Transplants of Fibroblasts Genetically Modified to Express BDNF Promote Regeneration of Adult Rat Rubrospinal Axons and Recovery of Forelimb Function

Abstract

Adult mammalian CNS neurons do not normally regenerate their severed axons. This failure has been attributed to scar tissue and inhibitory molecules at the injury site that block the regenerating axons, a lack of trophic support for the axotomized neurons, and intrinsic neuronal changes that follow axotomy, including cell atrophy and death. We studied whether transplants of fibroblasts genetically engineered to produce brain-derived neurotrophic factor (BDNF) would promote rubrospinal tract (RST) regeneration in adult rats. Primary fibroblasts were modified by retroviral-mediated transfer of a DNA construct encoding the human BDNF gene, an internal ribosomal entry site, and a fusion gene of lacZ and neomycin resistance genes. The modified fibroblasts produce biologically active BDNF in vitro. These cells were grafted into a partial cervical hemisection cavity that completely interrupted one RST. One and two months after lesion and transplantation, RST regeneration was demonstrated with retrograde and anterograde tracing techniques. Retrograde tracing with fluorogold showed that approximately 7% of RST neurons regenerated axons at least three to four segments caudal to the transplants. Anterograde tracing with biotinylated dextran amine revealed that the RST axons regenerated through and around the transplants, grew for long distances within white matter caudal to the transplant, and terminated in spinal cord gray matter regions that are the normal targets of RST axons. Transplants of unmodified primary fibroblasts or Gelfoam alone did not elicit regeneration. Behavioral tests demonstrated that recipients of BDNF-producing fibroblasts showed significant recovery of forelimb usage, which was abolished by a second lesion that transected the regenerated axons.

Neurotrophins: Potential Therapeutic Tools for the Treatment of Spinal Cord Injury

“…These cells can be noninvasively isolated from a patient, rapidly expanded in vitro, transduced to express therapeutic agents and transplanted without the need for immunosuppression. Numerous studies have used these non-neural cells as a means of bridging the lesion cavity, as well as providing extracellular matrix proteins and trophic support for regeneration [23,[59][60][61][62][63][64][65][66][67][68]. Although these cells are unable to initiate myelination of regenerated axons themselves, neurotrophin production by transplanted cells leads to robust graft infiltration by endogenous Schwann cells, which may act to promote or stabilize axonal regeneration in a secondary manner [63,65,69,70].…”

“…NGF promotes the sprouting and regeneration of cholinergic local motor axons, primary nociceptive sensory axons, and cerulospinal axons [104,105]. BDNF-secreting bone marrow stromal cell grafts promote regeneration of a number of neuronal populations including raphaespinal, cerulospinal, rubrospinal, local motor and propprioceptive sensory axons [63,67,94,96]. NT-3 expression, similar to BDNF, promotes the regeneration of ascending sensory neurons across the dorsal root entry zone and within the dorsal columns [60,64,[106][107][108][109].…”

Fate of rubrospinal neurons after unilateral section of the cervical spinal cord in adult macaque monkeys: Effects of an antibody treatment neutralizing Nogo-A

“…In the rat, after a spinal cord lesion at cervical levels, axotomized reticulospinal (RS) neurons shrink and display modification in the expression of various molecules (Egan et al, 1977;Kwon et al, 2002Kwon et al, , 2004Novikova et al, 2000;Tetzlaff et al, 1991). In this species, the RNm regains a normal aspect if brain derived neurotrophic factor (BDNF) is provided (Kwon et al, 2002;Liu et al, 1999;Novikova et al, 2000). Because neurotrophic substances are uptaken at synapses, the amount of neurotrophic factors accessible to a neuron relates ultimately to the number of synapses that this neuron possesses.…”

Freeze-dried poly(d,l-lactic acid) macroporous guidance scaffolds impregnated with brain-derived neurotrophic factor in the transected adult rat thoracic spinal cord

“…One approach to promote axonal regeneration in the injured spinal cord, which has been investigated extensively, is the grafting of three-dimensional cellular transplants bridging the injury gap or cyst (e.g., Schwann cells, peripheral nerves; olfactory ensheathing cells; reviewed by [6][7][8]). Many of these repair strategies limit spinal tissue loss [9,10], promote regeneration/ sparing of spinal and supraspinal axons [9][10][11][12][13], and result in some functional recovery [9][10][11][12][13][14][15][16][17][18]. Due to the ongoing (secondary) tissue loss following an injury, it seems inevitable that repair of the (sub-) chronically injured cord will require strategies that include the transplantation of cellular grafts to replace lost spinal nervous tissue.…”