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Stroke is primarily a neurodegenerative disease but can also severely impact the functions of other vital organs and deteriorate disease outcomes. A malfunction of the gastrointestinal tract (GIT), commonly observed in stroke patients, is often characterized by severe bowel obstruction, intestinal microbiota changes and inflammation. Over-activated immune cells after stroke are the major contributors to endorse intestinal inflammation and may induce damage to single-layer epithelial cell barriers. The post-stroke leakage of intestinal barriers may allow the translocation and dissemination of resident microflora to systemic organs and cause sepsis. This overshooting systemic immune reaction fuels ongoing inflammation in the degenerating brain and slows recovery. Currently, the therapeutic options to treat these GIT-associated anomalies are very limited and further research is required to develop novel treatments. In this mini-review, we first discuss the current knowledge from clinical studies and experimental stroke models that provide strong evidence of the existence of post-stroke GIT complications. Then, we review the literature regarding novel therapeutic approaches that might help to maintain GIT homeostasis and improve neurological outcomes in stroke patients.

Post-stroke early activation of neutrophils contributes to intensive neuroinflammation and worsens disease outcomes. Other pre-existing patient conditions can modify the extent of their activation during disease, especially hypercholesterolemia. However, whether and how increased circulating cholesterol amounts can change neutrophil activation responses very early after stroke has not been studied. In this study, we investigated the effect of high-fat diet (HFD) induced hypercholesterolemia on neutrophil activation and stroke outcome. Mice were fed with HFD or normal diet (ND) for six weeks and then induced stroke by transient occlusion of the middle cerebral artery. The activation receptors on immune cells and plasma levels of cytokines were analyzed using flow cytometry. The amount of plasma neutrophil extracellular traps (NETs) was measured using citH3-DNA complex ELISA. We found that HFD-induced cholesterolemia increased the number of circulating and splenic neutrophils in stroke mice but reduced bone marrow neutrophils compared to sham controls. After stroke neutrophils in HFD mice expressed higher levels of activation markers Ly6G and PSGL-1 (CD162) compared to ND mice. In addition, stroke led to an increased expression of the activation markers Ly6C and CD68 on monocyte/macrophages (MΦ) in HFD mice but not in ND mice. Compared to ND, HFD increased plasma levels of the proinflammatory cytokines TNF-α, IL-6, IL-23, and MCP-1 in stroke mice. Remarkably, HFD mice showed higher amounts of circulating NETs, brain-infiltrated neutrophils, and larger infarcts after stroke compared to ND mice. The existence of hypercholesterolemia with a stroke can trigger a stronger activation of neutrophils and MΦ, causing deteriorating disease outcomes.

Abstract Autonomous migration is essential for the function of immune cells such as neutrophils and plays an important role in numerous diseases. The ability to routinely measure or target it would offer a wealth of clinical applications. Video microscopy of live cells is ideal for migration analysis, but cannot be performed at sufficiently high-throughput (HT). Here we introduce ComplexEye, an array microscope with 16 independent aberration-corrected glass lenses spaced at the pitch of a 96-well plate to produce high-resolution movies of migrating cells. With the system, we enable HT migration analysis of immune cells in 96- and 384-well plates with very energy-efficient performance. We demonstrate that the system can measure multiple clinical samples simultaneously. Furthermore, we screen 1000 compounds and identify 17 modifiers of migration in human neutrophils in just 4 days, a task that requires 60-times longer with a conventional video microscope. ComplexEye thus opens the field of phenotypic HT migration screens and enables routine migration analysis for the clinical setting.