*3.2. Cells*

The involvement of cells other than neurons in the development of post-ischemic brain neurodegeneration has been evaluated in many experimental and clinical studies [1,26,27,37]. When neuroinflammation following brain ischemia begins, it starts with the release of pro-inflammatory factors that involve various cells. First, we observe the involvement of neuroglial cells in the brain. Next, leukocytes, monocytes, and other cells with immune functions enter the ischemic brain tissue. This mechanism may additionally exacerbate post-ischemic brain damage by increasing blood–brain barrier permeability, edema, and progressive neuronal death. The variety of cells involved in this process can have a beneficial or detrimental effect, depending on the post-ischemia period: early or delayed. Ultimately, we need to know which cells are involved in post-ischemic changes and when they are involved to establish ways to control them.

#### 3.2.1. Microglia

Microglia, the resident innate immune cells of the brain accounting for up to 20% of the neuroglial population, undergo morphological and phenotypic changes after brain ischemia [1]. Activated microglia function like macrophages during systemic inflammation and have the ability to remove foreign organisms and cellular debris. At rest, the microglia are referred to as small cells with wide protruding branches. However, after ischemia, microglial cells are activated, they change shape and function, but the exact mechanisms of this phenomenon are still unknown. They are activated after brain ischemia, as a result of which changes in their phenotypes can be observed [8,26,27,42–44,51]. Transient focal brain ischemia in the rat leads to microglia activation in the cerebral cortex of the ischemic hemisphere, and the severity and extent of the injury is reflected in the intensification of microglia activation [52]. The microglia around the post-ischemic parenchyma migrate towards the ischemic lesion and remain in close relationship with the neurons in a process called "capping", that is, after neuron death, the capping helps in early recognition and rapid phagocytic removal of dead neurons [53,54]. They become active a few minutes after the onset of brain ischemia, increasing in number in the following days, reaching a peak on the tenth day after transient local brain ischemia [55]. After ischemic injury, the microglia activating the phenotype become amoeboid and have a functional macrophage character. After this transformation, microglia look like macrophages not only in appearance but also in their behavior, they can release cytokines and secrete extracellular matrix metalloproteinases (MMPs) that are able to damage the blood–brain barrier and thus increase its permeability. This process facilitates the early transfer of leukocytes from the circulation to the ischemic brain, contributing to an increased level of pro-inflammatory factors that aggravates post-ischemic injury. Once activated, the microglial cells can take on two different phenotypes: classic pro-inflammatory (M1) and alternative anti-inflammatory (M2). The M1 phenotype releases the cytokines TNF-α, IL-6, and IL-1β and substances with oxidizing properties, such as nitric oxide [56]. The M2 phenotype has beneficial effects, it causes ischemic brain healing post-ischemia and the release of anti-inflammatory factors, such as IL-4 and IL-10, and secretes many factors with neurotrophic properties capable of preventing the development of neuroinflammation [56]. A recent study showed that microglia depletion by the dual colony-stimulating factor-1 inhibitor, PLX3397, exacerbates brain infarction and neurological deficits [57]. Following a transient focal brain ischemia, microglial depletion enhances leukocyte infiltration, expression of inflammatory factors, and neuronal loss in mice [57]. This pathological phenomenon is dependent not only on lymphocytes and monocytes, but also on astrocyte-mediated inflammatory factors. Hence, the presence of microglial cells prevents astrocytes from secreting inflammatory factors during and after ischemia [57]. Moreover, by supporting the above, the microglial cells produce different neurotrophic factors that animate neurogenesis and plasticity [58]. Thus, following brain ischemia, different subsets of microglial cells have different roles.
