Intraspinal Cavity Injection of Human Mesenchymal Stem Cells and Tracking their Migration into the Rat Brain

Summary

Several routes of administration can be used to deliver mesenchymal stem cells (MSCs) to the brain. In the present study, MSCs were delivered throughout the neuraxis and brain via intra-spinal cavity injection. MSCs were injected into the spinal cavities of rats, and stem cell migration was tracked and quantified.

Abstract

Mesenchymal stem cells (MSCs) have been studied for the treatment of various diseases. In neurodegenerative diseases involving defects in both the brain and the spinal cord, the route of administration is very important, because MSCs must migrate to both the brain and the spinal cord. This paper describes a method for administering MSCs into the spinal canal (intraspinal cavity injection) that can target the brain and spinal cord in a rat model. One million MSCs were injected into the spinal canals of rats at the level of lumbar vertebrae 2-3. After administration, the rats were euthanized at 0, 6, and 12 h post-injection. Optical imaging and quantitative real-time polymerase chain reaction (qPCR) were used to track the injected MSCs. The results of the present study demonstrated that MSCs administered via the spinal cavity could be detected subsequently in both the brain and spinal cord at 12 h. Intraspinal cavity injection has the advantage of not requiring general anesthesia and has few side effects. However, the drawback of the low migration rate of MSCs to the brain must be overcome.

Introduction

Mesenchymal stem cells
Under disease conditions, MSCs secrete disease-specific therapeutic substances via paracrine actions¹ that have been reported to regulate immune responses, restore damaged tissues, and remove toxic substances². Therefore, MSC therapy is considered more effective than single-target therapy in treating multifactorial diseases such as Alzheimer's disease and sarcopenia^3,4,5,6. Additionally, in contrast to pharmaceuticals, MSCs have a homing effect,....

Protocol

NOTE: This study was approved by the Institutional Animal Care and Use Committee (Approval number: 20170125001, Date: January 25, 2017) of the Samsung Biomedical Research Institute (SBRI) at Samsung Medical Center. As an accredited facility of the Association for Assessment and Accreditation of Laboratory Animal Care International, the SBRI acts in accordance with the guidelines set forth by the Institute of Laboratory Animal Resources.

1. Preparation of human Wharton's jelly-derived MSCs

1. Cultivation of human Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs)
   1. Thaw a vial of human WJ-MSCs quickly in a 37....

Results

To evaluate the efficacy of intraspinal cavity injection of MSCs, DiD-labeled MSCs were used in the present study. Before injecting MSCs into the spinal cavity, the labeling efficacy was assessed in vitro using optical imaging and fluorescence microscopy (Figure 1). After staining the MSCs with the DiD labeling reagent using the procedure described in protocol section 3.1, optical images were taken of the culture plates on which DiD-labeled MSCs were seeded (Figure 1A

Discussion

The optimal route of administration for treatment with MSCs should be chosen depending on the target disease, the patient's condition, and the type of drug to be delivered. In cell therapies, including MSC therapy, direct injection of stem cells into the brain or intrathecally via the CSF must be considered as the cells cannot pass through the BBB¹⁹. Intraspinal cavity injection is relatively non-invasive and does not cause neuronal damage in the brain, unlike intracerebroventricular injection.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
0.25% Trypsin-EDTA	Gibco-invitrogen	25200114	Cell culture
Fetal bovine serum	biowest	S1520	Culture medium supplement
gentamicin	Gibco-invitrogen	15710-072	Culture medium supplement
Gentra Puregene Tissue Kit	QIAGEN	158689	gDNA isolation
MEM, no glutamine, no phenol red	Gibco	51200038	WJ-MSC fomulation for injection
Miminum Essential Medium alpha	Gibco-invitrogen	12571063	WJ-MSC culture medium
Power SYBR Green PCR Master Mix	Applied Biosystems	4368577	quantitative real time PCR reagent
QuantStudio 6 Flex Real-Time PCR System	Thermo fisher	4485694	quantitative real time PCR
trypan blue	Gibco	15250061	Injection
Vybrant DiD Cell-Labeling Solution	invitrogen	V22887	Stem cell labeling solution
Xenogen IVIS Spectrum system	Perkin Elmer	124262	Optical imaging device