Cerebral ischemia model induces a severe damage of neuron, and it causes behavioral dysfunction in rats (Arvidsson et al 2002; Zuk et al., 2001). The criteria of functional recovery in stroke patients being the main issue, this gradual enlargement of infarction can be limited by a variety of interventions that do not interfere with cerebral blood flow. Therefore, we performed an analysis to investigate the evolution of infarct after middle cerebral artery occlusion (MCAO). We induced this analysis in rats, a model system that has been less well characterized. In order to analyze the changes following transient MCAO, several different histochemical methodologies can be utilized. 2,3,5-Triphenyltetrazolium chloride (TTC) staining is one of the most common histochemical stains used to assess cerebral injury. In ischemic tissue, lack of TTC staining is considered infracted with white color and defined as core and viable tissue is stained red (Iihoshi et al., 2004; Jin et al., 2005; Kim et al., 2007).

We examined the effects of the transplantation of mouse embryonic stem (mES) cells on behavioral function induced by focal ischemia in rats. Stem cell transplantation has established as a potential effective therapy for CNS disorders such as ischemic stroke and spinal cord injury. Embryonic stem (ES) cells are capable of proliferating and differentiation into neural progenitor cells with the use of induction protocols leading to the development of functionally mature neurons and glial cells (Barone and Feuerstein, 1999). Self-renewing, totipotent ES cells may use a virtually unlimited donor source for tissue transplantation (Benedek et al., 2006). We have focused on determining the appropriate culture condition to induce neural cell differentiation of ES cells with good cell viability. A cell based-therapy may have the advantage of exerting multiple therapeutic effects at various sites and times within the lesion as the cells respond to a particular pathological microenvironment. A single injection of mESCs several hours after ischemia onset can reduce infarction size and improve functional outcome in rodent cerebral ischemia models (Chiba et al., 2004; Englund et al., 2002). Endogenous neurogenesis and migration of precursor cells may help to replace some lost neurons in brain structures such as striatum (Ide et al., 1999). Transplantation of embryonic stem cells remains to be the most liable way to repair the massive damage in the cerebral cortex after ischemic stroke. Previous studied suggested that stem cell therapy is a powerful method for neurological disorders (Ikeda et al., 2005). We evaluated whether mES cells transplantation reduces the infarct size with functional improvement in the experimental models.

Stem cell transplantation can restore function in rodent models of diabetes, immunodeficiency, and myocardial infarction, by stimulating the production of pancreatic b-cells, lymphocytes, or endothelial cells from endogenous precursors. It has been reported that transplantation of mouse ES cells into rat brain following experimental stroke reduced infarct volume and improved behavioral outcome (Kim et al., 2007). In the present study, it is reported that transplantation also stimulated neurogenesis in the SVZ ipsilateral to stroke (Lin et al., 1998). Because the magnitude of the inflammatory response and its harmful effects as well as the types of released cytokines change with time after ischemia (Liu et al., 2009; Modo et al., 2000). The timing of the transplant could significantly influence graft survival, and longer survival could be predicted if cells are transplanted once inflammation has subsided. The present study shows that transplantation of mESC not only increased early survival of transplanted cells but also accelerated behavioral recovery following stroke. This is consistent with previous report that grafted embryonic stem cells develop into functional neurons and could integrate into host cortical circuitry (Nakatomi et al., 2002; Omori et al., 2008; Theus et al., 2008). The functional recovery 7 days after transplantation suggests the therapeutic advantage of accelerated repair processes and functional restoration.

The motor functional benefit, however, was more persistent in mESC transplantation, showing much better performance than control animals at 28 days after MCAO. More long-term investigation may be needed to verify the persistence of the morphological and functional benefits of the transplantation strategy (Barone and Feuerstein, 1999). Our finding showed that the infarct size was significantly reduced in both cell-transplanted groups and the post-ischemic exercise group, compared with the sham-operated group. Although the basis for the propagation of injury is unclear, it is well known that an ischemic brain infarct progresses over time (Wei et al., 2006). Therefore, we postulate that the reduction of the infarct area in the transplanted group, compared with the control operated group was induced by reduction of secondary damage in the penumbra area initiated by ischemia and cell death. Cell transplantation may induce certain functional recovery of the brain tissue by endogenous cell mediated effect (Zhao et al., 2002). Our study suggest that intracerebrally injected mES cells survived and migrated into the infarct area from inoculation site and neuroglially differentiated in the ischemic brain area of adult rats.