**5. Pre-Clinical Neuroinflammation Treatment in the Post-Ischemic Brain**

Anti-inflammatory actions are aimed at reducing neuroinflammatory reactions by inhibiting the factors and phenomena that increase inflammation, and at the same time stimulating anti-inflammatory factors and phenomena naturally occurring in the body (Table 1). One of the many cytokines most involved in the development of post-ischemic brain neurodegeneration is IL-1 [104]. IL-1 deficient mice showed a significant reduction in infarct volume of approximately 70% after focal brain ischemia compared to that of wild-type mice [104]. It has been shown that inhibition of microglia activation by 2% isoflurane in transient focal brain ischemia in rats reduced the infarct size, attenuated apoptosis, and significantly decreased microglia activation in ischemic penumbra [105]. Within 7 days after focal brain ischemia, edaravone, a free radical scavenger that mimics glutathione peroxidase, reduces microglia activation and early accumulation of oxidative products in rats [106]. Similarly, multiple exposures to hyperbaric oxygen reduced infarct volume by decreasing microglia activation (Table 1) [107]. Inactivation of NF-kB in astrocytes promoted survival of the neurons after ischemic damage in mice [108]. Treg immunodepletion using a specific CD-25 antibody aggravated tissue injury and impaired neurological deficits on day 7 after local brain ischemia in mice (Table 1) [94]. A beneficial effect of administration of a recombinant human interleukin-1 receptor antagonist on the reduction of neuronal death in rats after hypoxia-ischemia and focal cerebral ischemia has also been shown (Table 1) [109,110]. Administration of anti-TNF-α antibody (polyclonal rabbit anti-mouse TNF-α neutralizing antibody) improved neurological outcomes in rats after reversible local brain ischemia [111]. An approximately 40% reduction in infarct volume was observed in IL-10 overexpressing transgenic mice after local brain ischemia with a parallel decrease in caspase 3 levels compared to that of wild-type mice [112]. The administration of insulin-like growth factor-1 reduced infarct volume and improved sensitivity and mobility in mice [113]. In microglia after ischemia, Toll-like receptors (TLR) induce the expression of genes for cytokines with pro-inflammatory properties [114–116]. Knockout mice for TLR4, but not for TLR3 or TLR9, showed a significant reduction in infarct volume after ischemia compared to that of wild-type mice [114]. It has been found that deficiency of T and B lymphocyte cells in animal models of ischemic stroke resulted in a smaller lesion size and reduced neuroinflammation [117]. It has been shown that inhibition of T lymphocyte migration to the brain reduces infarct volume and post-ischemic inflammation [118]. Reduction in infarct size and improvement in neurological outcomes following local brain ischemia were noted after administration of heme oxygenase-1 and in heme oxygenase-1 overexpressing transgenic mice due to anti-inflammatory and antiapoptotic properties [119,120]. Additionally, heme oxygenase-1 deficient mice have been shown to exhibit increased infarct size following local permanent brain ischemia compared to that of wild-type controls (Table 1) [121].


**Table 1.** Preclinical studies in the prevention/treatment of neuroinflammation in post-ischemic brain neurodegeneration.

IL-1: Interleukin 1, TNF-α: Tumor necrosis factor-α, IL-10: Interleukin 10, IGF-1: Insulin-like growth factor 1, TLR4: Toll-like receptor 4, T cells: T lymphocytes, HO-1: Heme oxygenase 1.

### **6. Neuroinflammation: Good or Bad?**

Accumulating evidence suggests that neuroinflammation plays a key role in the pathogenesis of ischemic stroke and has become an interesting target for therapeutic intervention. Numerous reports indicate that neuroinflammatory cells play multiphase roles (beneficial and harmful) in which inhibiting the same pathway at the wrong time may exaggerate pathogenesis. Thus, better characterizing the pathophysiology of ischemic stroke together with timed treatment may provide the ultimate protective strategy of benefit.

A large amount of important data regarding the connection of inflammatory cells and their mediators that are released into the ischemic neurodegenerative brain has been accumulated in recent years. However, it is unclear whether the malfunctioning of inflammatory cells initiates the pathophysiological events of post-ischemic neurodegeneration, or whether the dysfunction of the inflammatory cells is a consequence of other adverse changes that occur in the early stages of the disease. Therefore, more data are needed on the origin of the inflammatory cell dysfunction. Recent research suggests that factors released from healthy inflammatory cells may also play a protective role when neurodegenerative conditions occur [122]. In conclusion, it is very interesting, but at the same time extremely difficult to understand how inflammatory cells exert various actions in different tissues under physiological and pathological conditions. Their functional complexity clearly requires an interdisciplinary approach to develop novel therapeutic interventions that will benefit from the multi-faceted nature of inflammatory cells, including their ability to facilitate crosstalk between the systemic environment and the brain.

Processes related to post-ischemic neurodegeneration of the brain are a strong support for the contribution of neuroinflammation to the progress of ischemic neuropathology (Figure 1). However, which aspects of this contribution seem positive or negative are still under debate. In this review, we suggest that there is a delicate balance in the response of neuroinflammation to post-ischemic brain neurodegeneration that may have both beneficial and harmful effects

(Figure 1) [122]. We suggest that certain aspects of neuroinflammation in the post-ischemic brain are necessary and beneficial, and may limit or prevent post-ischemic neuropathology. By exploiting the benefits of post-ischemic neuroinflammation, we can work to find a causal therapy for post-ischemic neurodegeneration. Scientists have focused too long on the negative consequences of astrogliosis and microgliosis, judging the neuroglial cells as excited. However, recent evidence has demonstrated the enormous heterogeneity of neuroglial cells and the variability of their functioning. Although our primary focus is on the involvement of the innate immune system in post-ischemic brain neurodegeneration, evidence suggests that the adaptive immune system also plays a role in this pathology, which is an area that requires further research. Although it is clear that neuroinflammation contributes to the pathogenesis of post-ischemic injury, it is too broad a word to describe the mixture of various elements of the innate immune processes that become active as the disease progresses [122]. Microgliosis involves phagocytosis of amyloid plaques, neurofibrillary tangles, dysfunctional synapses, and the release of trophic factors for cell plasticity and growth (Figure 1) [123]. In contrast, microgliosis increases the amounts of chemokines and cytokines, which become toxic and harmful to neurons (Figure 1). Astrogliosis is also beneficial because it causes the propagation of calcium currents to increase signal conduction and improve repair and protection. In contrast, excessive astrogliosis can increase the amount of various neurotoxic substances. Therefore, the neuroinflammatory and immune responses following ischemic brain injury are both good and bad. Treatment options based on these data suggest early use of antibiotics to prevent infection (in stroke patients) and inhibition of the early inflammatory/immune response, although it may be likely that after brain ischemia develops, it is too late for this treatment, and treatment should focus on enhancing the protective immune response to accelerate the repair of neurons that survived the primary ischemic episode. It is clear that neuroinflammation is strongly involved with post-ischemic neurodegeneration of the brain (Figure 1), but a more thorough understanding of the ischemic-specific immune/inflammatory response will be critical in developing causal therapy for post-ischemic brain injury.

**Figure 1.** There is a delicate balance between the beneficial and harmful effects of neuroinflammation in post-ischemic brain neurodegeneration. Some neuroinflammation phenomena are protective, such as phagocytosis, but others, such as pro-inflammatory mediators, are detrimental.
