Glycoproteomic Analysis of the Aortic Extracellular Matrix in Marfan Patients.

Xiaoke Yin ,Carlie J.M De Vries,Marjan Jahangiri,Dieter P Reinhardt,Shaynah Wanga,Philipp Skroblin,Vivian De Waard,Manuel Mayr,Rosa Viner,David R Koolbergen,Marika Fava,Javier Barallobre-Barreiro,Sanjay Sinha,Aeilko H Zwinderman,Barbara J.M Mulder,Ron Balm,Qiuru Xing,Ferheen Baig,Ruifang Lu,Hongorzul Davaapil,Maarten Groenink,Romy Franken,Adam Fellows

doi:10.1161/atvbaha.118.312175

Abstract

Marfan syndrome (MFS) is caused by mutations in FBN1 (fibrillin-1), an extracellular matrix (ECM) component, which is modified post-translationally by glycosylation. This study aimed to characterize the glycoproteome of the aortic ECM from patients with MFS and relate it to aortopathy. Approach and Results: ECM extracts of aneurysmal ascending aortic tissue from patients with and without MFS were enriched for glycopeptides. Direct N-glycopeptide analysis by mass spectrometry identified 141 glycoforms from 47 glycosites within 35 glycoproteins in the human aortic ECM. Notably, MFAP4 (microfibril-associated glycoprotein 4) showed increased and more diverse N-glycosylation in patients with MFS compared with control patients. MFAP4 mRNA levels were markedly higher in MFS aortic tissue. MFAP4 protein levels were also increased at the predilection (convexity) site for ascending aorta aneurysm in bicuspid aortic valve patients, preceding aortic dilatation. In human aortic smooth muscle cells, MFAP4 mRNA expression was induced by TGF (transforming growth factor)-β1 whereas siRNA knockdown of MFAP4 decreased FBN1 but increased elastin expression. These ECM changes were accompanied by differential gene expression and protein abundance of proteases from ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family and their proteoglycan substrates, respectively. Finally, high plasma MFAP4 concentrations in patients with MFS were associated with a lower thoracic descending aorta distensibility and greater incidence of type B aortic dissection during 68 months follow-up. Our glycoproteomics analysis revealed that MFAP4 glycosylation is enhanced, as well as its expression during the advanced, aneurysmal stages of MFS compared with control aneurysms from patients without MFS.

Highlights

MethodsThe data that support the findings of this study are available from the corresponding author upon reasonable request.Sample Collection From Patients With Marfan syndrome (MFS)For glycoproteomic analysis, aortic aneurysmal tissue was collected during surgery from 11 patients with MFS and 6 patients without MFS who had undergone aortic root and ascending aorta replacement (Table I in the online-only Data Supplement)
Xiaoke Yin,* Shaynah Wanga,* Adam Fellows, Javier Barallobre-Barreiro, Ruifang Lu, Hongorzul Davaapil, Romy Franken, Marika Fava, Ferheen Baig, Philipp Skroblin, Qiuru Xing, David R Koolbergen, Maarten Groenink, Aeilko H Zwinderman, Ron Balm, Carlie J.M de Vries, Barbara J.M Mulder, Rosa Viner, Marjan Jahangiri, Dieter P Reinhardt, Sanjay Sinha, Vivian de Waard,* Manuel Mayr*
In human aortic smooth muscle cells, MFAP4 mRNA expression was induced by TGF-β1 whereas siRNA knockdown of MFAP4 decreased FBN1 but increased elastin expression

Summary

Methods

The data that support the findings of this study are available from the corresponding author upon reasonable request.Sample Collection From Patients With MFSFor glycoproteomic analysis, aortic aneurysmal tissue was collected during surgery from 11 patients with MFS and 6 patients without MFS who had undergone aortic root and ascending aorta replacement (Table I in the online-only Data Supplement). The same patients with MFS, who were from a subset of the COMPARE trial (NTR1423)[20] and included in a study on plasma TGF-β1 in aortic disease,[21,22] were used to measure plasma MFAP4 concentration. The exclusion criteria were angiotensinconverting enzyme inhibitor usage and previous replacement of more than one part of the aorta. In this substudy, all patients were included irrespective of the medication they used, and patients with prior aortic surgery were allowed.[20] At the time of inclusion, plasma samples (n=96) from patients with MFS were obtained, and aortic root diameters were measured. Written consent was obtained from all patients included in this study

Results

Discussion

Conclusion