/or interleukin-22. Some ILC3s also activate IFNγ. IL-17 stimulates a variety of different signaling cascades that lead to the production of chemokines— signaling proteins that help move important immune cells, like monocytes and neutrophils, to areas where they are most needed. IL-22 is also closely tied to the inflammatory response, functioning to moderate cell survival and stimulate antimicrobial compounds including defensins and S100 proteins.

Group 3 innate lymphoid cells, SARS-CoV-2, and the gut

One of the many mysteries of SARS-CoV-2 is why it isn’t more of an intestinal virus. If laid out flat, the inner lining of our gastrointestinal tract would cover half a badminton court— roughly 430 square feet.5 Endothelial cells act as the physical barrier of this inner lining, helping to prevent potentially harmful microbes from entering. Their surface is covered in angiotensin-converting enzyme-2 (ACE2) receptors— the primary receptor by which SARS-CoV-2 enters our cells. And yet, despite such an abundance of intestinal epithelial cells and ACE2 receptors, symptoms of the gut are much less frequently reported than respiratory symptoms. As it turns out, this may have something to do with ILC3s.

Paneth cells are a special kind of intestinal epithelial cell. They produce and secrete a host of antimicrobial compounds that help stop invading pathogens, including γ-defensins. IL-22 and IL-13, both products of ILC3s, play an important role in regulating the differentiation of Paneth cells, and by extension, the production of γ-defensins.6

But, do γ-defensins offer any particular protection against SARS-CoV-2 infection? A group of researchers, based at The Army Medical University in Chongqing, China, tackled this question head on.7

Wang et al. zeroed in on human defensin-5, the most abundant γ-defensin secreted by Paneth cells. Through immunofluorescence microscopy, they discovered that HD-5 closely surrounds the ACE2 receptors of our enterocytes, the most common kind of intestinal epithelial cell. Not only this, but HD-5 can actually bind to ACE2. As seen in figure 2, when HD-5 binds ACE2 it “cloaks” part of the receptor known as the ligand-binding domain (LBD).

cloaking figure 2
Figure 2. Schematic showing the cloaking of the ligand-binding domain (pink) of the ACE2 receptor (green) by human defensin-5 (cyan). FROM: “Human Intestinal Defensin 5 Inhibits SARS-CoV-2 Invasion by Cloaking ACE2” WANG ET AL. 2020

Recognizing that the LBD plays an important role in SARS-CoV-2 entry into cells, the researchers were curious to see if this relationship between HD5 and ACE2 receptors had any impact on SARS-CoV-2 infection. To test their hunch, they exposed Caco-2 cells —a type of cell often used to model the intestinal epithelial barrier— to SARS-CoV-2 Spike (S) pseudovirions. When compared to a control group, those Caco-2 cells pre-treated with HD5 for one hour before infection displayed a significant reduction in SARS-CoV-2 invasion.

This was tested three times across three different days, and held true each time. Further, it was also shown to be the case in human renal proximal tubular epithelial cells, a kind of epithelial cell found in a different part of the gut.

Moving forward, it would be interesting to see if supplementation with HD-5 could work therapeutically to reduce the intensity of SARS-CoV-2 infection in the gut. Especially for those patients suffering from chronic inflammatory diseases of the gut which, as Wang et al. note, often run the risk of being HD-5 deficient.

ILCs, disease tolerance, and Covid-19 severity

Researchers with the University of Massachusetts Medical School recently discovered that ILCs may also act as a key factor in determining Covid-19 severity.8

When confronted with an invading pathogen, our body’s immune system kicks in. The first priority is always to contain the infection. This involves all of the processes we usually associate with our immune response, including increased inflammation, the recruitment of immune cells —macrophages, lymphocytes, and so on— and direct confrontation between our immune cells and the microbe in question. All of these are part of “disease resistance”, but there’s an equally important second half to the equation: “disease tolerance”. This is the work our body does to keep us healthy during infection, including the maintenance and repair of tissue. Importantly, the mechanisms underlying disease resistance usually don’t influence pathogen fitness, instead focusing on host health.

In the case of SARS-CoV-2, viral load doesn’t neatly correlate with Covid-19 severity; some people, children in particular, can have very high viral loads while remaining asymptomatic or only mildly symptomatic. To Silverstein et al., this suggested that disease severity in Covid-19 is determined more directly by age-dependent, disease tolerance mechanisms than by viral replication and viral loads.

Prior studies have indicated that ILCs may contribute to disease tolerance, with one specific subset of the ILC family, group 2 innate lymphoid cells (ILC2s), playing an especially active role.9 In response to tissue damage, ILC2s can produce a protein called amphiregulin (AREG). By binding to epidermal growth factor receptors (EGFR), amphiregulin helps stimulate cell growth, cell survival, and cell migration. In doing so, ILC2-induced amphiregulin helps maintain the integrity of the epithelial lining in the lungs and in the intestine.

Spurred on by this, Silverstein et al. measured blood lymphocyte levels through age. Of the lymphocytes measured —CD4+ T Cells, CD4+ B Cells, ILCs, and CD16+ natural killer cells (NK cells)— ILCs were the only subset to show a significant and consistent decrease with age. This age-dependent decrease in ILC levels, a 2-fold drop every twenty years, was very closely mirrored by the age-dependent increase in Covid-19 mortality (Figure 3).

Blood ILC levels figure 3
Figure 3. Blood ILC levels by age (left) next to Covid-19 mortality rate by age (right) FROM: “Innate lymphoid cells and COVID-19 severity in SARS-CoV-2 infection” SILVERSTEIN ET AL. 2022

With the exception of CD4+ T Cells, ILCs were also the only subset to show a marked decrease across sex. In both cases, natural levels were markedly lower in males than in females (Figure 4). Again, this correlates with the fact that males are more likely to suffer from severe Covid-19 than their female counterparts.

Lymphoid-subsets-sex figure 4
Figure 4. Blood ILC levels by sex (left) and blood CD4+ T cell levels by sex (right).

Even when accounting for age, sex and the global lymphopenia often seen during infection, hospitalized Covid-19 patients displayed 1.8-fold fewer blood ILCs compared to the control group (Figure 5). Only CD16+ NK cells showed a similar decrease.

Importantly, as ILCs decreased, both the risk of hospitalization and the duration of hospitalization increased considerably. Every 2-fold decrease in ILCs meant a 55% higher chance of hospitalization and an extra 9.4 days of hospital stay. None of the other lymphocyte subsets influenced rate and duration of hospitalization in the same way.

patient status figure 5
Figure 5. Comparison of ILC and CD16+ NK cell blood levels across control, hospitalized, and outpatient groups. SILVERSTEIN ET AL. 2022

Even when accounting for age, sex and the global lymphopenia often seen during infection, hospitalized Covid-19 patients displayed 1.8-fold fewer blood ILCs compared to the control group (Figure 5). Only CD16+ NK cells showed a similar decrease.

Importantly, as ILCs decreased, both the risk of hospitalization and the duration of hospitalization increased considerably. Every 2-fold decrease in ILCs meant a 55% higher chance of hospitalization and an extra 9.4 days of hospital stay. None of the other lymphocyte subsets influenced rate and duration of hospitalization in the same way.


Innate lymphoid cells (ILCs) are increasingly implicated in a number of crucial immune functions, including in response to SARS-CoV-2 infection. In large part, however, they continue to fly under the radar. Research by the likes of Wang et al. and Silverstein et al. indicates that this might be quite the oversight. Not only do ILCs contribute to the protection of our gut from SARS-CoV-2 infection, but they also play a key role in modulating Covid-19 severity via disease tolerance mechanisms. In the face of yet another wave of infections, knowing which patients will require hospitalization and which will only suffer mild symptoms can be the difference between life and death. More work needs to be done to see how else ILCs may be involved in the defense against, and tolerance of, SARS-CoV-2.


1. Shin, S. B., & McNagny, K. M. (2021). ILC-you in the thymus: A fresh look at innate lymphoid cell development. Frontiers in Immunology, 12.
2. Panda, S. K., & Colonna, M. (2019). Innate lymphoid cells in mucosal immunity.
Frontiers in Immunology, 10.
3. Spits, H., Artis, D., Colonna, M. et al. Innate lymphoid cells — a proposal for uniform nomenclature. Nat Rev Immunol 13, 145–149 (2013).
4. Seyfizadeh, N., Seyfizadeh, N., Gharibi, T., & Babaloo, Z. (2015). Interleukin-13 as an important cytokine: A review on its roles in some human diseases. Acta Microbiologica
Et Immunologica Hungarica, 62(4), 341–378.
5. Helander, H. F., & Fändriks, L. (2014). Surface area of the digestive tract – revisited. Scandinavian Journal of Gastroenterology, 49(6), 681–689.
6. Kamioka, M., Goto, Y., Nakamura, K., Yokoi, Y., Sugimoto, R., Ohira, S., Kurashima, Y., Umemoto, S., Sato, S., Kunisawa, J., Takahashi, Y., Domino, S. E., Renauld, J.-C., Nakae, S., Iwakura, Y., Ernst, P. B., Ayabe, T., & Kiyono, H. (2022). Intestinal commensal microbiota and cytokines regulate fut2+ paneth cells for Gut Defense. Proceedings of the National Academy of Sciences, 119(3).
7. Wang, C., Wang, S., Li, D., Wei, D.-Q., Zhao, J., & Wang, J. (2020). Human intestinal defensin 5 inhibits SARS-COV-2 invasion by cloaking ACE2. Gastroenterology, 159(3).
8. Silverstein, N. J., Wang, Y., Manickas-Hill, Z., Carbone, C., Dauphin, A., Boribong, B. P., Loiselle, M., Davis, J., Leonard, M. M., Kuri-Cervantes, L., Meyer, N. J., Betts, M. R., Li, J. Z., Walker, B. D., Yu, X. G., Yonker, L. M., & Luban, J. (2022). Innate lymphoid
cells and COVID-19 severity in SARS-COV-2 infection. ELife, 11.
9. Branzk, N., Gronke, K., & Diefenbach, A. (2018). Innate lymphoid cells, mediators of tissue homeostasis, adaptation and disease tolerance. Immunological Reviews, 286(1), 86–101.


William R. Haseltine, PhD, is chair and president of the think tank ACCESS Health
International, a former Harvard Medical School and School of Public Health professor and founder of the university’s cancer and HIV/AIDS research departments. He is also the founder of more than a dozen biotechnology companies, including Human Genome Sciences.

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