Aorta in Pathologies May Function as an Immune Organ by Upregulating Secretomes for Immune and Vascular Cell Activation, Differentiation and Trans-Differentiation-Early Secretomes may Serve as Drivers for Trained Immunity

Abstract

To determine whether aorta becomes immune organ in pathologies, we performed transcriptomic analyses of six types of secretomic genes (SGs) in aorta and vascular cells and made the following findings: 1) 53.7% out of 21,306 human protein genes are classified into six secretomes, namely, canonical, caspase 1, caspase 4, exosome, Weibel-Palade body, and autophagy; 2) Atherosclerosis (AS), chronic kidney disease (CKD) and abdominal aortic aneurysm (AAA) modulate six secretomes in aortas; and Middle East Respiratory Syndrome Coronavirus (MERS-CoV, COVID-19 homologous) infected endothelial cells (ECs) and angiotensin-II (Ang-II) treated vascular smooth muscle cells (VSMCs) modulate six secretomes; 3) AS aortas upregulate T and B cell immune SGs; CKD aortas upregulate SGs for cardiac hypertrophy, and hepatic fibrosis; and AAA aorta upregulate SGs for neuromuscular signaling and protein catabolism; 4) Ang-II induced AAA, canonical, caspase 4, and exosome SGs have two expression peaks of high (day 7)-low (day 14)-high (day 28) patterns; 5) Elastase induced AAA aortas have more inflammatory/immune pathways than that of Ang-II induced AAA aortas; 6) Most disease-upregulated cytokines in aorta may be secreted via canonical and exosome secretomes; 7) Canonical and caspase 1 SGs play roles at early MERS-CoV infected ECs whereas caspase 4 and exosome SGs play roles in late/chronic phases; and the early upregulated canonical and caspase 1 SGs may function as drivers for trained immunity (innate immune memory); 8) Venous ECs from arteriovenous fistula (AVF) upregulate SGs in five secretomes; and 9) Increased some of 101 trained immunity genes and decreased trained tolerance regulator IRG1 participate in upregulations of SGs in atherosclerotic, Ang-II induced AAA and CKD aortas, and MERS-CoV infected ECs, but less in SGs upregulated in AVF ECs. IL-1 family cytokines, HIF1α, SET7 and mTOR, ROS regulators NRF2 and NOX2 partially regulate trained immunity genes; and NRF2 plays roles in downregulating SGs more than that of NOX2 in upregulating SGs. These results provide novel insights on the roles of aorta as immune organ in upregulating secretomes and driving immune and vascular cell differentiations in COVID-19, cardiovascular diseases, inflammations, transplantations, autoimmune diseases and cancers.

Keywords: DAMPs; canonical and noncanonical secretomes; coronavirus infection; endothelial cell; inflammation.

Figure 1

The 53.7% out of 21,306 human protein genes can be classified into six secretomes. (A) Six secretomes, canonical (with signal peptide), caspase-1 (secretome mediated via N-terminal gasdermin D protein-formed channel when caspase-1 is activated), caspase-4 (secretome mediated via N-terminal gasdermin D protein-formed protein channel when caspase-4 (humans)/caspase-11 (mice) is activated) and exosomes, are the large secretomes with > 900 proteins, Weibel-Palade bodies (WPB), and autophagy secretome. (B) Venn Diagram Analysis was used to classify all the secretory protein genes into the secretome-shared genes and secretome-specific genes in six secretomes, which are ranked from the highest specificity to the lowest as exosome (83%) > canonical (68%) > caspase-1 (56%) > caspase-4 (49%) > autophagy (37.5%) > WPB (8%).

Figure 2

The three major aortic diseases including atherosclerosis, chronic kidney disease (CKD), and abdominal aortic aneurysm (AAA) significantly modulate the expression of six secretomes in aortas (A–C), suggesting that aorta is a new immune and endocrine organ in pathologies. Middle East Respiratory Syndrome Coronavirus (MERS-CoV) infection in human endothelial cells and Angiotensin-II (Ang-II) treatment in vascular smooth muscle cells also modulate the expression of six secretomes (A–C), suggesting that in response to danger associated molecular pattern (DAMPs, Ang-II) pathogens-associated molecular pattern (PAMPs, MERS-CoV) major aortic cell types such as endothelial cells and vascular smooth muscle cells modulate the expression of six secretomes. (D) Atherosclerosis and chronic kidney disease modulate the expression of secretomic genes much more than abdominal aortic aneurysm. (E) Angiotensin-II upregulates the expression of secretomic genes much more than downregulating the expression of secretomic genes where MERS-CoV infection downregulates more than upregulates the expression of secretomic genes. P < 0.05 |LogFC|>1. LogFC>1 or < -1.

Figure 3

Atherosclerosis (AS) aortas upregulate T and B cell immune secretomic genes (SGs); chronic kidney disease (CKD) aortas upregulate SGs for cardiac hypertrophy and hepatic fibrosis; and abdominal aortic aneurysm (AAA) aortas upregulate SGs for neuromuscular signaling and protein catabolism. (A) Ingenuity Pathway Analysis (IPA) analysis of upregulated genes in Fig. 2B. Atherosclerotic aortas upregulate expression of T cell and B cell adaptive immune secretomic genes; (B) chronic kidney disease (CKD) aortas upregulate expression of secretomic genes for cardiac hypertrophy, hepatic fibrosis, and senescence; and (C) Aortic aneurysm aortas upregulate expression of secretomic genes for neuromuscular signaling, protein catabolic process, and immunoglobulin Fc gamma (g) receptor-mediated phagocytosis. Z score>1.

Figure 4

Ang-II induced AAA, canonical, caspase 4, and exosome SGs in aortas have two expression peaks of high (Day 7)-low (Day 14)-high (Day 28) patterns suggesting the evidence of trained immunity response. (A) Canonical, caspase 4-GSDMD, and exosome pathways have two peak (three-phase) of high (Day 7)-low (Day 14)-high (Day 28) patterns; and caspase 1-GSDMD, Weibel-Palade body, and autophagy secretomes are only functional at the Day 7 AAA aortas. P < 0.05, LogFC>1 or < -1. (B) The signal pathways of secretomic genes are different in AAA progression. Z score>1.

Figure 5

Elastase induced AAA aortas have more inflammatory/immune pathways than that of Ang-II induced AAA aortas. (A) The signal pathways of secretomic genes are different in AAA progression. P < 0.05, |LogFC|>1. (B) IPA analysis of elastase induced AAA (GSE51229) in each secretory pathway. Z score>1. (C) The Venn diagram of upregulated pathways in Day 7 Ang II induced AAA and elastase induced AAA. Z score>1 .

Figure 6

Canonical secretome and caspase 1 secretome play more roles at early MERS-CoV (COVID-19 homoglous virus) infection of human microvascular endothelial cells whereas caspase 4 secretome and exosome secretome play more roles in the late and chronic phase infection; and these results have suggested novel therapeutic targets. (A) Canonical secretion and exosomes are most important secretory pathways in MERS condition. Number of differentially modulated genes in each pathway. P <0.05, |LogFC|>1. (B) MERS 36 hour is a transition time point in this process. Number of pathways which are regulated in MERS condition. (|Z score|>1 ). (C) Percentage of up/down gene number in each group based on (A). (D) Percentage of upregulated genes in each group. (E) Percentage of upregulated pathways in each group based on (B). (F) IPA analysis of upregulated genes in each group. Top 10 pathways are shown. Z score>1.

Figure 7

Venous ECs from arteriovenous fistula (AVF) upregulate SGs in five secretomes. (A) Experimental model of AVF with vascular anatomy (Red and Blue) and pathologic factors (Black). (B) Secretome analysis of venous endothelium during the formation of neointimal hyperplasia in a rat AVF model for kidney dialysis from GEO database (GSE46126) P <0.05 |LogFC|>1. (C) IPA analysis of each secretory pathway gene in AVF. Z score>1. (D, E) Venn diagram of upregulated canonical (D) and exosome (E) pathways in AVF.

Figure 8

Increased trained immunity genes and decreased trained tolerance regulator immune-responsive gene 1 (IRG1) participate in upregulations of SGs in atherosclerotic, Ang-II induced AAA and CKD aortas, and MERS-CoV infected Ecs. (A) Model of trained immunity. (B) Trained immunity gene list from http://www.ieom-tm.com/tidb/browse. (After remove 17 duplicated gene by Excel, totally 101 genes involved) IPA analysis of 101 trained immunity genes. (C) Number of upregulated trained immunity gene in different time course disease and condition. P < 0.05, LogFC>1. (D) The LogFC of IRG1 in different time course and condition.(P < 0.05). (E) The expression of IL-1 family genes in different condition. IL-1 family genes from PMID: 34844039, P < 0.05, LogFC was showed in table. (F) Number of downregulated secretomic genes in HIF1a-/- (GSE29765), SET7 SiRNA (GSE53038) and mTOR inhibitor (GSE138558) condition. P < 0.05, LogFC>1. (G) Working model of trained immunity.

Figure 9

Under pathological conditions, immune cells in aortas undergo maturation, activation, phenotypic switch and trans-differentiation, which justifies that aorta serves as an immune organ. (A) Literature based findings indicate that aorta may play a role as immune organ. (B) A new working model (created in BioRender.com) has indicated that similar to lymph nodes, a prototypic immune organ, which provide a niche for immune cell maturation, differentiation and activation, aortas sin pathologies serve as a novel immune organ for immune cells and vascular cells to get activated, matured, differentiated and trans-differentiated.