Endogenous Deoxyribonuclease Activity and Cell-Free Deoxyribonucleic Acid in Acute Ischemic Stroke: A Cohort Study.

BACKGROUND: Cell-free DNA (cfDNA) and endogenous deoxyribonuclease activity are opposing mediators and might influence the inflammatory response following acute ischemic stroke. In this cohort study, we investigated the relation between these markers, circulating inflammatory mediators and clinical course including occurrence of stroke-associated infections (SAI) in patients with acute stroke. METHODS: Ninety-two patients with stroke due to large vessel occlusion undergoing mechanical thrombectomy were prospectively recruited at Hannover Medical School from March 2018 to August 2019. Deoxyribonuclease activity, cfDNA, damage-associated molecular patterns, and circulating cytokines were measured in venous blood collected immediately before mechanical thrombectomy and 7 days later. Reperfusion status was categorized (sufficient/insufficient). Clinical outcome was evaluated using the modified Rankin Scale after 90 days, where a score of 3 to 6 was considered unfavorable. To validate findings regarding SAI, another stroke cohort (n=92) was considered with blood taken within 24 hours after stroke onset. RESULTS: Patients with unfavorable clinical outcome had higher cfDNA concentrations. After adjustment for confounders (Essen Stroke Risk Score, National Institutes of Health Stroke Scale, and sex), 7-day cfDNA was independently associated with clinical outcome and especially mortality (adjusted odds ratio: 3.485 [95% CI, 1.001-12.134] and adjusted odds ratio: 9.585 [95% CI, 2.006-45.790]). No association was found between reperfusion status and cfDNA or deoxyribonuclease activity. While cfDNA concentrations correlated positively, deoxyribonuclease activity inversely correlated with distinct biomarkers. Baseline deoxyribonuclease activity was lower in patients who developed SAI compared with patients without SAI. This association was confirmed after adjustment for confounding factors (adjusted odds ratio: 0.447 [95% CI, 0.237-0.844]). In cohort 2, differences of deoxyribonuclease activity between patients with and without SAI tended to be higher with higher stroke severity. CONCLUSIONS: The interplay of endogenous deoxyribonuclease activity and cfDNA in acute stroke entails interesting novel diagnostic and potential therapeutic approaches. We confirm an independent association of cfDNA with a detrimental clinical course after stroke due to large vessel occlusion. This study provides first evidence for lower endogenous deoxyribonuclease activity as risk factor for SAI after severe stroke.

Circulating Inflammatory Biomarkers in Early Prediction of Stroke-Associated Infections.

(1) Background: Patients with acute ischaemic stroke (AIS) are at high risk for stroke-associated infections (SAIs). We hypothesised that increased concentrations of systemic inflammation markers predict SAIs and unfavourable outcomes; (2) Methods: In 223 patients with AIS, blood samples were taken at ≤24 h, 3 d and 7d after a stroke, to determine IL-6, IL-10, CRP and LBP. The outcome was assessed using the modified Rankin Scale at 90 d. Patients were thoroughly examined regarding the development of SAIs; (3) Results: 47 patients developed SAIs, including 15 lower respiratory tract infections (LRTIs). IL-6 and LBP at 24 h differed, between patients with and without SAIs (IL-6: p < 0.001; LBP: p = 0.042). However, these associations could not be confirmed after adjustment for age, white blood cell count, reduced consciousness and NIHSS. When considering the subgroup of LRTIs, in patients who presented early (≤12 h after stroke, n = 139), IL-6 was independently associated with LRTIs (OR: 1.073, 95% CI: 1.002−1.148). The ROC-analysis for prediction of LRTIs showed an AUC of 0.918 for the combination of IL-6 and clinical factors; (4) Conclusions: Blood biomarkers were not predictive for total SAIs. At early stages, IL-6 was independently associated with outcome-relevant LRTIs. Further studies need to clarify the use of biochemical markers to identify patients prone to SAIs.

Autoantibodies against the NMDAR subunit NR1 are associated with neuropsychiatric outcome after ischemic stroke.

BACKGROUND AND PURPOSE: Preexisting autoantibodies against N-methyl-D-aspartate-receptor subunit NR1 (NMDAR1-AB) in acute ischemic stroke patients with previously intact blood-brain-barrier were associated with smaller evolution of lesion size. Effects of chronic exposure to NMDAR1-AB long after stroke, however, have remained unclear. We investigated in a prospective follow-up study whether long-term neuropsychiatric outcome after stroke differs depending on NMDAR1-AB status. METHODS: Blood samples for NMDAR1-AB analysis were collected within 24 h after ischemic stroke from n = 114 patients. Outcome was assessed 1-3 years later using NIHSS, modified Rankin-scale, Barthel-Index, RBANS (Repeatable Battery for the Assessment of Neuropsychological Status) subcategories (immediate/delayed memory, attention, visuoconstruction), anamnesis evaluating neuropsychiatric symptoms (e.g. hallucinations, psychomotor slowing, reduced alertness, depressiveness, fatigue) and questionnaires (Beck's Depression Inventory-BDI, Fatigue Impact Scale-FIS). Scores were generated to cover RBANS plus neuropsychiatric symptoms (Score A; n = 96) or only neuropsychiatric symptoms (Score B; n = 114, including patients unable to conduct RBANS). Depression/fatigue were measured in patients, capable to perform questionnaires (n = 86). RESULTS: NMDAR1-AB (IgM, IgA, IgG) were detected in n = 27 patients (23.7%). NMDAR1-AB seropositive patients showed inferior results in Score A (p = 0.006), Score B (p = 0.004), BDI (p = 0.013) and FIS (p = 0.018), compared to seronegative patients. Multiple regression analysis including covariates age, NIHSS at day 7 post-stroke, and days from stroke to follow-up, showed NMDAR1-AB seropositivity associated with worse outcome in Scores A (b: 1.517, 95%CI: 0.505-2.529, p = 0.004) and B (b: 0.803, 95%CI: 0.233-1.373; p = 0.006). Also FIS was unfavorably associated with NMDAR1-AB seropositivity (binary logistic regression: OR: 3.904, 95%CI: 1.200-12.695; p = 0.024). CONCLUSIONS: Even though the numbers of included patients are low, our data apparently indicate that NMDAR1-AB seropositivity at the time point of acute ischemic stroke is associated with neuropsychiatric symptoms including cognitive dysfunction and fatigue years after stroke. Preclinical proof of a causal relation provided, targeted immunosuppression may be a future prophylactic option to be clinically evaluated.

Plasma Dimethylarginine Levels and Carotid Intima-Media Thickness are related to Atrial Fibrillation in Patients with Embolic Stroke.

A relevant part of embolic strokes of undetermined source (ESUS) is assumed to be due to non-detected atrial fibrillation (AF). In this study, we aimed to investigate if markers of endothelial dysfunction and damage may indicate AF risk in embolic stroke. Eighty-eight patients with ischemic stroke confirmed by imaging were assigned to one of three groups: ESUS, AF, or micro-/macroangiopathy. ESUS patients underwent prolonged Holter electrocardiography scheduled for three days. The National Institutes of Health Stroke Scale (NIHSS), the CHA2DS2VASC score, and the carotid intima⁻media thickness (CIMT) were obtained. Markers of endothelial (dys)function (L-arginine, asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA)) were measured at day seven after stroke. ESUS patients were younger and had fewer cardiovascular risk factors than patients with determined stroke etiology. Compared with AF patients, ESUS patients showed significantly lower values of SDMA (p = 0.004) and higher values of L-arginine (p = 0.031), L-arginine/ADMA ratio (p = 0.006), L-arginine/SDMA ratio (p = 0.002), and ADMA/SDMA ratio (p = 0.013). Concordant differences could be observed comparing ESUS patients with those with newly diagnosed AF (p = 0.026; p = 0.03; p = 0.009; p = 0.004; and p = 0.046, respectively). CIMT was significantly larger in AF than in ESUS patients (p < 0.001), and was identified as an AF risk factor independent from CHA2DS2VASC in the regression analysis (p = 0.014). These findings may support future stratification for AF risk in patients who have suffered embolic stroke.

Early Post-Stroke Infections Are Associated with an Impaired Function of Neutrophil Granulocytes.

To investigate whether neutrophil granulocytes' function relates to post-stroke infections and clinical outcome after stroke, we prospectively recruited 95 patients after ischemic stroke and tested them for their microbiocidal neutrophil functions in this exploratory study. Additionally, 24 age-adjusted controls were examined regarding neutrophil function. Phagocytic capacity and the ability of the neutrophil granulocytes to produce reactive oxygen species (ROS) as well as CD11b and CD16 receptor expression profile were measured by flow cytometry at days 1, 3, 7, and 90 after symptom onset. Primary outcome was the development of an infection within the first week after stroke. Results of neutrophil functional measurements were compared between patients with and without infection as well as between all stroke patients and controls. Further risk factors for the development of infections were summarized in an infection-risk score for the purpose of multivariate statistical analysis. The ROS production in neutrophils after stimulation with formyl-methionyl-leucyl-phenylalanine (fMLP) was reduced at baseline in patients with post-stroke infections compared to those without (p = 0.013). This difference proved to be independent from the infection-risk score in the binary logistic regression (p = 0.011). Phagocytosis and oxidative bursts were not significantly reduced in the whole stroke patient group compared to controls. Dysfunction of neutrophil granulocytes seems to play a significant role in the development of post-stroke infections. Further studies are warranted to investigate neutrophil granulocytes´ function as a potential biomarker of post-stroke infections.