Bridging innate and adaptive immunity in cardiovascular disease

Cardiovascular disease is the leading cause of death and morbidity in the world. Myocardial infarction and stroke constitute the main manifestations of atherosclerosis that lead to the majority of cardiovascular events. Calcific aortic valve stenosis is the most common valve pathology. Atherosclerosis and aortic valve stenosis share common risk factors such as hypercholesterolemia. Lipid lowering treatment has ameliorated the incidence of fatal events; however, residual risk remains indicating the need to address the inflammatory component in cardiovascular disease. Many inflammatory mediators, such as cytokines and receptors have been implicated to play important role in the pathogenesis and the endpoints caused by an atherosclerotic plaque rupture. The role of pattern recognition receptors has been highlighted in several experimental studies. Both protective and detrimental effects have been described for the members of Tolllike receptor (TLR) family, a class of pattern recognition receptors. Several studies have focused on the role of the cell surface TLRs. The aim of the current thesis is to investigate the role of the intracellular pattern recognition receptor, TLR7. To gain information of the pathophysiological mechanisms that TLR7 is involved, both human cohorts of atherosclerosis and aortic valve stenosis as well as experimental models of atherosclerosis have been utilized.

In Paper I, mRNA expression of TLR7 in human carotid plaques was associated with patients´ outcome. Patients that expressed higher levels of TLR7 in their removed plaque had fewer future adverse cardio- and cerebrovascular events. Macrophages and T cells were co-localized with TLR7 in carotid plaques. Furthermore, carotid plaque tissue responded with increased cytokine secretion upon ex vivo stimulation with a synthetic TLR7 ligand. Paper II showed TLR7 mRNA expression in calcified aortic valves. TLR7 mRNA was increased in calcified areas of the aortic valves compare to intermediate and healthy areas. In addition, TLR7 expression was associated with M2 macrophage markers in all parts of the aortic valve. Stimulation of calcified aortic valves ex vivo with a synthetic TLR7 ligand elicited cytokine response that was possibly derived directly or indirectly by macrophages. In Paper III, we investigated the in vivo effects of a synthetic TLR7 ligand in experimental atherosclerosis. Locally, treatment with the synthetic TLR7 ligand led to decrease in lesion size and changes in plaque composition. The lesions of the treated mice presented lesions with smaller necrotic core and fewer apoptotic cells compare to the control. The treatment had effect in the spleen, leading to marginal zone B and regulatory T cell expansion. In the plasma, we observed decrease in cholesterol levels and increase in IgM antibodies against oxidized lowdensity lipoprotein.

The three studies presented in this thesis illustrate the protective role of TLR7 in atherosclerosis and aortic valve stenosis. TLR7 was expressed in both myeloid cells and lymphocytes indicating a role of the receptor in bridging innate and adaptive immune. The current results can encourage the investigation of TLR7 ligands as therapeutic intervention in cardiovascular disease.

List of scientific papers

I. Karadimou G, Folkersen L, Berg M, Perisic L, Discacciati A, Roy J, Hansson GK, Persson J and Paulsson-Berne G. Low TLR7 gene expression in atherosclerotic plaques is associated with major adverse cardio- and cerebrovascular events. Cardiovasc Res. 2017;113:30-39.
https://doi.org/10.1093/cvr/cvw231

II. Karadimou G, Persson O, Carracedo M, Eriksson P, Franco-Cereceda A, Paulsson-Berne G and Bäck M. TLR7 expression is associated with M2 macrophage subset in calcific aortic valve stenosis. [Manuscript]

III. Karadimou G, Gisterå A, Gallina AL, Caravaca AS, Centa M, Salagianni M, Andreakos E, Hansson GK, Malin S, Olofsson PS, Paulsson-Berne G. Treatment with a Toll-like Receptor 7 ligand evokes protective immunity against atherosclerosis in hypercholesterolemic mice. J Intern Med. May, 2020.
https://doi.org/10.1111/joim.13085