The Role of Toll-like Receptor-4 in Macrophage Imbalance in Lethal COVID-19 Lung Disease, and Its Correlation with Galectin-3.

To the current data, there have been 6,955,141 COVID-19-related deaths worldwide, reported to WHO. Toll-like receptors (TLRs) implicated in bacterial and virus sensing could be a crosstalk between activation of persistent innate-immune inflammation, and macrophage's sub-population alterations, implicated in cytokine storm, macrophage over-activation syndrome, unresolved Acute Respiratory Disease Syndrome (ARDS), and death. The aim of this study is to demonstrate the association between Toll-like-receptor-4 (TLR-4)-induced inflammation and macrophage imbalance in the lung inflammatory infiltrate of lethal COVID-19 disease. Twenty-five cases of autopsy lung tissues were studied by digital pathology-based immunohistochemistry to evaluate expression levels of TLR-4 (CD 284), pan-macrophage marker CD68 (clone KP1), sub-population marker related to alveolar macrophage Galectin-3 (GAL-3) (clone 9C4), and myeloid derived CD163 (clone MRQ-26), respectively. SARS-CoV-2 viral persistence has been evaluated by in situ hybridation (ISH) method. This study showed TLR-4 up-regulation in a subgroup of patients, increased macrophage infiltration in both Spike-1(+) and Spike-1(-) lungs (p < 0.0001), and a macrophage shift with important down-regulation of GAL-3(+) alveolar macrophages associated with Spike-1 persistence (p < 0.05), in favor of CD163(+) myeloid derived monocyte-macrophages. Data show that TLR-4 expression induces a persistent activation of the inflammation, with inefficient resolution, and pathological macrophage shift, thus explaining one of the mechanisms of lethal COVID-19.

The Role of TLR-2 in Lethal COVID-19 Disease Involving Medullary and Resident Lung Megakaryocyte Up-Regulation in the Microthrombosis Mechanism.

Patients with COVID-19 have coagulation and platelet disorders, with platelet alterations and thrombocytopenia representing negative prognostic parameters associated with severe forms of the disease and increased lethality. METHODS: The aim of this study was to study the expression of platelet glycoprotein IIIa (CD61), playing a critical role in platelet aggregation, together with TRL-2 as a marker of innate immune activation. RESULTS: A total of 25 patients were investigated, with the majority (24/25, 96%) having co-morbidities and dying from a fatal form of SARS-CoV-2(+) infection (COVID-19+), with 13 men and 12 females ranging in age from 45 to 80 years. When compared to a control group of SARS-CoV-2 (-) negative lungs (COVID-19-), TLR-2 expression was up-regulated in a subset of patients with deadly COVID-19 fatal lung illness. The proportion of Spike-1 (+) patients found by PCR and ISH correlates to the proportion of Spike-S1-positive cases as detected by digital pathology examination. Furthermore, CD61 expression was considerably higher in the lungs of deceased patients. In conclusion, we demonstrate that innate immune prolonged hyperactivation is related to platelet/megakaryocyte over-expression in the lung. CONCLUSIONS: Microthrombosis in deadly COVID-19+ lung disease is associated with an increase in the number of CD61+ platelets and megakaryocytes in the pulmonary interstitium, as well as their functional activation; this phenomenon is associated with increased expression of innate immunity TLR2+ cells, which binds the SARS-CoV-2 E protein, and significantly with the persistence of the Spike-S1 viral sequence.