Mosaic Loss of Chromosome Y Is Associated With Functional Outcome After Ischemic Stroke

BACKGROUND: Mosaic loss of chromosome Y (LOY) is associated with cardiovascular and neurodegenerative diseases in men, and genetic predisposition to LOY is associated with poor poststroke outcome. We, therefore, tested the hypothesis that LOY itself is associated with functional outcome after ischemic stroke. METHODS: The study comprised male patients with ischemic stroke from the cohort studies SAHLSIS2 (Sahlgrenska Academy Study on Ischemic Stroke Phase 2; n=588) and LSR (Lund Stroke Register; n=735). We used binary logistic regression to analyze associations between LOY, determined by DNA microarray intensity data, and poor 3-month functional outcome (modified Rankin Scale score, >2) in each cohort separately and combined. Patients who received recanalization therapy were excluded from sensitivity analyses. RESULTS: LOY was associated with about 2.5-fold increased risk of poor outcome in univariable analyses (P<0.001). This association withstood separate adjustment for stroke severity and diabetes in both cohorts but not age. In sensitivity analyses restricted to the nonrecanalization group (n=987 in the combined cohort), the association was significant also after separate adjustment for age (odds ratio, 1.6 [95% CI, 1.1–2.4]) and when additionally adjusting for stroke severity and diabetes (odds ratio, 1.6 [95% CI, 1.1–2.5]). CONCLUSIONS: We observed an association between LOY and poor outcome after ischemic stroke in patients not receiving recanalization therapy. Future studies on LOY and other somatic genetic alterations in larger stroke cohorts are warranted.


Methods
Anonymized data will be shared upon reasonable request from a qualified academic investigator as long as data transfer agrees with EU legislation on the general data protection regulation (GDPR) and with decisions by the Ethical Review Board of Sweden and the University of Gothenburg, the latter which should be regulated in a data transfer agreement.

Study populations
The ongoing Sahlgrenska Academy Study on Ischemic Stroke phase 2 (SAHLSIS2) and Lund Stroke Register (LSR) study consecutively include adult first-ever or recurrent stroke patients at stroke units at the Sahlgrenska University Hospital in Gothenburg and first-ever stroke patients from the local catchment area and hospitalized at Skåne University Hospital in Lund, respectively, both Sweden. In the present study male participants with acute ischemic stroke recruited between 2015 and 2020 in SAHLSIS2, and between 2013 and 2020 in LSR, were included. In line with the World Health Organization stroke definition, ischemic stroke was diagnosed in patients with rapidly developing clinical signs of focal disturbance of cerebral function lasting >24 hours and without hemorrhage or signs of another cause on neuroimaging. All patients included in this study underwent computed tomography (CT) and/or magnetic resonance (MR) imaging of the brain as part of the clinical routine investigation. If clinically indicated, patients were also evaluated by CT or MR angiography and perfusion measures. Patients received recanalization therapy with intravenous thrombolysis only (IVT, n=97 in SAHLSIS2 and n=97 in LSR) or endovascular therapy with or without bridging IVT (n=92 in SAHLSIS2 and n=45 in LSR) according to national guidelines issued by the Swedish Board of Health and Welfare in 2015, and before that according to local hospital guidelines. Data regarding these treatments were registered at each hospital and reported to national quality registers. The patients also underwent additional work-up according to national guidelines, and patients were excluded from this study if further evaluation showed another etiology of the presenting symptoms than ischemic stroke. Written informed consent, or consultant consent from next-of-kin, was obtained prior to enrollment. The studies were approved by the respective local Regional Ethics Review Board (Etikprövningsnämnden; EPN) or the Swedish Ethical Review Authority (Etikprövningsmyndigheten; EPM). The registration numbers for SAHLSIS2 are EPN Dnr 823-13 and 823-13 T1110-16 and EPM Dnr 2022-00012-02. For LSR the registration numbers are EPN Dnr 2016/179 and 2016/999.

Clinical variables and outcome
In Sweden, all hospitals treating patients with acute stroke report comprehensive data on for instance demographics, vascular risk factors, acute treatments and severity of the stroke, and outcomes to the national quality register for stroke, Riksstroke. Data are entered into this register by licensed healthcare personnel at each hospital, and the coverage rate is about 95% (www.riksstroke.org/general-information/). For both SAHLSIS2 and LSR data on diabetes mellitus, hypertension, smoking at baseline and mortality within 3 months were obtained from Riksstroke. We also used data from Riksstroke on self-reported dependency in specified domains (mobility, dressing, and toileting), living conditions, and need of help or support from next of kin 3 months post-stroke to estimate the modified Rankin Scale (mRS) scores 0-2, 3, 4 or 5 using a validated translation algorithm. Good outcome was defined as independency (mRS 0-2) and poor outcome as dependency or death (mRS 3-6). For SAHLSIS2 data on the admission stroke severity, i.e. National Institutes of Health Stroke Scale (NIHSS) score, was obtained from Riksstroke. For LSR the initial NIHSS score from admission, as close to day 1 as possible, was registered.

Single nucleotide polymorphism (SNP) intensity data
The Illumina Infinium Global Screening Array-24 v3.0 BeadChip was used for genotyping of both SAHLSIS2 and LSR. The array contains 5,006 single nucleotide polymorphisms in the male specific part of the Y chromosome. The microarray intensity data of the Y chromosome were normalized against cluster files created from male samples only. The pre-processing of the data was accomplished using GenomeStudio, and clustering was performed based on the signal intensities of the male specific part of the Y chromosome. Samples with a call rate lower than 98% and SNPs with a gene call score < 0.15 were excluded. The normalization was performed at the SNP&SEQ Technology Platform in Uppsala, and a Nexus plugin was used to generate the copy number variation (CNV) calls.

Statistics
For the final multivariable regression model, we used the R-package missForest (v 1.5) to impute missing data from the clinical variables NIHSS scores and diabetes mellitus in order to retain power in the final model.

Results
Participants 13* (a) Report numbers of individuals at each stage of study-eg numbers potentially eligible, examined for eligibility, confirmed eligible, included in the study, completing follow-up, and analysed 4 and   Tables   Table S1. Baseline characteristics, LOY and functional outcome (mRS score) for male ischemic stroke patients in the non-recanalization therapy and the recanalization therapy groups in the exploratory (SAHLSIS2) and validation (LSR) cohorts.