Serum Proteome Alterations in Human Cystathionine β-Synthase Deficiency and Ischemic Stroke Subtypes

Abstract

Ischemic stroke induces brain injury via thrombotic or embolic mechanisms involving large or small vessels. Cystathionine β-synthase deficiency (CBS), an inborn error of metabolism, is associated with vascular thromboembolism, the major cause of morbidity and mortality in affected patients. Because thromboembolism involves the brain vasculature in these patients, we hypothesize that CBS deficiency and ischemic stroke have similar molecular phenotypes. We used label-free mass spectrometry for quantification of changes in serum proteomes in CBS-deficient patients (n = 10) and gender/age-matched unaffected controls (n = 14), as well as in patients with cardioembolic (n = 17), large-vessel (n = 26), or lacunar (n = 25) ischemic stroke subtype. In CBS-deficient patients, 40 differentially expressed serum proteins were identified, of which 18 were associated with elevated homocysteine (Hcy) and 22 were Hcy-independent. We also identified Hcy-independent differentially expressed serum proteins in ischemic stroke patients, some of which were unique to a specific subtype: 10 of 32 for cardioembolic vs. large-vessel, six of 33 for cardioembolic vs. lacunar, and six of 23 for large-vessel vs. lacunar. There were significant overlaps between proteins affected by CBS deficiency and ischemic stroke, particularly the cardioembolic subtype, similar to protein overlaps between ischemic stroke subtypes. Top molecular pathways affected by CBS deficiency and ischemic stroke subtypes included acute phase response signaling and coagulation system. Similar molecular networks centering on NFκB were affected by CBS deficiency and stroke subtypes. These findings suggest common mechanisms involved in the pathologies of CBS deficiency and ischemic stroke subtypes.

Cystathionine β-synthase deficiency: different changes in proteomes of thrombosis-resistant Cbs−/− mice and thrombosis-prone CBS−/− humans

“…Total Hcy levels in CBS −/− patients and Cbs −/− mice were severely elevated compared to corresponding unaffected controls (71.2 ± 55.5 vs. 9.7 ± 5.9 μM in human plasma 12 ; 236 ± 80 vs. 10.2 ± 5.3 μM in mouse plasma), as expected 13,14 . In each group of subjects, label-free nanoLC-MS/MS mass spectrometry identified 196-198 proteins in human and mouse plasma with a minimum of two peptides and 1% false discovery rate (FDR).…”

“…We identified 117 proteins differentiating between Cbs −/− vs. Cbs +/+ in mice in the present study while 41 proteins differentiating between CBS −/− vs. CBS +/+ were identified in humans 12 (Fig. 2).…”

“…To identify biological pathways linked to proteins affected by Cbs deficiency in mice and CBS deficiency in humans, we performed bioinformatics analyses using IPA resources. Analyses for the human CBS deficiency have been described before 12 . We have found that in the mouse Cbs deficiency, proteins from 19 pathways were overrepresented among differentially expressed proteins, while proteins from 17 pathways were enriched in the human.…”

“…Graphical illustrations of interactions between proteins in these networks are shown in Fig. 5A-D. Data for human plasma are from Sikora et al 12 .…”

“…Different susceptibilities of CBS −/− patients and Cbs −/− mice to thrombosis might be reflected in their proteomes. Previous work has identified CBS deficiency-responsive proteins in human plasma 12 . In the present work we have used label-free mass spectrometry and the Ingenuity Pathway Analysis resources to interrogate plasma proteomes of thrombosis-resistant Cbs −/− mice and compared them with plasma proteomes of thrombosis-prone CBS −/− humans.…”