In this study, we aimed to analyze the role of lymphocyte subsets in the immunopathogenesis of COVID-19 and severe influenza A, and examined the clinical significance of their alterations, especially in determining the prognosis and recovery duration. Our analyses revealed significant dynamic variations in total lymphocytes and lymphocyte subsets, which get activated in the early stages of COVID-19 and severe influenza A infections, and further demonstrated that severe immune injury tended to be more prominent in patients with severe form of the disease. The recovery rate of patients with severe COVID-19 was comparatively longer than those who received immediate antiviral treatment for severe influenza A and those with non-severe COVID-19.
We retrospectively reviewed the clinical data of 99 patients who were confirmed to have COVID-19, 37 patients with severe influenza A and 94 healthy blood donors who were previously recruited in 2018. All the patients with COVID-19 were divided into two groups, according to the abovementioned diagnostic criteria, including 80 non-severe cases (80.8%) and 19 severe cases (19.2%). Several reports and studies have clearly indicated that older or elderly people are more prone to COVID-19 as their immune systems are likely to get overwhelmed by infections due to their advanced age. Similarly, the elderly who are 65 years or older are particularly at risk for influenza infection, hospitalization, and death due to influenza-related complications, such as pneumonia. In our study, the median age of patients with non-severe COVID-19 was 37 years, while that of those with severe COVID-19 was 67 years. Consistent with the previous studies, our data indicate that the ages of the severe patient group are higher than those of the non-severe COVID-19 group. Further, our study showed that the median age of patients with severe influenza A infection was higher ( 67 years) than that of those with non-severe COVID-19 (37 years) and healthy controls ( 47years). Notably, this difference in age was found to be statistically significant (P < 0.05) among the groups, which suggests that the elderly people represent a large at-risk population.
The WHO-China joint report on COVID-19 provided a comprehensive symptomatology of COVID-19 (n = 55,924).A previous study showed that patients with COVID-19 present with pyrexia in 85% of cases during their illness course, but only 45% are febrile on early presentation. In addition to cough (67.7%) and sputum (33.4%), respiratory symptoms, such as dyspnea, sore throat, and nasal congestion were reported to be present in 18.6%, 13.9%, and 4.8% of cases, respectively. Both COVID-19 and influenza present with common clinical manifestations including fever, cough, rhinitis, sore throat, headache, dyspnea, and myalgia. [7,14,15]In our study, all the patients with severe COVID-19 presented with fever, cough, and dyspnea, whereas only 73% and 48% of patients with non-severe COVID-19 had fever and cough, respectively. Similar to the severe COVID-19 group, all patients with severe influenza A infection presented with fever and cough. However, only 76.0% of the patients had dyspnea. These clinical manifestations of COVID-19 and influenza infections were consistent with other studies. As previously reported the mortality rate also increases in patients with additional comorbidities. Specifically, 63% and 89% patients with severe COVID-19 and severe influenza A, respectively, presented with one or more pre-existing chronic medical conditions. While the mortality rate was 5% due to severe COVID-19, it was even higher (11%) for severe influenza A.
Lymphocytes and their subsets play a crucial role in maintaining immune homeostasis and inflammatory response in the host. As in case of immune disorders and other infections, a viral infection impairs the host’s immune defenses and results in decreased levels of lymphocytes and their subsets.[16,17] Lymphocyte subsets, namely CD4+ T cells, CD8+ T cells, B cells, and NK cells are primarily involved in the humoral and cytotoxic immunity against viral infection. Therefore, this necessitates the need to understand the mechanism of reduced blood lymphocyte levels and characterize the dynamic alterations of lymphocyte subsets to provide novel insights for an effective treatment and prognosis of COVID-19.As lymphopenia is frequently observed during the initial stages of respiratory viral infection.[3,18] We identified and analyzed different variations in leukocytes, lymphocytes, and T lymphocyte subsets in patients with non-severe and severe COVID-19 and severe influenza A. . The counts of WBCs and lymphocytes, but not the counts of T cells, nor CD4+ or CD8+ T cells of non-severe COVID-19 patients decreased significantly compared with healthy controls in group one during the first week of illness. No significant changes were seen CD4+ T, CD8+ T cells and CD4+/CD8+ ratios of patients with non-severe COVID-19 during the four weeks following the infection. On the contrary, the counts of T cells, CD4+ and CD8+ T cells in patients with severe COVID-19 were significantly lower than those of healthy controls in group two from week1 to week3 after illness onset. Further, CD8+ T cells did not improve significantly at week4 of illness compared with healthy controls. Although the counts of WBCs, lymphocytes and T lymphocyte subpopulations were not compared between non-severe and severe COVID-19 patients due to the age mismatch, this results still suggested that T lymphocytes and their subsets were damaged more drastically in severe COVID-19 patients. These findings are corroborated by a previous study which hypothesized that the virus might directly infect lymphocytes resulting in their apoptosis, thus leading to causing a sharp decline in total lymphocyte population and subsequent lymphopenia. Moreover, lymphocytes express the coronavirus receptor angiotensin-convertingenzyme 2(ACE-2), and therefore are a direct target for the virus. Another retrospective study suggested that lymphopenia might be one of the predictive factors for progression to respiratory failure during early stages following Middle East Respiratory Syndrome coronavirus (MERS-Cov) infection. Giving further strength to our study, a study by Geng et al. demonstrated the decline in the populations of T lymphocytes and their subsets, after influenza A virus infection, to be positively correlated with prognosis.Meantime, our results reiterate the fact that CD8+ T cell responses play a major role in antiviral immunity. Taken together, lymphopenia was common in the patients with COVID-19, indicating a significant impairment in the host’s immune system following SARS-CoV-2 and influenza A infections. Our findings are in line with other studies which also detected these alterations in patients with pneumonia caused by MERS-CoV and Severe acute respiratory syndrome coronavirus (SARS-CoV).[20, 23] In addition, a significant reduction in both CD4+ T cells and CD8+ T cells were specifically observed in patients with severe COVID-19 and severe influenza A. Therefore, this indicates a more severe immune insult in patients with the severe form of the disease. Consequently, the alteration would be more profound, and leads to adverse clinical outcome in these patients. Thus, lymphocytes and their subsets, especially CD8+ T cells, might be a potential predictor for disease severity and clinical efficacy in COVID-19.
Next, we focused on the dynamics of T lymphocytes and their subsets, which played a vital role in cellular immune responses. compared the absolute counts of leukocytes, total lymphocytes, and lymphocyte subsets of the non-severe and severe COVID-19 patient groups at weeks 2, 3, and 4 with that of those observed during the initial stages of infection. The total leukocyte, lymphocyte, and T cell counts significantly improved at week 3 in patients with non-severe COVID-19. Lymphocyte also recovered markedly at week 3 in severe COVID-19 patients. However, T cell and CD4+ T cell subset population significantly increased at week 4 in patients with severe COVID-19. Our results are consistent with a previous study by He et al. that showed a sharp decline (below normal) in the cell counts of CD45+, CD3+ T cell subsets, CD4+ T cell subsets, and CD8+ T cell subsets during the first week of SARS-Cov infection; their values further declined during the second week before increasing during the third week and returning to normal by the fifth week. Moreover, CD4+ T and CD8+ T cell counts were found to be extremely low in critically ill and deceased patients. Taken together, the alterations in lymphocytes and their subsets gradually improved at later time points in patients with COVID-19. Collectively, our results indicate that the recovery duration of patients with severe COVID-19 is longer than those with the mild form of the disease.
Our study results further revealed a noticeable difference in the time taken for the cell counts to improve among the severe and non-severe COVID-19 and the severe influenza groups. The cell counts of total lymphocytes and their subsets recovered only around week 4 in severe COVID-19; the recovery time was almost delayed by a week compared with those having non-severe COVID-19. On the contrary, the cell counts of total lymphocytes and their subsets in patients with severe influenza A increased and improved drastically at week2; this rapid recovery rate could be attributed to the early initiation of treatment with the neuraminidase inhibitor, oseltamivir, or peramivir, which interfere with virus release from host cells by blocking the viral nucleic acid function, thus preventing infection of new host cells.
The relative role(s) played by the immune response to SARS-CoV-2 versus direct viral effects in the respiratory system and other organ systems has been questioned, with the possibility of immunopathogenesis being a major causal component of severe COVID-19 [25, 26]. Limited by research conditions, our study only demonstrated that severe COVID-19 cases had significant lymphopenia, especially the helper (CD3+CD4+) and cytotoxic T cells (CD3+CD8+), in the early stage of illness onset, consistent with previous research results, and lymphocytes and their subgroups in severe COVID-19 patients require a longer recovery period, compared with severe influenza patients. These results can give more points to other studies about T lymphocyte dynamic. The percentage of naïve helper T cells increased and memory helper T cells decreased in severe cases may be an important phenomenon in lymphopenia of severe COVID-19 patients . Moreover, the function of CD4+ T cells, CD8+ T cells and NK cells was within normal range which indicated by PMA/Ionomycin stimulated IFN-γ positive cells in these three subsets, and there was no significant difference between severe and non-severe COVID-19 cases. At the same time, T cell counts recovered was related to the concentration of cytokines in plasma likes IL-6, IL-10, and TNF-α, which was decreased in the disease resolution stage of COVID-19 patients . They also observed that increasing exhausted marker PD-1 expression on T cells was tested as COVID-19 patients progressed from prodromal to overtly symptomatic stages, which demonstrated that the surviving T cells appear functionally exhausted, not just a decrease in T cell counts. In summary, we would continue further studies on the function of T cell subsets across the spectrum of COVID-19 disease severity to gain insights into SARS-CoV-2 potential immunopathogenesis.
Our study has several limitations. Firstly, this study was retrospective, small single-center, cross-sectional clinical study. Only a small sample of 99 patients with COVID-19 admitted to Beijing Ditan Hospital was included, which may confound the results and potentially introduce selection bias. This may limit the generalizability of the study. At the same time, the study cohort was limited by the actual conditions of the admitted patients. Among the 99 COVID-19 patients, there were 80 non-severe cases and 19 severe ones. Non-severe COVID-19 patients were mainly young people, while severe COVID-19 patients were mainly elderly patients. The age data of the non-severe and severe COVID-19 cohorts could not be matched. In addition, inconsistencies in time periods between illness onset and admission might have led to missing data which could result in observation biases in the dynamic variations in immune cells.