2025
PhD Thesis
The long-term cardiovascular complications of SARS-CoV-2 infection and COVID-19 vaccines
University of Queensland, Australia
Service type: Stock strains
Abstract
Long COVID or post-acute sequelae of COVID-19 (PASC) (used interchangeably throughout this thesis) continues to be a serious health issue around the globe, currently affecting an estimated 11% of adults who have ever suffered from COVID-19 with over 200 symptoms across 10 different organ systems over 12 weeks post-infection. Of particular concern are complications affecting the cardiovascular system, which have been reported both during acute infection and in long COVID (termed PASC-CVS). Proposed mechanisms are hotly debated, with emerging evidence suggesting a role for a persistent aberrant pro-inflammatory response, with significant but very low-level (~10-20pg/mL) plasma inflammatory marker increases detected up to eight months post-infection in patients suffering from long COVID compared to fully-recovered patients. These circulating proinflammatory mediators may be recognised by different cell types around the body, including cardiomyocytes, and cause downstream effects that promote further cytokine release, electrical remodelling, and apoptosis. This possible role of chronic inflammation in cardiovascular sequelae, particularly in individuals with symptoms persisting for >1 year after SARS-CoV-2 infection, remains to be clearly defined. In the cross-sectional study presented in Chapter 2, blood samples were obtained from three different sites in Australia from individuals with i) a resolved SARS-CoV-2 infection (and no persistent symptoms i.e. ‘Recovered’), ii) individuals with prolonged PASC-CVS (i.e. chest pain and heart palpitations) and iii) SARS-CoV-2 negative individuals. Individuals with PASC-CVS, relative to Recovered individuals, had a blood transcriptomic signature associated with inflammation. This was accompanied by elevated levels of pro-inflammatory cytokines (IL-12, IL1β, MCP-1 and IL-6) at approximately 18 months post-infection. These cytokines were present in trace amounts, such that they could only be detected with the use of novel nanotechnology. Importantly, these trace-level cytokines had a direct effect on the functionality of induced pluripotent stem cell-derived cardiomyocytes in vitro. This effect was not observed in the presence of dexamethasone. Plasma proteomics demonstrated further differences between PASC-CVS and Recovered patients at approximately 18 months post-infection including enrichment of complement and coagulation-associated proteins in those with prolonged cardiovascular symptoms. Together, these data provide a new insight into the role of chronic inflammation in PASC-CVS and presenting nanotechnology as a possible novel diagnostic approach for the condition. If chronic inflammation is driving disease progression in some patients, anti-inflammatory drugs may cure or at least reduce symptoms. While there are a few anti-inflammatory agents in clinical trials for this, the current best protection from long COVID is still prevention, which may be aided with vaccination and antivirals administered during acute infection. The use of vaccines as treatment for long COVID is also a possibility, however many studies on this have been limited by small cohort sizes and self-reporting methods, and results need to be validated. One reason that we still struggle to understand the effects of vaccination on long COVID is a lack of sufficient animal models. While a limited number of animal models have been generated to study post-COVID sequelae, these often require access to non-typical lab animals like hamsters or humanised mice, which can get very expensive and isn't easily scalable to large studies, and they largely focus on the neurological complications of SARS-CoV-2 infection. No scalable animal model to date has been established to reflect long COVID pulmonary complications, significantly impairing the development of novel therapeutics for this condition. In Chapter 3, Beta variant B.1.351 SARS-CoV-2 strain was mouseadapted via serial passaging and used to characterise a murine model of long COVID in which treatments might be tested. Obese female mice infected with 104 PFU of mouse-adapted virus showed systemic and pulmonary inflammation at 28 and 56 dpi, a phenotype not observed in younger male mice fed a standard chow diet. Interestingly, high-density microarray patch (HD-MAP) BetaHexaPro (-HP) vaccination at seven dpi partially attenuated inflammation in obese mice 28 dpi. These data present a practical murine model for pulmonary long COVID incorporating pertinent clinical risk factors, while HD-MAP delivery of the Beta-HP vaccine shows potential promise as a therapeutic strategy against long COVID. An additional issue with long COVID is that it was not considered when developing pathways out of the pandemic, and continues to be overlooked when making important public health decisions. The immense number of infections paired with the uncertain prevalence amongst those infected means that long COVID may profoundly impact the Australian population with post-infection disability and compromised quality-of-life, and many may require continuing medical care that can pressure the healthcare system. Thus, it is vital to develop a tool that may aid in evaluation of risk of long COVID development given SARS-CoV-2 infection. Such a tool should also consider the risks and benefits associated with receiving a COVID-19 vaccine to help individuals to make an informed decision on whether to get vaccinated, and for priming public health policy regarding the recommendation of different vaccine types for different population subgroups. Two Bayesian networks (BNs) were developed by integrating Australian and international data, 1) to calculate probabilities of outcomes for the delta (since updated to omicron) variant under different scenarios of Pfizer COVID-19 vaccine coverage, age groups (≥12 years), sex, community transmission intensity and vaccine effectiveness, and 2) to aid in assessing the risk of long-term adverse outcomes from COVID-19 under different scenarios of vaccine coverage and effectiveness, sex, age, comorbidities, number of previous SARSCoV-2 infections, and drug treatments administered during acute infection. The first model estimates that in a population where 5% were unvaccinated, 5% had one dose, 60% had two doses and 30% had three doses, there was a substantially greater probability of developing (239–5847 times) and dying (1430–384,684 times) from COVID-19-related than vaccine-associated myocarditis (depending on age and sex). For one million people with this vaccine coverage, where transmission intensity was equivalent to 10% chance of infection over 2 months, 68,813 symptomatic COVID-19 cases and 981 deaths would be prevented, with 42 and 16 expected cases of vaccine-associated myocarditis in males and females, respectively. These results justify vaccination in all age groups as vaccine-associated myocarditis is generally mild in the young, and there is unequivocal evidence for reduced mortality from COVID-19 in older individuals. Calculated outcomes from the second model show incomplete vaccination, missed opportunity for drug treatment during acute infection, and repeat infections to be the greatest controllable influences of an increased risk of long COVID. The models may easily be updated to include emerging best evidence, data pertinent to specific countries, vaccines, and outcomes. These models form the basis of the Pfizer vaccine and long COVID iterations of the COVID-19 Risk Calculator (CoRiCal) (https://corical.immunisationcoalition.org.au), a userfriendly online tool that enables scenario-analysis based on user inputs. These models (and their subsequent adaption to an online tool) can be used by individuals alone or in conjunction with clinicians for shared decision making on vaccination, pursuing early treatment during acute infection, and continuing infection-avoidant behaviors such as masking and social distancing. It may also assist health managers to assess such effects at a population level, contributing to better-informed public health policies. Overall, this study advances knowledge by elucidating the role of chronic inflammation in PASCCVS and presenting nanotechnology as a possible novel diagnostic approach for the condition, developing and characterising an accessible, inexpensive, and replicable non-humanised murine model of pulmonary long COVID in which therapeutic vaccination was tested, and developing models to aid in personal and public health decision-making on COVID-19 vaccination, pursuing early treatment during acute infection, and continuing infection-avoidant behaviors such as masking and social distancing. It is the hope of the authors that these findings may act as shoulders on which ongoing research may stand, into underlying disease mechanisms of, and diagnostic, prevention and treatment options for helping those with long COVID.
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