The survival, proliferation, and differentiation of freshly isolated and cultured cells were studied after absorbing film-assisted laser-induced forward transfer. Rat Schwann and astroglial cells and pig lens epithelial cells were used for transfer and the cells were cultured for 2 weeks after laser-pulsed transfer. All three cell types survived, proliferated, and differentiated under cell culture conditions and regained their original phenotype a few days after cell transfer. Time resolution studies have shown that the time required to accelerate the jets and droplets containing the cells was less than 1 micros and that the estimated minimum average acceleration of those ejected cells that reached a constant velocity was approximately 10(7) x g. This suggests that the majority of studied cells tolerated the extremely high acceleration at the beginning of the ejection and the deceleration during impact on the acceptor plate without significant damage to the original phenotype. These results suggest that the absorbing film-assisted laser-induced forward transfer technique appears to be suitable for several potential applications in tissue engineering and the biomedical tissue repair technologies.

Synapse loss precedes cell death in Alzheimer's disease, but the timing of axon degeneration relative to these events, and the causal relationships remain unclear. Axons become so severely dystrophic near amyloid plaques that their interruption, causing permanent loss of function, extensive synapse loss, and potentially cell death appears imminent. However, it remains unclear whether axons are truly interrupted at plaques and whether cell bodies fail to support their axons and dendrites. We traced TgCRND8 mouse axons longitudinally through, distal to, and proximal from dystrophic regions. The corresponding neurons not only survived but remained morphologically unaltered, indicating absence of axonal damage signalling or a failure to respond to it. Axons, no matter how dystrophic, remained continuous and initially morphologically normal outside the plaque region, reflecting support by metabolically active cell bodies and continued axonal transport. Immunochemical and ultrastructural studies showed dystrophic axons were tightly associated with disruption of presynaptic transmission machinery, suggesting local functional impairment. Thus, we rule out long-range degeneration axons or dendrites as major contributors to early synapse loss in this model, raising the prospect of a therapeutic window for functional rescue of individual neurons lasting months or even years after their axons become highly dystrophic. We propose that multi-focal pathology has an important role in the human disease in bringing about the switch from local, and potentially recoverable, synapse loss into permanent loss of neuronal processes and eventually their cell bodies.

Wallerian degeneration of the CNS is accompanied by axonal dystrophy or swelling. To understand the mechanisms by which swellings arise, we studied their spatiotemporal dynamics, ultrastructure, composition, and the conditions that affect their formation in vivo and ex vivo. In contrast to peripheral nerve axons, lesioned optic nerve (ON) axons in vivo developed focal swellings asynchronously within 6 hours, long before there is any axon fragmentation. Axons in ON, spinal cord dorsal column, and corpus callosum all showed marked gradients with more swellings in proximal regions of their distal stumps early after lesion. Time-lapse imaging of a validated ex vivo system showed that multiple focal swellings arise after around 1 hour close to the injury site, followed by anterograde wave-like progression on continuous ON axon stumps. Swellings were largely stable but occasionally seemed to fuse with neighboring swellings. Their ultrastructural appearances resembled disease-associated spheroids. Although accumulation of axonal markers suggested transport deficits, large accumulations of mitochondria were not observed. Early swelling formation was decreased in Wld gene-expressing rodents and by removing extracellular calcium. Several pharmacologic agents that inhibit axon loss in vitro and/or in vivo also prevented early formation of axonal spheroids in acute ON explants. Because axonal swellings are hallmarks of many neurodegenerative conditions, these data suggest that they are a manifestation of Wallerian-like degeneration in some cases. Thus, Wallerian-like degeneration may be a more common component mechanism in CNS diseases than previously thought.