Abstract
Spinal cord injury or amyotrophic lateral sclerosis damages spinal motor neurons and forms a glial scar, which prevents neural regeneration. Signal transducer and activator of transcription 3 (STAT3) plays a critical role in astrogliogenesis and scar formation, and thus a fine modulation of STAT3 signaling may help to control the excessive gliogenic environment and enhance neural repair. The objective of this study was to determine the effect of STAT3 inhibition on human neural stem cells (hNSCs). In vitro hNSCs primed with fibroblast growth factor 2 (FGF2) exhibited a lower level of phosphorylated STAT3 than cells primed by epidermal growth factor (EGF), which correlated with a higher number of motor neurons differentiated from FGF2-primed hNSCs. Treatment with STAT3 inhibitors, Stattic and Niclosamide, enhanced motor neuron differentiation only in FGF2-primed hNSCs, as shown by increased homeobox gene Hb9 mRNA levels as well as HB9+ and microtubule-associated protein 2 (MAP2)+ co-labeled cells. The increased motor neuron differentiation was accompanied by a decrease in the number of glial fibrillary acidic protein (GFAP)-positive astrocytes. Interestingly, Stattic and Niclosamide did not affect the level of STAT3 phosphorylation; rather, they perturbed the nuclear translocation of phosphorylated STAT3. In summary, we demonstrate that FGF2 is required for motor neuron differentiation from hNSCs and that inhibition of STAT3 further increases motor neuron differentiation at the expense of astrogliogenesis. Our study thus suggests a potential benefit of targeting the STAT3 pathway for neurotrauma or neurodegenerative diseases.
Highlights
Acute spinal cord injury (SCI) and amyotrophic lateral sclerosis (ALS) are characterized by death of cholinergic motor neurons accompanied by reactive astrogliosis, i.e. hypertrophy and proliferation of astrocytes and alterations in their gene expression patterns
We found that unprimed human neural stem cells (hNSCs) grown as free-floating neurospheres (Fig. 1B) had considerable amounts of pSTAT3-Y705, pSTAT3-S727 and total Signal transducer and activator of transcription 3 (STAT3) (Fig. 1A, sphere)
The main findings of our study are that 1) hNSCs primed with fibroblast growth factor 2 (FGF2) had reduced levels of phosphorylated/activated STAT3; 2) Treatment of FGF2-primed, but not epidermal growth factor (EGF)-primed, hNSCs with specific pharmacological inhibitors of STAT3 enhanced motor neuron fate specification at the expense of astroglial differentiation; 3) The STAT3 inhibitors, Stattic and Niclosamide, do not inactivate STAT3 signaling by affecting STAT3 expression or its phosphorylation, but by blocking nuclear transport of activated STAT3
Summary
Acute spinal cord injury (SCI) and amyotrophic lateral sclerosis (ALS) are characterized by death of cholinergic motor neurons accompanied by reactive astrogliosis, i.e. hypertrophy and proliferation of astrocytes and alterations in their gene expression patterns. It is clear that increased astrogliosis resulting from acute spinal injury or chronic neurodegenerative conditions creates a highly gliogenic cellular environment, which is not conducive to the formation or long-term survival of motor neurons. In such patients, potential therapy should employ a two-pronged approach: 1) reduce the local gliogenic environment and 2) switch the environmental milieu such that it promotes/sustains neurogenesis
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