Proteomic profiling of plasma biomarkers in acute ischemic stroke due to large vessel occlusion

Abstract

Background: Acute ischemic stroke (AIS) due to large vessel occlusion (LVO) is a devastating cerebrovascular disorder, which could benefit from collateral circulation. Proteins associated with acute LVO pathogenesis and endothelial function may appear in blood samples of AIS patients due to LVO, thus permitting development of blood-based biomarkers for its diagnosis and prognosis.

Methods: This study is a single-center, retrospective, observational case-control trial. Consecutive patients who presented at the Department of Neurology of Tongji Hospital were recruited from July 2016 to April 2018. In the discovery phase, a proteomic approach with iTRAQ-based LC-MS/MS was used to investigate the altered proteomic pattern in plasma from patients with AIS due to LVO. In the validation study, Western blots was used to identify biomarkers associated with stroke diagnosis as well as their prognostic value associated with different collateral statuses.

Results: For this exploratory study, the proteomic analysis of plasma from 40 patients with AIS due to LVO and 20 healthy controls revealed seven differentially expressed proteins with a 1.2/0.83-fold or greater difference between groups. The four elevated proteins, PPBP (1.58 ± 0.78 vs 0.98 ± 0.37; P < 0.001), THBS1 (1.13 ± 0.88 vs 0.43 ± 0.26; P < 0.001), LYVE1 (1.61 ± 0.55 vs 0.97 ± 0.50; P < 0.001), and IGF2 (1.19 ± 0.42 vs 0.86 ± 0.24; P < 0.001), were verified by Western blots analysis in an independent cohort including 33 patients and 33 controls. A strong interaction was observed between the four-protein panel and the diagnosis of AIS due to LVO (AUC 0.947; P < 0.001). Furthermore, IGF2, LYVE1, and THBS1 were closely associated with collateral status (IGF2 0.115, 95% CI 0.016-0.841, P = 0.033; LYVE1 0.183, 95% CI 0.036-0.918, P = 0.039; THBS1 4.257, 95% CI 1.273-14.228, P = 0.019), and proved to be independent predictors of good outcome (IGF2 0.115, 95% CI 0.015-0.866, P = 0.036; LYVE1 0.028, 95% CI 0.002-0.334, P = 0.005; THBS1 3.294, 95% CI 1.158-9.372, P = 0.025) at a 3-month follow-up.

Conclusions: The identified 4-biomarker panel could provide diagnostic aid to the existing imaging modalities for AIS due to LVO, and the prognostic value of IGF2, LYVE1, and THBS1 was proved in predicting functional outcomes related to collateral status. Trial registration ClinicalTrials.gov NCT03122002. Retrospectively registered April 20, 2017. URL of trial registry record: https://www.clinicaltrials.gov/ct2/show/NCT03122002?term=NCT+03122002&rank=1.

Keywords: Biomarkers; Collateral circulation; Ischemic stroke; Large vessel occlusion; Proteomics.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1

Flow diagram of the study design. Discovery phase: proteomic analysis of AIS patients due to LVO (4 groups, 10 individuals pooled in a group) versus healthy controls (2 groups, 10 individuals pooled in a group). Validation phase: Western blot analysis of selected plasma biomarkers in healthy controls (n = 33), patients with AIS due to non-LVO (n = 33), and patients with AIS due to LVO (n = 33). Patients with AIS due to LVO were further divided into two subgroups, AIS with good collaterals (n = 17) and poor collaterals (n = 16)

Fig. 2

Validation phase of potential biomarkers by Western blots analysis. a Western blots representation of each potential biomarker. b Quantitative analysis was performed. *P < 0.05 **P  < 0.01 versus Control, †P < 0.05 ††P < 0.01 versus AIS due to non-LVO, n = 33 for Control, n = 33 for AIS due to non-LVO group and n = 33 for AIS due to LVO group. ‡P < 0.05 ‡‡P < 0.01 versus SGC, n = 17 for SGC, n = 16 for SPC. Control healthy controls. AIS acute ischemic stroke patients, SGC stroke with good collaterals, SPC stroke with poor collaterals

Fig. 3

Diagnostic efficiency of the potential biomarkers. a Receiver operating characteristic curve of biomarkers in diagnosis of AIS due to LVO. IGF2, LYVE1, PPBP and THBS1 could be efficient in diagnosis, and b 4-protein panel combined has better performance