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Cellprofiler cell area6/13/2023 ![]() Recent advances in imaging and processing power have led to the development of high-content image (HCI) analysis that enables high-throughput assessment of multiple single cell features, reduces time of analysis, and decreases subjectivity ( Chen et al., 2012 Riordan et al., 2015). However, these standard approaches are limited by the number of cellular features that can be assessed, accompanied by decreased spatial information, slow data processing speeds, and subjectivity. Modern histological approaches have led to significant breakthroughs in understanding thrombus etiology ( Sporns Peter et al., 2017 Heo et al., 2020), patterns of immune cell activation ( Savchenko et al., 2016 Rayasam et al., 2018), and other important pathological and recovery cellular changes. The development of new therapeutic targets and more effective treatments hinge on having a more complete understanding of cellular dynamics after stroke to assess changes in key processes such as the immune response and glial scar formation, which may also serve as biomarkers of injury severity and recovery. Stroke remains a leading cause of death and long-term disability worldwide despite numerous preclinical and clinical trials to develop novel treatments ( Virani Salim et al., 2020). These identified features can provide information of disease pathogenesis and evolution with high resolution, as well as be used in therapeutic screening applications. Ultimately, this unbiased, semi-automated HCI and CellProfiler histopathological analysis approach revealed regional and cell specific morphological signatures of immune and neural cells after stroke in a highly translational porcine model. Principal component analysis (PCA) and correlation analyses demonstrated that stroke-induced significant and predictable morphological changes that demonstrated strong relationships between IBA1 +, GFAP +, and NeuN + areas. Out of 19 morphological parameters assessed in the stroke perilesional and ipsilateral hemisphere regions (38 parameters), a significant change in 38 38 measured IBA1 + parameters, 34 38 GFAP + parameters, 32 38 NeuN + parameters, 31 38 FactorVIII + parameters, and 28 38 DCX + parameters were observed in stroked vs. Evaluation of 19 morphological parameters in IBA1 + microglia/macrophages, GFAP + astrocytes, NeuN + neuronal, FactorVIII + vascular endothelial, and DCX + neuroblast cell areas was conducted on porcine brain tissue 4 weeks post pMCAO. Here, a semi-automated high-content imaging (HCI) and CellProfiler histological analysis method was developed and used in a Yucatan miniature pig permanent middle cerebral artery occlusion (pMCAO) model of ischemic stroke to overcome these limitations. ![]() However, commonly used manual approaches are hindered by limitations in speed, accuracy, bias, and the breadth of morphological information that can be obtained. Histopathological analysis of cellular changes in the stroked brain provides critical information pertaining to inflammation, cell death, glial scarring, and other dynamic injury and recovery responses.
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