Although there is a growing body of research that suggest sex differences in health outcomes of COVID-19, it is not understood whether immune responses to COVID-19 differ based on sex. In this study, the researchers evaluated sex differences in virus levels, SARS-CoV-2 specific antibodies, cytokines, and blood phenotype in COVID-19 patients with mild to moderate disease. The researchers found that male patients had more innate immune response activation, with higher cytokine and chemokine levels, as well as higher monocyte activation. On the other hand, female patients had more adaptive immune response activation, through T-cell activation, than male patients, even in old age. In addition, the researchers found that poor T-cell activation was associated with older age and worse disease outcomes in male patients, but not female patients. In contrast, higher cytokine activation in female patients was associated with worse disease outcomes, but not in male patients. These findings may explain sex differences in COVID-19 and influence sex-based approaches to COVID-19 treatment.
The researchers wanted to understand how and if the immune system was responding differently against COVID-19 based on sex.
A growing body of research is suggesting that male sex is a risk factor for more severe COVID-19. Previous studies have demonstrated that sex can impact health outcomes for infectious diseases due to sex differences in immune responses to infection, such as in hepatitis A, tuberculosis, hepatitis C, and HIV. These studies suggest that the immune systems of women have better capabilities to control infectious disease. However, the mechanism of how SARS-CoV-2 causes more severe disease in male patients is unknown. To understand this mechanism, the researchers analyzed sex differences in immune responses against SARS-CoV-2 by evaluating viral levels, specific antibody levels, cytokine/chemokine levels, and blood phenotypes.
Overview of Study Design
The researchers enrolled patients admitted to Yale-New Haven Hospital who were positive for SARS-CoV-2 infection by RT-PCR. 198 patients were enrolled in the study, and the researchers obtained peripheral blood mononuclear cells (PBMCs), plasma, nasal swabs or saliva samples from 93 of these patients. Uninfected healthcare workers were enrolled as a control group.
Baseline analyses were performed on patients who were not in the intensive care unit, had not received tocilizumab, and had not received high dose corticosteroids, in order to make sure the measurements were accurate and reflective of COVID-19 disease rather than the effects of treatments.
The researchers also performed analyses on all patients, whether they were excluded or included.
Male patients have higher levels of key innate immune cytokines
The researchers compared viral levels in male and female patients, and found that there was no significant difference between sexes.
The researchers also found that SARS-CoV-2 spike protein-specific antibodies, such as IgG and IgM, were activated in many patients. IgG levels tended to be higher in female patients, although the difference was not significant.
Next, the researchers analyzed 71 cytokines and chemokines that have been shown to have elevated levels in COVID-19 patients. Aligning with previous studies, the researchers found that these cytokines/chemokines had higher levels in COVID-19 patients than in controls. Although cytokine/chemokine levels were similarly increased in men and women, IL-8 was higher in male patients and IL-18 was higher in female patients, but not significantly. CCL5 and CXCL10 were increased in male patients. These results indicate that inflammatory cytokines are more elevated in male COVID-19 patients.
Phenotypic differences in monocytes between male and female COVID-19 patients
The researchers investigated immune cell types in the blood of COVID-19 patients, including T-cells, B cells, NK-T cells, NK cells, monocytes, macrophages, and dendritic cells. T-cells were significantly reduced in patients, but levels were comparable between sexes. In addition, there was an increase in monocyte levels in both sexes, but CD14+CD16+ monocytes were elevated in female patients. In contrast, CDloCD16+ was elevated in male patients, as well as non-classical monocytes (ncMono). The researchers investigated the relationship between elevated IL-8 and CCL5 with elevation of ncMono in male patients. They found no correlation with IL-8 but a significant correlation between CCL5 and ncMono in male patients. These results suggest that the progression from classical monocytes (which differentiate into other immune cells) to non-classical monocytes (which are only anti-inflammatory) stops at the intermediate stage of COVID-19 in women. They also suggest that inflammatory cytokines/chemokines are associated with increased activation of innate immune cells in male patients.
Female COVID-19 patients induce more robust T cell response than male patients
The researchers evaluated the T-cell phenotype in COVID-19 patients. They found that the composition and phenotype of many kinds of T-cells were similar. However, they observed CD38 and HLA-DR-positive activated T cells, as well as PD-1 and TIM-3-positive T cells, were more commonly activated in female patients than in male patients for both CD4 and CD8 T cells. These results suggest that female COVID-19 patients had more activated and differentiated T cells than male patients.
Sex-differences in immune phenotype associated with worsening of COVID-19 disease
Lastly, the researchers investigated whether certain immune phenotypes and their sex differences could predict severity of COVID-19. They evaluated disease progression of all enrolled patients and divided patients into two groups, stabilized and deteriorated, and further categorized these groups by sex. In terms of demographics, deteriorated males were significantly older than the stabilized males, and had significantly higher BMI. There was no difference between stabilized and deteriorated female patients for age and BMI. However, the stabilized female patients had lower virus levels compared to the deteriorated groups and the two male groups. In addition, higher IgG antibody levels were observed in stabilized female patients, indicating that high IgG levels were associated with women being able to control disease progression.
The researchers further investigated if the components of the immune response explored above were associated with disease progression in male and female patients. They observed that regardless of sex, many chemokines were elevated in patients who developed severe disease. However, some cytokines such as CCL5, TNFSF10, and IL-15 were specifically elevated in female patients who progressed to worse disease, but not in male patients. After adjusting for age, CXCL10, a chemokine, was only elevated in female patients that progressed to to worse disease.
In terms of T-cell phenotypes, male patients who progressed to worse disease had lower levels of activated T-cells, and fewer differentiated T-cells compared to male patients who did not deteriorate. In women, both deteriorated and stabilized groups had similar levels of activated and differentiated T-cells.
Finally, the researchers investigated relationships between age, BMI, virus levels, specific antibodies, cytokines/chemokines, activated/differentiated T-cells, and disease progression. This analysis showed that in women, higher cytokine levels were significantly associated with disease progression, but not activated/differentiated T-cells. On the other hand, in male patients, severe disease was associated with higher age, higher BMI, and poor T-cell activation/differentiation. In male patients, poor T-cell activation/differentiation was significantly correlated with higher age, but this trend was not observed in female patients.
In conclusion, these sex differences in immune responses to SARS-CoV-2 may reveal potential predictive factors for COVID-19 disease progression.