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The Advanced Therapeutic Effect of MY145-enhanced Mesenchymal
Stem Cells on Functional Recovery and Neurogenesis in Stroke Model
1,2
1,2
1,2
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HyeRyeong Sim , Hyun-Jin Kim , JeeIn Choi , MinYoung Kim *
1 Department of Rehabilitation Medicine, CHA Bundang Medical Center, CHA University School of Medicine, Republic of Korea
2 Rehabilitation and Regeneration Research Center, CHA University, Republic of Korea
Abstract Results
Ischemic stroke had limited treatment options due to the irreversibility of neuronal • The results show that MY145-transfected MSC induced better neurological recovery
tissue damage. Recent studies suggested stem cell-based therapy as a promising than non-transfected MSC in the MCAO model.
treatment for stroke; however, its therapeutic effect has been regarded as insufficient. • In-vitro assay revealed increments of MY145 and angiogenic markers by MY145-
According to our previous clinical study, we found MY145, a marker of efficacy, transfected MSC treatment.
was increased in protein screening after stem cell therapy. The aim of this study was • Intravenous administration of MY145-transfected MSC resulted in increased neurogenic
to investigate the role of MY145 which might enhance therapeutic efficacy of stem and angiogenic effects in the ipsi-lesional brain tissue compared to non-transfected MSC.
cells. The neurobehavioral score results showed that MY145-transfected stem cell
induced further amelioration of neurological impairment compared to non-transfected Figure 3. Co-cultured MSCs were increased cell proliferation in OGD-induced cells.
stem cell. In addition, protein expressions of neuronal nuclei and doublecortin, the 0hr 6hr 9hr 9hr + 1day
representative neurogenesis markers, were increased in the brain by stem cell (cell seeding) (OGD) (reperfusion 3hr) (MSC treat)
administration and upregulated most by MY145-transfected stem cell therapy. To OGD
confirm the role of MY145, we measured cell proliferation and MY145 mRNA group
levels after stem cell co-culture of OGD-induced bEnd.3 cell. The stem cell treatment 400μm 400μm 400μm 400μm
significantly upregulated mRNA levels of vascular endothelial growth factor (VEGF)
and basic fibroblast growth factor (bFGF), which are angiogenesis markers. In
conclusion, this study suggests that MY145-enhaced MSC has synergistic effects and OGD+MSC
that MY145 is a potential marker for predicting the therapeutic effect of stem cell group
therapy.
400μm 400μm 400μm 400μm
Materials & Methods
Figure 4. MSCs increased mRNA expression of the MY145, VEGF and bFGF in OGD-
• Animals induced cells.
- Sprague-Dawley rats (160~180g, male)
✱✱ ✱ 8 ✱✱
• Transient middle cerebral artery occlusion 6 10 Control
- Occlusion time: 90 min 8 6 OGD
OGD+MSC
• Neurological functional tests 4 6 4
- modified Neurological Severity Scores MY145 mRNA Expression (Fold change) VEGF mRNA Expression (Fold change) 4 bFGF mRNA Expression 2 (Fold change)
- Statistical analysis: 2
Mean±standard error of the mean, One-way ANOVA 2
• Cell dose of administered MSC, Intravenous injection 0 0 0
- 3  10 6 (*P<0.05, **P<0.01 / One-way ANOVA)
• Transfection Figure 5. Bigger amelioration of neurological impairment by MY145-transfected MSCs
- MSCs were plated into 6 wells plates in density 7x10 cells per well in Opti-MEM
4
medium 30min before experiments mNSS ΔD7-D28
✱✱✱
- MY145 was transfected into MSCs 14 Saline (n=8) 5 ✱
12
Figure 1. Middle Cerebral Artery Occlusion Model (MCAO) 10 MSC (n=5) 4
mNSS scores 8 3
MSC+MY145 (n=9)
infarct 6 Improved mNSS scores 2
2 4 1
0
Pre D1 D3 D7 D14 D21 D28 0
Day(s) after MCAO (*P<0.05, ***P<0.005 / One-way ANOVA)
Figure 6. Elevated protein expressions representing neurogenesis by MY145-transfected
MSCs treatment in the ipsi-lesional brain tissue
Figure 2. Schematic of experimental setup 5 ✱✱ 2.0 2.5
DCX protein Expression (Fold change) Nestin protein Expression (Fold change) 1.0 NeuN protein Expression (Fold change) 1.0
◀ DCX 4 2.0
◀ Nestin 3 1.5 1.5
◀ NeuN 2 1 0.5 0.5
◀ β-actin 0 0.0 0.0
Saline MSC+MY145 Saline MSC+MY145 Saline MSC MSC+MY145
MSC
MSC
Table 1. Modified Neurological Severity Scores (mNSS) ( **P<0.01 / One-way ANOVA)
Figure 7. Elevated CD31 protein expression by MY145-transfected MSCs treatment
Motor tests Points in the ipsi-lesional brain tissue
Raising the rat by the tail 3
1=Flexion of forelimb 2.0
1=Flexion of hindlimb Saline
1=Head moved >10° to the vertical axis within 30s 1.5 MSC
Walking on the floor (normal=0; maximum=3) 3 ◀ CD31 MSC+MY145
0=Normal walk CD31 protein Expression (Fold change) 1.0
1=Inability to walk straight ◀ β-actin
2=Circling toward the paretic side 0.5
3=Fall down to the paretic side
Sensory tests 2 0.0
1=Placing test (visual and tactile test)
1=Proprioceptive test (deep sensation, pushing the paw against the table edge) Conclusion
Beam balance tests (normal=0; maximum=6)
0=Balances with steady posture 6 • In conclusion, we have shown that administration of MY145 transfected MSCs exhibits
1=Grasps side of beam
2=Hugs the beam and one limb falls down from the beam the therapeutic effects by improving recovery of neurological function after tMCAO.
3=Two limbs fall down from the beam or spins on the beam (>60s) • MY145 is a potential marker for predicting the therapeutic effect of stem cell therapy
4=Attempts to balance on the beam but falls off (>40s) such as MSCs.
5=Attempts to balance on the beam but falls off (>20s)
6=Falls off : no attempt to balance or hang on to the beam (<20s) Reference
Total 14
• Choi, J. I., Choi, J. W., Shim, K. H., Choung, J. S., Kim, H. J., Sim, H. R., ... & Kim, M. (2021). Synergistic Effect in
Acknowledgement Neurological Recovery via Anti-Apoptotic Akt Signaling in Umbilical Cord Blood and Erythropoietin Combination
Therapy for Neonatal Hypoxic-Ischemic Brain Injury. International journal of molecular sciences, 22(21), 11995
• This work was supported by a grant of the Korea Health Technology R&D Project through the Korea Health • Suh, M. R., Lim, I., Kim, J., Yang, P. S., Choung, J. S., Sim, H. R., ... & Kim, M. (2021). Efficacy of Cord Blood Cell
Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant . Therapy for Hutchinson–Gilford Progeria Syndrome—A Case Report. International journal of molecular
number : HI16C1559) sciences, 22(22), 12316.