Role of IL-27 in COVID-19: A Thin Line Between Protection and Disease Promotion

1. Introduction

The COVID-19 pandemic has caused a global health crisis with millions of confirmed cases and deaths worldwide [1]. COVID-19 is caused by the SARS-CoV-2 virus, which enters the body through the respiratory system and attaches to ACE2 (angiotensin converting enzyme II), widely present on cells in different parts of the body: the respiratory system, intestines, blood vessels, heart tissue, thyroid gland, epididymis, pancreas, liver, and even placenta [2]. Upon entry, the virus initiates immune reactions, which can cause inflammatory responses to become hyperergic. This hyperinflammation has the potential to cause a cytokine storm and widespread inflammation throughout the body, resulting in tissue damage to various organs such as the respiratory tract, heart, kidneys, and nervous system [3]. Cytokine storm is usually described as one of the main reasons behind COVID-associated mortality [4].

The virus primarily affects the respiratory system; therefore, symptoms of infection vary in their severity from the common cold to pneumonia with severe acute respiratory syndrome [5]. However, the pathophysiology behind COVID-19 is rather complex; it involves multiple organ systems, including the immune system.

Cytokines are essential protein molecules engaged in immune responses; they play a critical role in protection against infections. However, they also contribute to inflammatory reactions and tissue damage, becoming a double-edged sword in the context of COVID-19. Recent studies have suggested various cytokines and chemokines that play a crucial role in the immune response to SARS-CoV-2 infection. One such cytokine is Interleukin 27 (IL-27), which has been found to be elevated in the blood plasma of patients with COVID-19.

Within this study, we will explore the role of IL-27 in immune responses and address our own findings concerning this cytokine in COVID-19.

2. Interleukin 27 Conformation and Its Biological Function in Immunity

IL-27 consists of two subunits, IL-27p28 and EBI3, that combine to form a heterodimer. IL-27p28 (also known as IL-30) was initially discovered as an orphan homologue of the IL-6/12 family in 2002 through a computational analysis of expressed sequence tags [6]. In contrast, EBI3 was first identified in 1996 as an erythropoietin receptor-like protein induced by Epstein-Barr virus infection in B cells [7]. Antigen-presenting cells typically express both subunits of IL-27, and the secretion of human p28 is reliant on the secretion of human EBI3. However, mouse p28 can be secreted without relying on other factors [8]. The receptor for IL-27 (IL-27R) is also a heterodimer; it is composed of IL-27Rα and glycoprotein 130 (gp130). IL-27Rα is unique to IL-27R in the IL-6/IL-12 family, while gp130 is also a subunit of the IL-6 receptor and IL-35 receptor [9]. The IL-27 receptor is present on various cell types, such as DCs, monocytes, macrophages, neutrophils, mast cells, eosinophils, T, B, and NK cells.

The IL-30/EBI3 combination, known as IL-27, has both regulatory and anti-inflammatory functions. At first, it was believed to be similar to IL-12, which promotes Th1 immunity by inducing T-bet and IL12R-beta-2 expression through STAT1 activation during Th1 differentiation [10]. However, it was later found that IL-27 also has a strong inhibitory effect during Th17 differentiation [11].

IL-27 uses a STAT1-dependent mechanism to stop activated T cells from developing ROR-gamma-t and IL-17 expression [12]. It also inhibits Th2 immunity by preventing Th2 differentiation and cytokine production or by directly suppressing ILC2s [13]. IL-27 is involved in Treg suppression; it suppresses TGF-beta-induced Foxp3+ regulatory T cell differentiation. Therefore, IL-27 has a wide range of functions that control T-cell immunity by targeting multiple pathways [14]. IL-27 can also function in collaboration with other cytokines, including IL-6, IL-10, IL-11, and interferons [15,16,17]. The immunomodulatory effects of IL-27 on a number of adaptive and innate immune cells are presented in Figure 1. On the spectrum between pro- and anti-inflammatory effects, it tends to lean toward anti-inflammation [18].

Interestingly, IL-27 has both pro-inflammatory and anti-inflammatory properties that are mediated by STAT1 and STAT3 activation, respectively. However, its pro-inflammatory effects rely on the induction of T-bet and IL-12Rβ2 expression. IL-27 has been found to have a dual function in preventing tissue damage caused by excessive inflammation. Its impact on NK cells and their ability to regulate tumor growth have been observed in certain mouse models, but not in all of them, suggesting that the effects of IL-27 on mouse NK cells may vary depending on the type of tumor [19].

In other words, IL-27 has a varying spectrum of functions, and it may be difficult to predict the overall effects of this cytokine as its functional activity relies on several factors, such as signaling pathways and target cells.

In oncology, IL-27 is often discussed as a potential agent capable of stimulating anti-tumor immunity [20]. Various preclinical murine models have provided evidence that IL-27 has strong antitumor effects against different types of tumors. These effects are achieved through several mechanisms and do not seem to cause any negative effects. These involve CD8+ T cells, natural killer cells, and macrophages, as well as antibody-dependent cell-mediated cytotoxicity and inhibition of cyclooxygenase-2 and prostaglandin E2 expression, as well as direct effects on tumor cells, including inhibition of cell proliferation, tumor cell apoptosis [21] and suppression of epithelial-mesenchymal transition.

However, anti-tumor immunity is rarely achieved through the activity of only IL-27. In the review by Kourko et al., IL-27 effects are investigated in combination with other cytokines (IL-30, IL-35). Interactions of these so-called ‘sister’ cytokines have the ability to hinder the development of tumors by either interacting with cancer cells directly or indirectly by activating various types of immune cells. For instance, IL-27 and other cytokines increase NK cell-mediated cytotoxic activity, promote the cytotoxic activity of CD8+ effector T cells, and stimulate the generation of CD8+ memory T cells. At the same time, IL-27 provides certain pro-tumor effects: for instance, it is known to be produced by tumor cells and inhibits chemotherapeutic effects [22]. When tumor cells undergo apoptosis, they are endocytosed by antigen-presenting cells, which stimulate regulatory T cell activation [23].

Overall, in the context of cancer, IL-27 is mostly discussed as a pro-inflammatory cytokine that stimulates the anti-tumor effects via activation of tumor-specific cytotoxicity. Unlike the stimulation of tumor immunity, IL-27 is often suggested as a therapeutic option in autoimmune diseases because it has anti-inflammatory effects.

In vivo studies of autoimmune encephalomyelitis showed that the introduction of IL-27 affected T helper 17 cells and suppressed autoinflammation [24]. In the review by Meka et al., IL-27 is described as one of the most important cytokines in autoimmune diseases. In rheumatoid arthritis, IL-27 is often found in the synovial tissue from affected joints. injections of IL-27 cause disease improvement when injected in a murine model of rheumatoid arthritis. Overall, a similar picture is seen for other autoimmune pathologies, i.e., systemic lupus erythematosus, colitis, psoriasis, and diabetes mellitus.

As mentioned above, IL-27 can play an ambiguous role in terms of inflammatory reactions. It is known to augment antigen-specific generation of cytotoxic lymphocytes [25], specifically via activation of T-bet (T-box transcription factor) and EOMES (Eomesodermin/Tbr2). In addition, IL-27 plays a key role in the generation of Th1 immune responses. IL-27 is produced by the antigen-presenting cells upon activation and promotes the rapid proliferation of CD4+ T cells that have previously encountered the antigen. It also works together with IL-12 to induce IFN-gamma production by CD4+T cells. However, IL-27 does not have the same effect on memory CD4+ T cells [27]. Lucas et al. showed that IL-27 is insufficient to stimulate full-scale IFN production by the innate cells, but it plays a contributing role to Th1 responses [28]. At the same time, IL-27 suppresses Th2 cell differentiation by blocking the expression of GATA-3, a transcription factor deeply involved in Th2 responses [29]. In the study conducted by Schneider R et al., it was demonstrated that IL-27 presence empowered CD8+ T cell proliferation and their development into Tc1 cells [29]; IL-27 promotes the expansion of self-renewing CD8+ T cells [30].

Overall, IL-27 interacts with cells via the IL-27 receptor and activates or inhibits their activity through signaling molecules STAT1/3.

3. Interleukin 27 in Infectious Pathology

Due to the abovementioned diversity of IL-27 effects, its role in infectious diseases is often a matter of debate. It launches pro-inflammatory and protective effects by stimulating Th1-mediated immune responses; at the same time, it elicits regulatory effects by causing the expression of co-inhibitory receptors, including PD-L1, LAG-3, TIM-3, CTLA-4, and TIGIT on T cells [31]. During infections, IL-27 levels are commonly increased and have been associated with the generation of IL-10, which could create a mechanism for controlling inflammation while still allowing the infectious agent to thrive [32].

In bacterial infections, especially those that are often followed by sepsis, IL-27 is usually elevated in the blood plasma [33]. The same tendency was noted in tuberculosis: IL-27 showed elevated concentrations in the plasma of patients with tuberculosis when compared to healthy donors [34]. In one of the studies, IL-27 was named ‘a wolf in sheep’s clothing’, due to the fact that in its absence, better control of mycobacterium growth was noted, but at the same time, its absence resulted in chronic hyperinflammation in later stages [35]. In another study on M. Tuberculosis, it was shown that IL-27 inhibits the anti-mycobacterial activity of macrophages [36]. In respiratory chlamydia infections, IL-27 is known to stimulate Th1-mediated inflammatory responses [37].

In viral infections, for instance, influenza, IL-27 plays an ambiguous role. In murine experiments conducted by Liu et al., IL-27 was found to limit immunopathology, neutrophil accumulation, and dampened Th1 or Th17 responses via IL-10-dependent and -independent pathways. And in the absence of IL-27 signals, a more severe disease course was observed [38]. The results indicate that IL-27 plays a crucial role in regulating immunopathology and that administering IL-27 at appropriate times could be an effective treatment for severe inflammation caused by infections.

At the same time, in parasitic infections, Th1 responses mediated by IL-27 signaling are directed towards the limitation of inflammation [39].

4. Interleukin 27 in COVID-19

As shown in other pathologies, IL-27 demonstrates a diversity of functions. We believe IL-27 to be a potential key player in the development of COVID-19 and protection against it. There are several studies concerning the involvement of IL-27 in COVID-associated immunity. IL-27 is a protein that is produced by immune cells when they are activated by pathogens or TLR ligands. It can stimulate the release of inflammatory cytokines from various types of cells, including mast cells, monocytes, and keratinocytes [40].

Initially, our investigation was aimed at analyzing the role of this cytokine in COVID-19 patients. Upon comparing acute-phase COVID-19 patients with healthy donors, we observed a statistically significant increase in IL-27 plasma levels in acute COVID-19 patients. Conversely, we also noted an opposite trend in convalescent patients, where concentrations of IL-27 were lower than those in healthy donors [41]. This tendency was seen independently of disease severity in the first-wave COVID patients in early 2020.

In a study performed by Zamani et al., IL-27 was proposed as a potential predictor for the severity and outcome of COVID-19 [42], along with IL-32 and neutrophil-to-lymphocyte ratio (NLR). IL-27 was identified as a factor indicating the necessity for hospitalization in patients who have a higher risk due to demographic factors such as being male, of black or Hispanic ethnicity, and having a median age over 63 years [43]. Another research used machine learning to predict the outcome of COVID-19 and identified a few cytokines, including IL-27, IL-9, IL-12p40, and MCP-3, as potential indicators for mild cases of the disease [49]. The 2023 publication authored by Valdés-López and Urcuqui-Inchima revealed that the identification of mRNA encoding IL-27 subunits in PBMCs and monocytes from individuals with COVID-19 was linked to the severity of the disease [45]. Moreover, mRNA expression levels of IL-27 signaling components in COVID-derived PBMCs showed positive correlations between the expression level of mRNA encoding the IL-27p28 subunit and IL-27 signaling components in COVID-derived PBMCs, including Gp130, JAK2, STAT1, STAT3, and SOCS3. In addition to our previous studies on the role of cytokines in COVID-19, we conducted another investigation to explore the association between Interleukin 27 (IL-27) concentrations and disease outcome. Our research showed that there was a significant increase in concentrations among both COVID-19 survivors and non-survivors when compared to healthy individuals. However, there was no statistical significance found when comparing the survivors and non-survivors [46]. And while it is possible that IL-27 may have predictive value, our analysis showed no sufficient sensitivity or specificity to support this claim. We used ROC analysis to assess the predictive value of IL-27 and found that it was not a reliable marker for prognosis.

In studies concerning other respiratory pathologies, IL-27 has shown a positive correlation with the severity of pulmonary inflammation [47]. In patients with community-associated pneumonia (CAP), serum levels of IL-27 were elevated in comparison with healthy donors [48].

However, as we delved further into the investigation of IL-27 and its involvement in COVID-19, we also analyzed the correlation between IL-27 and the SARS-CoV-2 genetic variant. In one of our studies, we performed multiplex cytokine profiling in plasma samples from patients infected with different SARS-CoV-2 variants (i.e., 2019-nCoV strain or Wuhan strain, B.1.1.7 or Alpha variant, B.1.617.2 or Delta variant, and B.1.1.529 or Omicron variant) [49]. Our results suggest that IL-27 plasma levels are not a reliable indicator of the severity of COVID-19 infection, as they do consistently vary with the viral variant. However, the four cytokines (IL-6, IL-10, IL-18, and IL-27) that were consistently elevated in COVID-19 patients may provide valuable insights into the equilibrium behind disease progression and host infection. Further research is needed to confirm these findings and determine their clinical significance.

When analyzing CD8+ T cell subpopulations in acute COVID-19 patients, convalescents, and healthy donors, we noted a negative correlation with the CCR6+CD8+ T cell frequencies within effector memory T cells and terminally differentiated effector memory T cells re-expressing CD45RA in patients with acute COVID-19 [50]. This tendency was not evident in COVID-19 convalescents or healthy controls. Moreover, serum IL-27 levels also showed a negative correlation with the absolute numbers of central memory and terminally differentiated effector memory cells (Tc17 cells) from patients with acute COVID-19 but not from convalescent or healthy donors. In the study by Schneider R et al., IL-27 presence was affecting CD8+ T cell proliferation and inducing their development into Tc1 cells [51]. In viral infections, it was highlighted that IL-27 promotes the expansion of self-renewing CD8+ T cells [52].

All findings concerning IL-27 in COVID-19 are presented in Table 1.

IL-27, among other cytokines, is deeply involved in immune-mediated processes associated with COVID-19. As its receptor is widely present on a variety of cells, its function is strongly interconnected with other cytokines and chemokines. We present our view on how IL-27 is integrated into immune processes in COVID-19 in Figure 2.

5. Conclusions

Interleukin 27 is a cytokine endowed with diverse functional activities. It can play both pro- and anti-inflammatory roles, depending on the signaling pathway and the target cells it affects. Overall, regardless of the pathological process it is involved in, IL-27 is critical for upholding the necessary balance between tissue damage and cytotoxicity against infectious agents and/or tumors.

In COVID-19, its role is also ambiguous. Being one of the few cytokines that show statistically significant changes in concentrations in the acute phase of the infection, IL-27 is directly linked to the markers of cell-mediated immunity. And even though its full role in the immune responses behind COVID-19 is yet to be explored, it is now clear that IL-27 may be one of the most interesting cytokines involved in this infection.

6. Future Directions

We are open to discussing this topic further and welcome any insights or perspectives that may help us better understand the role of IL-27 and other cytokines in COVID-19. By gaining a better understanding of the immune response to SARS-CoV-2, we hope to identify new therapeutic targets and develop more effective treatments for this devastating disease.