Review 2: "Outcome of SARS-CoV-2 infection linked to MAIT cell activation and cytotoxicity: evidence for an IL-18 dependent mechanism"

RR:C19 Evidence Scale rating by reviewer:

• Strong. The main study claims are very well-justified by the data and analytic methods used. There is little room for doubt that the study produced has very similar results and conclusions as compared with the hypothetical ideal study. The study’s main claims should be considered conclusive and actionable without reservation.

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Review:

Mucosa-associated invariant T (MAIT) cells are potent innate-like T lymphocytes enriched in mucosal layers. They respond vigorously to bacterial infections and also indirectly, through cytokine receptor signaling, to viruses. MAIT cells are thought to play protective roles against several viral pathogens and also participate in tissue repair mechanisms that may potentially assist in long-term recovery from certain infections. On the other hand, MAIT cells may be viewed as double-edge swords whose uncontrolled activation in response to MHC-related protein 1 (MR1) ligands, inflammatory cytokines, or both, may result in a cytokine storm, immunopathology and tissue damage [1]. Given the abundance of MAIT cells in the human respiratory tract, the main port of entry for many viruses including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), it is important to reveal what functions MAIT cells play in various stages of coronavirus disease 19 (COVID-19), which has been the main subject of the investigation by Flament et al.

The authors assessed the numerical, phenotypic and functional changes human peripheral blood (PB) MAIT cells undergo during COVID-19 and their potential correlations with inflammatory markers and clinical measures of disease severity and outcomes. They recruited patients admitted to infectious disease units (IDU) or intensive care units (ICU) with moderate and severe COVID-19, respectively. Blood samples previously obtained from uninfected subjects were used as controls. Patient and control cohorts were carefully matched for age and body mass index (BMI). In addition, IDU and ICU patients were matched for the presence of diabetes as judged by HbA1c levels. MAIT cell frequencies decline as we age, and obesity and diabetes are among significant comorbidities that increase the risks of COVID-19 severity and mortality. Therefore, Flament and coworkers should be commended for taking these pre-existing conditions into consideration in designing their study. Future studies may include an additional control cohort of non-COVID ICU patients to enable investigation into the impact of critical illness per se on MAIT cell functions. In addition, given the sexual dimorphism of COVID-19 severity, it will be pertinent to match IDU and ICU patients more closely for sex.

As previously reported in other viral infections, MAIT cell frequencies were reduced in the peripheral blood, which may be due to their recruitment to the site(s) of infection/inflammation and/or their activation-induced cell death. This notion is supported by the authors’ observation that CCR6⁺CD127⁺ cell frequencies were reduced among MAIT cells that had remained in the circulation of COVID-19 patients. Also of note, according to a preprint from another prominent group, MAIT cells are enriched in the airways of COVID-19 patients [2].     

Flament et al. report that PB MAIT cells from COVID-19 patients exhibit an activated phenotype, as judged for instance by CD69 expression, which is not surprising and neither is granzyme B (GZM B) upregulation by MAIT cells. However, their work highlights several other important findings that deserve close attention. First, ICU patients had higher percentages of CD56⁺, CD69⁺ and CD56⁺CD69⁺ PB MAIT cells than did IDU patients. Intriguingly, the IFN-γ production capacity of MAIT cells was diminished in IDU patients but augmented in ICU patients. This was evaluated in ex vivo stimulation cultures using a combination of phorbol 12-myristate 13-acetate and ionomycin, which bypasses the requirement for T cell receptor (TCR) engagement. It will be interesting to examine whether this phenomenon can be recapitulated in response to MR1 ligands such as 5‐(2‐oxopropylideneamino)‐6-D‐ribitylaminouracil (5-OP-RU) and MAIT cell-activating cytokines such as IL-12 and IL-18. Second, a strong trend was found towards increased CD69 expression by MAIT cells from patients who had died of COVID-19 in comparison with survivors in either the IDU or the ICU cohort. Furthermore, IFN-γ⁺, TNF-α⁺ and GZM B⁺ cells were more frequent among stimulated MAIT cells from patients who had succumbed to COVID-19. These results suggest that the activation status of MAIT cells may help predict the probability of death versus survival. Additional analyses in which survivors and non-survivors are compared in each cohort (IDU and ICU patients independently) will be informative. Third, plasma levels of IL-6, IL-8, IL-10, IL-15 and IL-18 were elevated in non-survivors, suggesting that a cytokine storm may be responsible, at least partially, for COVID-19 mortality. In COVID-19 patients combined, the plasma levels of the above cytokines were correlated with CD69⁺ and CD56⁺CD69⁺ MAIT cell frequencies. Moreover, a strong correlation was evident between plasma IL-18 levels and GZM B⁺ MAIT cell percentages in non-surviving ICU patients. Whether increased GZM B expression by MAIT cells equals their enhanced cytotoxicity and what this may mean during COVID-19 are unclear at this point and may form the basis of future studies. MAIT cells’ ability to degranulate may be assessed, for instance by CD107a expression analysis. Last but certainly not the least, an activated MAIT cell phenotype (CD69 expression) was associated with lower PaO₂/FiO₂ (partial pressure of arterial oxygen over fraction of inspired oxygen) ratios, indicative of hypoxemia, and with higher SAPS (simplified acute physiology score) II scores, predictive of in-hospital mortality. These results further suggest that MAIT cell activation may be detrimental to COVID-19 patients. It is noteworthy that in a separate study, CD69 expression by MAIT cells on day 1 post-admission correlated positively with the PaO₂/FiO₂ ratio on day 7 [3]. Therefore, future studies will need to test blood and tissue MAIT cell functions at multiple snapshots in the course of COVID-19 when logistically possible. Clinically relevant animal models will prove useful as they enable longitudinal and mechanistic studies on MAIT cells as well as their therapeutic potentials.

In summary, the impressive work by Flament et al. has yielded several novel findings and provided important insight into MAIT cell roles in COVID-19. The results presented in this preprint, along with those of Parrot et al. [2] and Jouan et al. [3], advance the field of MAIT cell immunology and set the stage for additional MAIT cell analyses during infection with SARS-CoV-2 and likely other respiratory viruses with pandemic potentials. I apologize to other investigators whose relevant work in this area may have been overlooked.              

1.     Haeryfar SMM: MAIT cells in COVID-19: heroes, villains, or both? Crit Rev Immunol 2020, 40(2): 173-184 

2.     Parrot T, et al: https://www.medrxiv.org/content/10.1101/2020.08.27.20182550v1

3.     Jouan Y, et al: https://www.medrxiv.org/content/10.1101/2020.05.03.20089300v1