Source Paper
Neural stem cell mediated recovery is enhanced by Chondroitinase ABC pretreatment in chronic cervical spinal cord injury
Hidenori Suzuki, Christopher S. Ahuja, et al.
Materials & Equipment Checklist
8 items1 from ConductScience
Gather these items before starting the experiment. Check off items as you prepare.
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Protocol Steps
1
Animal acquisition and housing
Obtain 70 adult female wild-type C57BL/6 mice aged 8-10 weeks weighing 15-20g from Jackson Laboratories
View evidence from paper
“A total of 70 adult (8–10 weeks old), female, wild-type C57BL/6 mice (15 to 20g) from Jackson Laboratories”
2
Baseline grip strength measurement
Measure grip strength at baseline (6 weeks post-injury) before randomization into treatment groups
Note: No statistically significant differences between groups at this timepoint
View evidence from paper
“Grip strength measurements at six weeks were not statistically different between groups (ANOVA; p = 0.871)”
3
Randomization into experimental groups
Randomize 60 injured mice into four experimental groups: control, ChABC, iPS-NSCs, and ChABC + iPS-NSCs
View evidence from paper
“Six weeks after injury, mice were randomized into four experimental groups (control, ChABC, iPS-NSCs, & ChABC + iPS-NSCs)”
4
Mini-osmotic pump implantation
Implant mini-osmotic pump rostral to the injury site under anesthesia
View evidence from paper
“Seven weeks post-injury, a mini-osmotic pump was implanted rostral to the injury site”
5
Pump-delivered treatment
Deliver either ChABC in aCSF or aCSF alone via implanted mini-osmotic pump
One week
View evidence from paper
“The pump delivered either Chondroitinase ABC (ChABC) in artificial cerebral spinal fluid (aCSF) or aCSF alone”
6
Pump explantation
Remove mini-osmotic pump after one week of treatment delivery
View evidence from paper
“One week later the pump was explanted”
7
Cell transplantation
Transplant iPS-NSCs into spinal cord parenchyma at four sites (two rostral, two caudal), delivering 50,000 cells in 1 μL per site
View evidence from paper
“induced pluripotent stem cell-derived neural stem cells (iPS-NSC) were transplanted into the cord parenchyma at two rostral and two caudal sites. 50,000 cells in 1 μL were delivered to each site”
8
Immunosuppression administration
Administer Cyclosporine A immunosuppression starting 2 days prior to cell transplantation
From 2 days prior to transplantation until end of study (16 weeks post-injury)
View evidence from paper
“Cyclosporine A immunosuppression was delivered from 2 days prior to transplantation until the end of the study”
9
Behavioral assessments
Perform behavioral assessments including grip strength measurements throughout the study period
Across all 16 weeks of the study
View evidence from paper
“Behavioral assessments were performed across all 16 weeks of the study”
10
Electrophysiologic assessments
Conduct electrophysiologic assessments at end of study
View evidence from paper
“At 16 weeks electrophysiologic assessments were completed”
11
Tissue collection and analysis
Perform whole-animal perfusion, fixation, and immunohistochemical analyses
View evidence from paper
“followed by whole-animal perfusion, fixation and immunohistochemical analyses”
Subjects / Specimens
- Species
- mouse
- Strain
- C57BL/6
- Age
- 8-10 weeks old
- Sex
- female
- Count
- 70
- Weight
- 15 to 20g
Adult wild-type mice