Resolving the developmental mechanisms of coagulation abnormalities characteristic of SARS-CoV2 based on single-cell transcriptome analysis (preprint)

The COVID-19 outbreak caused by the SARS-CoV-2 virus has developed into a global health emergency. In addition to causing respiratory symptoms following SARS-CoV-2 infection, COVID-19-associated coagulopathy (CAC) is the main cause of death in patients with severe COVID-19. In this study, we performed single-cell sequencing analysis of the right ventricular free wall tissue from healthy donors, patients who died in the hypercoagulable phase of CAC, and patients in the fibrinolytic phase of CAC. Among these, we collected 61,187 cells, which were enriched in 24 immune cell subsets and 13 cardiac-resident cell subsets. We found that in response to SARS-CoV-2 infection, CD9highCCR2high monocyte-derived mo promoted hyperactivation of the immune system and initiated the extrinsic coagulation pathway by activating CXCR-GNB/G-PI3K-AKT. This sequence of events is the main process contributing the development of coagulation disorders subsequent to SARS-CoV-2 infection. In the characteristic coagulation disorder caused by SARS-CoV-2, excessive immune activation is accompanied by an increase in cellular iron content, which in turn promotes oxidative stress and intensifies intercellular competition. This induces cells to alter their metabolic environment, resulting in an increase in sugar uptake, such as that via the glycosaminoglycan synthesis pathway, in CAC coagulation disorders. In addition, high levels of reactive oxygen species generated in response elevated iron levels promote the activation of unsaturated fatty acid metabolic pathways in endothelial cell subgroups, including vascular endothelial cells. This in turn promotes the excessive production of the toxic peroxidation by-product malondialdehyde, which exacerbates both the damage caused to endothelial cells and coagulation disorders.

Effects of (Combined) Chinese Medicine on the Risk of Death From COVID-19: A Retrospective Data Analysis Based on 4567 Patients (preprint)

Background The coronavirus disease 2019 (COVID-19) epidemic is still ongoing, but the optimal treatment remains unclear. China adopted a series of measures, including widespread screening, strict quarantine and early treatment, combining western medicine with Chinese medicine, leading to rapid control of its spread. Nevertheless, the effects of ( combined ) Chinese medicine in reducing the toll of COVID-19 lack proof from statistics.Objective We conducted a retrospective data analysis to determine whether ( combined ) Chinese medicine is able to affect patient outcomes and to decrease the risk of death in COVID-19 patients.Methods The data were acquired by outputting the formatting information from the HIS system and then extracting and recording it in the database for complete cases. The demographics, disease onset, treatment, survival/death and all of the clinical classifications, groups and definitions were verified by specialists in the clinic, along with the research methodology and statistics, before conducting the statistical analysis. The characteristics of the cohort and the clinical symptoms and signs, prescriptions and outcomes were described and analyzed by the mean ± SD, median, interquartile range and composition ratio. Analysis of variance was used for comparisons between the measurement data sets; otherwise, the rank sum test was used. Counting data were compared between groups using the chi square test and Fisher’s exact test. Tendency matching was adopted to make the general data balance between groups. A Cox proportional hazard model was used to compare the risk of death among the different groups.Results Four centers were included in our study, and a total of 6,076 patients' clinical records were obtained after combining the data. We included 4567 cases for the descriptive statistics, and the crude case fatality rate was 3.0%. Compared with using only western medicine, (combined) Chinese medicine reduced the risk of death from COVID-19 after adjusting for other prognostic risk factors (HR = 0.135, 95% CI (0.088, 0.208)). Multivariate Cox regression also indicated that when applying the clinical classification of severe/critical, age ≥ 65 years old, coronary heart disease or chronic kidney disease and a time from onset to hospital admission of fewer than 14 days, all of these factors increased the risk of death.Conclusion (Combined) Chinese medicine can significantly reduce the risk of death from COVID-19, but the specific strategy/solution, effects and amount need further exploration in future studies.

Transcriptome analysis of PBMCs reveals distinct immune response in the asymptomatic and re-detectable positive COVID-19 patients (preprint)

The existence of asymptomatic and re-detectable positive COVID-19 patients presents the disease control challenges of COVID-19. Most studies on immune response of COVID-19 have focused on the moderately or severely symptomatic patients, however little is known about the immune response in asymptomatic and re-detectable positive patients. Here we performed a comprehensive analysis of the transcriptomic profiles of PBMCs from 48 COVID-19 patients which include 8 asymptomatic, 13 symptomatic, 15 recovering and 12 RP patients. Our analysis revealed a down-regulation of IFN response and complement activation in the asymptomatic patients compared with the symptomatic, indicating a weaker immune response of the PBMCs in the asymptomatic patients. In addition, we observed a lower expression of the cytokines and chemokines in the PBMC of asymptomatic and symptomatic patients. In contrast, the cytokines and chemokines level in the RP patients are higher than the recovering. GSEA analysis showed the enrichment of TNFa/NF-{kappa}B and influenza infection in the RP patients compared with the recovering patients, indicating a flu-like, hyper-inflammatory immune response in the PBMC of RP patients. Thus our findings could extend our understanding of host immune response during the progression COVID-19 disease and help the clinical management and the immunotherapy development for COVID-19.

T-cell Repertoire Characteristics of Asymptomatic and Re-detectable Positive COVID-19 Patients (preprint)

Background: The prevention of COVID-19 pandemic is highly complicated by the prevalence of asymptomatic and recurrent infection. Many previous immunological studies have focused on symptomatic and convalescent patients, while the immune responses in asymptomatic patients and re-detectable positive cases remain unclear. Methods: Here we comprehensively analyzed the peripheral T-cell receptor (TCR) repertoire of 54 COVID-19 patients in different phases, including asymptomatic, symptomatic, convalescent and re-detectable positive cases. Results: We found progressed immune responses from asymptomatic to symptomatic phase. Furthermore, the TCR profiles of re-detectable positive cases were highly similar to those of asymptomatic patients, which could predict the risk of recurrent infection. Conclusion: Therefore, TCR repertoire surveillance has the potential to strengthen the clinical management and the immunotherapy development for COVID-19.

Health-related quality of life of COVID-19 survivors at 6 months after hospital discharge: a cohort study (preprint)

Background: COVID-19 is a multi-systemic disease that is highly contagious and pathogenic. The long-term consequences of it are not yet clear, as is whether society and life can return to a healthy state. Long-term assessment of their health-related quality of life (HRQoL) is essential. This study aimed to investigate HRQoL and its risk factors in COVID-19 survivors at a follow-up of 6-month. Methods: A multicenter cross-sectional survey was conducted among 192 COVID-19 patients with confirmed age ≥ 18 years who were discharged from various hospitals in Wuhan from January to April 2020. The demographic characteristics, clinical characteristics, and laboratory results of the study subjects were obtained from the hospital's medical records. Survivors' HRQoL was assessed using the Short Form 36 (SF-36), cognition was assessed using the ascertain dementia eight-item informant questionnaire (AD8), and survivors' pulmonary function were examined. All participants in this study completed the survey and testing at Hubei Provincial Hospital of Chinese and Western Medicine. SF-36 scores were compared with the Chinese norm, and logistic regression and multivariate analysis were used to investigate the factors affecting HRQoL in COVID-19 survivors. Results: SF-36 showed significant differences in HRQoL between COVID-19 survivors and the general Chinese population ( P< 0.05).Multiple linear regression demonstrated that age was negatively correlated with physical functioning (PF), role-physical limitation (RP) and social functioning (SF) ( P <0.05). Bodily pain (BP), vitality (VT), SF and role-emotional limitation (RE) were negatively correlated with females ( P <0.05). Length from discharge to follow‐up was positively correlated with PF and RP ( P <0.05). Abnormal cognitive function was negatively correlated with PF, RP, general health (GH), VT, SF, RE and mental health (MH) ( P <0.05). Abnormal Carbon Monoxide Diffusing Capacity (DLCO%<80%) was significantly negatively correlated with PF and SF ( P <0.05).In addition, there was a significant negative correlation between Coronary heart disease and RP, GH, VT and RE ( P <0.05).Logistic regression analysis demonstrated that age(OR 1.032) and AD8 scores (OR 1.203)were risk factors associated with a low physical component summary (PCS) score. Length from discharge to follow‐up (OR 0.971) was the protective factor for PCS score. Abnormal cognitive function (OR 1.543) was a significant determinant associated with a mental component summary (MCS)<50 in COVID-19 patients. Conclusions: The HRQoL of COVID-19 survivors remains to be improved at six-month follow-up. Future studies should track HRQoL in older adults, women, patients with abnormal DLCO, and abnormal cognitive function for a long time and provide them with rehabilitation advice and guidance.

Long-Term Clinical Sequelae and Immunological Features of COVID-19 Survivors: A Cross-Sectional Study in Wuhan, China (preprint)

Objectives: To describe the clinical sequelae and immunological features of COVID-19 survivors who have been discharged from the hospital for 5-8 months. Methods: This study is a cross-sectional study of confirmed COVID-19 patients aged ≥18 years who were discharged from hospitals in Wuhan from January to April 2020. The demographics, clinical features, and laboratory findings of the participants were collected from medical records in the hospital. The participants from the study completed an investigation of clinical sequelae, blood tests, a pulmonary function examination and an unarmed rehabilitation evaluation at Hubei Provincial Hospital of Traditional Chinese & Western Medicine. A group of volunteers who were free of COVID-19 and lived in Wuhan during the outbreak were recruited as the comparison group.Results: The average age of the 574 COVID-19 survivors was 57.7±11.4 years, and 348 (60.6%) survivors were female. The average number of days from the onset of symptoms was 241.79±16.16. The average number of days from discharge was 194.3±14.4. Clinical sequelae were common, including general symptoms (n=321, 55.9%), respiratory symptoms (n=265, 46.2%), digestive symptoms (n=84, 14.6%), nervous symptoms (n=75,13.1%) and psychosocial symptoms (n=201, 35%). A total of 190 (33.7%) survivors reported reduced exercise capacity. Through the results of pulmonary function examination, anomalies were noted in carbon monoxide diffusion capacity (DLCO)% in 110 cases (32.4%), maximal mid-expiratory flow (MMEF)% in 105 cases (30.7%), forced expired flow at 50% of forced vital capacity (FEF 50 )% in 128 cases (37.4%), and forced expired flow at 75% of forced vital capacity (FEF 75 )% in 240 cases (70.2%). The counts of T lymphocyte, CD4+ T lymphocyte counts, CD8+ T lymphocyte counts, B lymphocyte and NK cell in the survival group was significantly lower than that in the comparison group(all P < 0.05).There were 252 (43.9%) survivors whose total T lymphocyte counts had dropped, 260 (45.3%) whose CD4+T lymphocyte counts had dropped, 231 (40.2%) whose CD8+ T lymphocyte counts had dropped, 119 (20.7%) whose B lymphocyte counts had dropped, and 54 (9.4%) whose NK cell counts had dropped. But there were no statistically significant differences in the incidence of lymphocyte subsets reduction between severe and nonsevere groups (all P > 0.05). The T lymphocyte counts, CD4+ T lymphocyte counts and CD8+ T lymphocyte counts of the patients before discharge were significantly higher than those in the early stage of admission (P < 0.025). There were 319 (55.6%) survivors with positive or weakly positive IgG antibodies and 17 (2.9%) survivors with positive or weakly positive IgM antibodies.Conclusion: Even after 5 to 8 months of discharge, many survivors still have clinical sequelae, and some of them have impaired immune function. Therefore, the long-term rehabilitation of COVID-19 survivors remains a concern.Funding Statement: This study was funded by the National Key R&D Plan of China (2020YFC0841600), Guangdong Provincial Key Laboratory of Research on Emergency in TCM (2017B030314176), R&D plan in key areas of Guangdong Province (2020B1111300005), and National Administration of Traditional Chinese Medicine (2020ZYLCYJ05-11).Declaration of Interests: The authors declare that they have no competing interests.Ethics Approval Statement: This study was approved by the ethical committees of Guangdong Provincial Hospital of Chinese Medicine, Guangzhou, China (GPHCM; No. BF2020-205-01). All participants signed informed consent forms.