Circulating anti-β2-glycoprotein I antibodies of peripheral arterial disease patients trigger a genomic overexpression of Toll-like receptor 4 in endothelial cells.
OBJECTIVE
Circulating anti-β2-glycoprotein I (ABGPI) antibodies are associated with peripheral arterial disease (PAD) and induce the expression of leukocyte adhesion molecules and proinflammatory cytokines by endothelial cells. Our aim is to study a transcriptional activation pathway of the innate immune system through the cellular signalling cascade triggered by receptors Toll-like receptor 2 (TLR2) and Toll-like receptor 4 (TLR4) of endothelial cells after the exposure of these cells to seropositive ABGPI human serum obtained from PAD patients.


METHODS
We obtained serum samples from PAD patients and controls without PAD. ABGPI serum titer was detected using indirect immunofluorescence. Our sample was stratified into three groups: group I (PAD and ABGPI titer ≥1:100; n = 15), group II (PAD and ABGPI titer <1:100; n = 15), and control participants (no PAD; n = 15). All serum samples were incubated with human aortic endothelial cell (HAEC) culture. Genomic expression of TLR2 and TLR4 receptors and their shared intracellular signalling molecules, myeloid differentiation primary response gene 88 (MyD88), and interleukin (IL)-1 receptor-associated kinase (1IRAK1), were measured after the exposure of HAECs to each serum.


RESULTS
HAEC genomic expression of TLR4 was higher after the exposure to group I serum than after the exposure to group II serum (log10×10-relative quantification [RQ]: 1.80 ± 0.42 vs 1.37 ± 0.39; P = .01) or control serum (log10×10-RQ: 1.80 ± 0.42 vs 1.09 ± 0.26; P < .01). TLR4 expression was higher in group II than in the control group (log10×10-RQ: 1.37 ± 0.39 vs 1.09 ± 0.26; P = .04). TLR4 expression correlated with MyD88 (r = 0.54; P < .01) and IRAK1 (r = 0.55; P < .01) expression. We recorded a positive correlation between MyD88 and IRAK1 genomic expression (r = 0.58; P < .01).


CONCLUSIONS
Our results suggest that serum from PAD patients with elevated ABGPI antibodies induces a genomic overexpression of TLR4 and its cellular signalling molecules in endothelial cells.
Department of Angiology and Vascular Surgery, Hospital Universitario de Getafe, Madrid, Spain. Electronic address: varelot@hotmail.com.
Cesar
C
Varela
Department of Angiology and Vascular Surgery, Hospital Universitario de Getafe, Madrid, Spain.
Joaquin
J
de Haro
Department of Angiology and Vascular Surgery, Hospital Universitario de Getafe, Madrid, Spain.
Silvia
S
Bleda
Department of Angiology and Vascular Surgery, Hospital Universitario de Getafe, Madrid, Spain.
Javier
J
Rodriguez-Padilla
Department of Research, Hospital Universitario de Getafe, Madrid, Spain.
Antonio
A
Ferruelo
Department of Angiology and Vascular Surgery, Hospital Universitario de Getafe, Madrid, Spain.
Francisco
F
Acín
D000368:Aged; D001323:Autoantibodies / Q000097:blood*; D016022:Case-Control Studies; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000276:immunology / Q000378:metabolism*; D005260:Female; D006801:Humans; D007113:Immunity, Innate; D053592:Interleukin-1 Receptor-Associated Kinases / Q000235:genetics / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D053594:Myeloid Differentiation Factor 88 / Q000235:genetics / Q000378:metabolism; D058729:Peripheral Arterial Disease / Q000097:blood* / Q000276:immunology; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014158:Transcription, Genetic; D015854:Up-Regulation; D053482:beta 2-Glycoprotein I / Q000276:immunology*
D001323:Autoantibodies; C507779:MYD88 protein, human; D053594:Myeloid Differentiation Factor 88; D012333:RNA, Messenger; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D053482:beta 2-Glycoprotein I; C507768:IRAK1 protein, human; D053592:Interleukin-1 Receptor-Associated Kinases