Comprehensive compartmental model and calibration algorithm for the study of clinical implications of the population-level spread of COVID-19: a study protocol.

INTRODUCTION: The complex dynamics of the coronavirus disease 2019 (COVID-19) pandemic has made obtaining reliable long-term forecasts of the disease progression difficult. Simple mechanistic models with deterministic parameters are useful for short-term predictions but have ultimately been unsuccessful in extrapolating the trajectory of the pandemic because of unmodelled dynamics and the unrealistic level of certainty that is assumed in the predictions. METHODS AND ANALYSIS: We propose a 22-compartment epidemiological model that includes compartments not previously considered concurrently, to account for the effects of vaccination, asymptomatic individuals, inadequate access to hospital care, post-acute COVID-19 and recovery with long-term health complications. Additionally, new connections between compartments introduce new dynamics to the system and provide a framework to study the sensitivity of model outputs to several concurrent effects, including temporary immunity, vaccination rate and vaccine effectiveness. Subject to data availability for a given region, we discuss a means by which population demographics (age, comorbidity, socioeconomic status, sex and geographical location) and clinically relevant information (different variants, different vaccines) can be incorporated within the 22-compartment framework. Considering a probabilistic interpretation of the parameters allows the model's predictions to reflect the current state of uncertainty about the model parameters and model states. We propose the use of a sparse Bayesian learning algorithm for parameter calibration and model selection. This methodology considers a combination of prescribed parameter prior distributions for parameters that are known to be essential to the modelled dynamics and automatic relevance determination priors for parameters whose relevance is questionable. This is useful as it helps prevent overfitting the available epidemiological data when calibrating the parameters of the proposed model. Population-level administrative health data will serve as partial observations of the model states. ETHICS AND DISSEMINATION: Approved by Carleton University's Research Ethics Board-B (clearance ID: 114596). Results will be made available through future publication.

Rac1 plays a role in CXCL12 but not CCL3-induced chemotaxis and Rac1 GEF inhibitor NSC23766 has off target effects on CXCR4.

Cell migration towards a chemotactic stimulus relies on the re-arrangement of the cytoskeleton, which is triggered by activation of small G proteins RhoA, Rac1 and Cdc42, and leads to formation of lamellopodia and actin polymerisation amongst other effects. Here we show that Rac1 is important for CXCR4 induced chemotaxis but not for CCR1/CCR5 induced chemotaxis. For CXCL12-induced migration via CXCR4, breast cancer MCF-7 cells are reliant on Rac1, similarly to THP-1 monocytes and Jurkat T-cells. For CCL3-induced migration via CCR1 and/or CCR5, Rac1 signalling does not regulate cell migration in either suspension or adherent cells. We have confirmed the involvement of Rac1 with the use of a specific Rac1 blocking peptide. We also used a Rac1 inhibitor EHT 1864 and a Rac1-GEF inhibitor NSC23766 to probe the importance of Rac1 in chemotaxis. Both inhibitors did not block CCL3-induced chemotaxis, but they were able to block CXCL12-induced chemotaxis. This confirms that Rac1 activation is not essential for CCL3-induced migration, however NSC23766 might have secondary effects on CXCR4. This small molecule exhibits agonistic features in internalisation and cAMP assays, whereas it acts as an antagonist for CXCR4 in migration and calcium release assays. Our findings strongly suggest that Rac1 activation is not necessary for CCL3 signalling, and reveal that NSC23766 could be a novel CXCR4 receptor ligand.

Cell migration towards CXCL12 in leukemic cells compared to breast cancer cells.

Chemotaxis or directed cell migration is mediated by signalling events initiated by binding of chemokines to their cognate receptors and the activation of a complex signalling cascade. The molecular signalling pathways involved in cell migration are important to understand cancer cell metastasis. Therefore, we investigated the molecular mechanisms of CXCL12 induced cell migration and the importance of different signalling cascades that become activated by CXCR4 in leukemic cells versus breast cancer cells. We identified Src kinase as being essential for cell migration in both cancer types, with strong involvement of the Raf/MEK/ERK1/2 pathway. We did not detect any involvement of Ras or JAK2/STAT3 in CXCL12 induced migration in Jurkat cells. Preventing PKC activation with inhibitors does not affect migration in Jurkat cells at all, unlike in the adherent breast cancer cell line MCF-7 cells. However, in both cell lines, knock down of PKCα prevents migration towards CXCL12, whereas the expression of PKCζ is less crucial for migration. PI3K activation is essential in both cell types, however LY294002 usage in MCF-7 cells does not block migration significantly. These results highlight the importance of verifying specific signalling pathways in different cell settings and with different approaches.

Dynamin function is important for chemokine receptor-induced cell migration.

The HIV viral entry co-receptors CCR5 and CXCR4 function physiologically as typical chemokine receptors. Activation leads to cytosolic signal transduction that results in a variety of cellular responses such as cytoskeletal rearrangement and chemotaxis (CTX). Our aim was to investigate the signalling pathways involved in CC and CXC receptor-mediated cell migration. Inhibition of dynamin I and II GTPase with dynasore completely inhibited CCL3-stimulated CTX in THP-1 cells, whereas the dynasore analogue Dyngo-4a, which is a more potent inhibitor, showed reduced ability to inhibit CC chemokine-induced CTX. In contrast, dynasore was not able to block cell migration via CXCR4. The same activation/inhibition pattern was verified in activated T lymphocytes for different CC and CXC chemokines. Cell migration induced by CC and CXC receptors does not rely on active internalization processes driven by dynamin because the blockade of internalization does not affect migration, but it might rely on dynamin interaction with the cytoskeleton. We identify here a functional difference in how CC and CXC receptor migration is controlled, suggesting that specific signalling networks are being employed for different receptor classes and potentially specific therapeutic targets to prevent receptor migration can be identified.