Modified mesenchymal stem cells for brain remodeling and motor recovery in a rodent stroke model

2018 Fiscal Year Research-status Report

• Project/Area Number: 18K16577
• Research Institution: University of Tsukuba
• Principal Investigator: プエンテス サンドラ 筑波大学, システム情報系, 助教 (00725765)
• Project Period (FY): 2018-04-01 – 2021-03-31
• Keywords: Mesenchymal stem cells / Stroke / Microglia polarization

Outline of Annual Research Achievements

During the last fiscal year, the project has advanced as planned. Adipose tissue derived mesenchymal stem cells from baby and adults were obtained and its modification with microvesicles from human endothelial progenitor cells was also achieved. Mice underwent transient middle cerebral artery occlusion (MCAO) using the filament model and reperfusion was allowed 1 hour after ischemia onset. Animals were randomly assigned to six different groups including 5 different cell conditions and one negative control (phosphate buffer saline injection). Only animals who exhibited Longa score (neurologic deficit assessment) 3 to 5 were included.
Mortality rate for MCAO did not vary from previous reports and experimental success rate (animals who exhibited Longa score 3-5 and survived after treatment) was above 70%.
From these animals, brain samples has been collected and histochemistry for infarct size quantification and immunohistochemistry to evaluate microglial polarization, vascular, astrocytic and neuronal status is undergoing. Cell quantification has been going on the penumbral area of cortical and subcortical areas and also infarct core for microglial polarization. Statistical analysis is still pending since animal surgeries and sample collection is not finished yet.

Current Status of Research Progress

3: Progress in research has been slightly delayed.

Reason

As expected, mice experiments have been going without trouble. Mortality rate was expected and the number of dead animals after treatment was small. Immunohistochemistry takes time since several randomized images are needed. Picture collection has been progressing as planned. Also, cell counting will be done by a researcher blinded to the experiments. Training for cell counting has been completed and image analysis will start soon. In vitro experiments were scheduled for the end of the previous fiscal year but it has not been started yet.

Strategy for Future Research Activity

Since the research is progressing according to out plans, we will keep collecting brain samples to complete 6 to 8 animals per group. Immunohistochemistry, cell counting and histology for infarct size evaluation also will be done. Additionally we will start in vitro experiments to evaluate the behavior of neurons under oxygen and glucose deprivation in the presence of the different cell types that are used for animal treatment.

Causes of Carryover

The non-used part is related to in vitro experiments. This budget part was supposed to be invested in culture reagents and culture disposables. Since in vitro experiments will be starting within this fiscal year, the plan was to carry over the budget so it can be used this year as planned before.