Biomechanical Forces Regulate Gene Transcription During Stretch-Mediated Growth of Mammalian Neurons.

Joseph R Loverde,Bryan J Pfister,Patricia Soteropoulos,Rosa E Tolentino

doi:10.3389/fnins.2020.600136

Abstract

At birth, there are 100 billion neurons in the human brain, with functional neural circuits extending through the spine to the epidermis of the feet and toes. Following birth, limbs and vertebrae continue to grow by several orders of magnitude, forcing established axons to grow by up to 200 cm in length without motile growth cones. The leading regulatory paradigm suggests that biomechanical expansion of mitotic tissue exerts tensile force on integrated nervous tissue, which synchronizes ongoing growth of spanning axons. Here, we identify unique transcriptional changes in embryonic rat DRG and cortical neurons while the corresponding axons undergo physiological levels of controlled mechanical stretch in vitro. Using bioreactors containing cultured neurons, we recapitulated the peak biomechanical increase in embryonic rat crown-rump-length. Biologically paired sham and “stretch-grown” DRG neurons spanned 4.6- and 17.2-mm in length following static or stretch-induced growth conditions, respectively, which was associated with 456 significant changes in gene transcription identified by genome-wide cDNA microarrays. Eight significant genes found in DRG were cross-validated in stretch-grown cortical neurons by qRT-PCR, which included upregulation of Gpat3, Crem, Hmox1, Hpse, Mt1a, Nefm, Sprr1b, and downregulation of Nrep. The results herein establish a link between biomechanics and gene transcription in mammalian neurons, which elucidates the mechanism underlying long-term growth of axons, and provides a basis for new research in therapeutic axon regeneration.

Highlights

Individual neurons span remarkable distance through peripheral nerves and white matter tracts, constituting the largest known mammalian cells
We found that 8 out of 18 significant cortical gene expression changes overlapped with significant dorsal root ganglia (DRG) findings, suggesting there is mechanistic overlap in the stretch-growth of both peripheral and central nervous system neurons
Despite dissimilar stretch-growth rates and final axon lengths, eight genes were mutually significant across our DRG and cortical neuron assays, including upregulation of Sprr1b, Mt1a, Hmox1, Hpse, cAMP responsive element modulator (Crem), glycerol-3-phosphate acyltransferase-3 (Gpat3), neurofilament medium chain (Nefm), and downregulation of neuronal regeneration related protein (Nrep), (Figure 7)

Summary

SIGNIFICANCE STATEMENT

This work establishes a link between biomechanics and gene transcription in the developmental growth of mammalian neurons. While links have been established between biomechanics and gene transcription in bone and muscle tissue, a link between physiological axon stretch and gene transcription has not been clearly established in nervous tissue. Transcriptional Changes of Axon Stretch-Growth analysis presented provide a conceptual advance in our fundamental understanding of axon growth after growth cone differentiation, whereby a dedicated “stretch-growth” mechanism accommodates for axonal elongation in response to the stimulus of biomechanical stretch. The identified transcriptional changes are poised to spur innovative research toward the regeneration of long axons within the spinal cord and peripheral nervous system

INTRODUCTION

MATERIALS AND METHODS

DISCUSSION

DATA AVAILABILITY STATEMENT

Findings

ETHICS STATEMENT