with antibodies against cell-type-specific membrane proteins that are conjugated to either magnet or fluorophores. Cell sorting instruments are then used to separate these labeled cells. [3] The labeling-based separation has an inherently high specificity by definition, ensuring good purification quality. However, the labeling-based methods have the disadvantages of high cost, technical complexity, and low yield. In addition, the labeling step during the isolation process may cause undesired changes in cellular properties. [2] Labelfree approaches, on the other hand, allow isolation of the target cells based on their physical properties such as size, density, deformability, and adhesiveness. [4,5] Since physical approaches to cell isolation provide greater accessibility and lower costs than biological approaches, cell separation methods based on the physical properties of cells are currently being widely used. Despite the better accessibility, the physical approach may be considered more delicate because its accuracy is critically dependent on the skill of individual researchers. Among different physical traits of cells, cellular adhesion properties, including cell-matrix adhesion and cell-cell adhesion, are considered the marked traits for identifying distinct cell types or physiological states. For example, in cell state classification, cells of mesenchymal phenotype mainly exhibit robust cell-matrix adhesion, whereas epithelial cells are characterized by prominent cell-cell adhesion. [6] In cancer cells, metastatic cells typically display relatively weaker cell-matrix adhesion than their non-metastatic counterparts. [7] Cell-matrix adhesion strength is also known to contribute to lineage determination during the differentiation of mesenchymal stem cells. [8] Accordingly, the adhesion strength has been utilized as a sorting strategy for cells of different origins and pathophysiological states.Glial cells, which are non-neuronal cells in the brain, have gained considerable attention as key players in neurological diseases. [9] Glial cell populations consist of three types of cells, namely astrocytes, oligodendrocytes, and microglia, each with distinct functions in the brain: astrocytes interact with neurons and blood vessels; [10] oligodendrocytes serve as the primary electrical insulator for neurons; [11] microglia are the critical immune regulators. [12] These three types of glial cells are derived from different origins, thereby exhibiting distinct Glial brain cells, including astrocytes, oligodendrocytes, and microglia, have received much attention as crucial players in neurological diseases. As a result of their critical roles, numerous in vitro studies are being conducted, necessitating the use of appropriate isolation methods for different glial cell types. The most effective glial isolation at the moment is labeling-based protocols that require expensive antibodies and equipment. More commonly used label-free methods are better known for higher accessibility but with compromised accuracy and longer isol...
Traumatic Brain Injury (TBI) by an external physical impact results in compromised brain function via undesired neuronal death. Following the injury, resident and peripheral immune cells, astrocytes, and neural stem cells (NSCs) cooperatively contribute to the recovery of the neuronal function after TBI. However, excessive pro-inflammatory responses of immune cells, and the disappearance of endogenous NSCs at the injury site during the acute phase of TBI, can exacerbate TBI progression leading to incomplete healing. Therefore, positive outcomes may depend on early interventions to control the injury-associated cellular milieu in the early phase of injury. Here, we explore electrical stimulation (ES) of the injury site in a rodent model (male Sprague-Dawley rats) to investigate its overall effect on the constituent brain cell phenotype and composition during the acute phase of TBI. Our data showed that a brief ES for 1h on day 2 of TBI promoted pro-healing phenotypes of microglia as assessed by CD206 expression and increased the population of NSCs and Nestin+ astrocytes at 7 days post-TBI. Also, ES effectively increased the number of viable neurons when compared to the unstimulated control group. Given the salience of microglia and neural stem cells for healing after TBI, our results strongly support the potential benefit of the therapeutic use of ES during the acute phase of TBI to regulate neuroinflammation and to enhance neuroregeneration.Significance StatementTraumatic brain injury (TBI) occurs when a head injury leads to a disruption of normal function in the brain and is a major cause of death and disability, worldwide. The authors used electrical stimulation during the acute phase of TBI, which promoted pro-healing phenotypes of microglia and increased the number of neural stem cells and Nestin+ astrocytes, thereby enhancing neuronal viability. These findings support further study of electrical stimulation to regulate neuroinflammation and to enhance neuroregeneration after TBI.Graphical AbstractFIGURE 1.
Brain cells are influenced by continuous fluid shear stress driven by varying hydrostatic and osmotic pressure conditions, depending on the brain's pathophysiological conditions. While all brain cells are sensitive to the subtle changes in various physicochemical factors in the microenvironment, microglia, the resident brain immune cells, exhibit the most dramatic morphodynamic transformation. However, little is known about the phenotypic alterations in microglia in response to the changes in fluid shear stress. In this study, we first established a flow-controlled microenvironment to investigate the effects of shear flow on microglial phenotypes, including morphology, motility, and activation states. Microglia exhibited two distinct morphologies with different migratory phenotypes in a static condition: bipolar cells that oscillate along their long axis and unipolar cells that migrate persistently. When exposed to flow, a significant fraction of bipolar cells showed unstable oscillation with an increased amplitude of oscillation and a decreased frequency, which consequently led to the phenotypic transformation of oscillating cells into migrating cells. Interestingly, the level of pro-inflammatory genes increased in response to shear stress, while there were no significant changes in the level of anti-inflammatory genes. Our findings suggest that an interstitial fluid-level stimulus can cause a dramatic phenotypic shift in microglia toward pro-inflammatory states, shedding light on pathological outbreaks of severe brain diseases. Given that the fluidic environment in the brain can be locally disrupted in pathological circumstances, the mechanical stimulus by a fluid flow should also be considered a crucial element in regulating the immune activities of the microglia in brain diseases.
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