Intravenous Infusion of Mesenchymal Stem Cells Alters Motor Cortex Gene Expression in a Rat Model of Acute Spinal Cord Injury

Abstract

Recent evidence has demonstrated that remote responses in the brain, as well as local responses in the injured spinal cord, can be induced after spinal cord injury (SCI). Intravenous infusion of mesenchymal stem cells (MSCs) has been shown to provide functional improvements in SCI through local therapeutic mechanisms that provide neuroprotection, stabilization of the blood-spinal cord barrier, remyelination, and axonal sprouting. In the present study, we examined the brain response that might be associated with the functional improvements induced by the infused MSCs after SCI. Genome-wide RNA profiling was performed in the motor cortex of SCI rats at 3 days post-MSC or vehicle infusion. Then, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) data revealed that the "behaviorally-associated differentially expressed genes (DEGs)" were identified by the Pearson's correlation analysis with the behavioral function, suggesting that the "behaviorally-associated DEGs" may be related to the functional recovery after systemic infusion of MSCs in SCI. These results suggested that the infused MSCs alter the gene expression signature in the brain and that these expression changes may contribute to the improved function in SCI.

Keywords: MSC; SCI; transcriptome.

FIG. 1.

Locomotor function assessment using the Basso, Beattie, and Bresnahan (BBB) behavioral score. Both groups displayed complete hindlimb paraplegia immediately prior to infusion (1 day after SCI induction) (left). The BBB scores at 3 and 7 days after mesenchymal stem cells (MSC) or vehicle infusion (4 and 8 days after SCI induction) are shown (right). **p < 0.01.

FIG. 2.

Experimental procedures for differentially expressed gene (DEG) analysis (A) Schematic drawing of the experimental procedure. (B) Volcano plot showing DEGs defined with the cutoff of fold change at 1.5 or −1.5, and p < 0.05 by one way ANOVA. The black dots represent the 38 screened DEGs. The 15 coding DEGs are displayed with the gene name. (C) Heat map of 15 coding DEGs showing the expression levels in each SCI+MSC and SCI+vehicle sample. The dendrogram was generated based on the Euclidean distance metric with Ward's method. (D) Charts showing the percentage of the 38 DEGs that belong to each type of RNA including coding, non-coding, precursor microRNA, and unassigned DEGs.

FIG. 3.

Correlation between mRNA expression and behavioral function. Pearson's correlation analysis of the Basso, Beattie, and Bresnahan (BBB) scores and mRNA expression (2-ΔΔCT) of the indicated genes as detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Circles and triangles indicate SCI+MSC and SCI+vehicle samples, respectively. The X-axis represents the BBB score and the Y-axis shows the 2-ΔΔCT of mRNA gene expression.

FIG. 4.

mRNA expression of “behaviorally-associated differentially expressed genes (DEGs).” Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) data of the “behaviorally-associated DEGs” are shown. Sham, SCI+vehicle, and SCI+MSC groups were used. The Y-axis shows the 2-ΔΔCT of mRNA gene expression. Asterisks indicate *p < 0.05, **p < 0.01 by the Tukey–Kramer test.

FIG. 5.

The neuron-restrictive silencer factor (NRSF) binding domains in some “behaviorally-associated differentially expressed genes (DEGs).” Genomic loci of NRSF binding domains within 20 kb from the transcription start of the genes including (A) Kcnip2 (Base genome: human chromosome: chr10 103,561,373–103,626,222), (B) Scn3b (Base genome: human chromosome: chr11 123,481,778–123,540,867), and (C) Pde10a (Base genome: human chromosome: chr6 165,727,008–166,409,519) (red arrows) are shown. The binding motifs of NRSF are indicated with red characters.

FIG. 6.

mRNA expression of non-“behaviorally-associated differentially expressed genes (DEGs).” Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) data of the non-“behaviorally-associated DEGs” are shown. The Y-axis represents the 2-ΔΔCT of mRNA gene expression. Asterisks indicate *p < 0.05, **p < 0.01 by the Tukey–Kramer test.

FIG. 7.

Differential gene expression is transient in the acute phase. Relative fold change of mRNA expression in all coding differentially expressed genes (DEGs) at 3 days post-infusion is shown as red (“behaviorally-associated DEGs”) and blue (non-“behaviorally-associated DEGs”). No significant differences were observed in the comparisons between SCI+MSC and SCI+vehicle groups at day 7 post-infusion.