Time-dependent translocational changes of Synapsin I (SyI), a synaptic vesicle-associated phosphoprotein and its involvement in the axonal transport were investigated in the regenerating axonal sprouts. A weak SyI immunoreactivity (IR) was found in the axoplasm of normal axons. Rat sciatic nerves were crush-injured by ligating with 1-0 silk thread at the mid-thigh level and released from the ligation 24 h later. At various times after release, immunocytochemistry was performed. SyI was translocated from the proximal to the distal site of ligation and also involved in the sprouting of regenerating axons. The distribution patterns of SyI IR were changed in the crush-injured nerves. SyI immunoreactive thin processes were strongly appeared in the proximal region from 1 h after release. After 3 h, a very strong IR was expressed. The intense SyI immunoreactive thin processes were elongated distally and were changed the distribution pattern by time-lapse. After 12 h, strong immunoreactive processes were extended to the ligation crush site. At 1 day, a very intense IR was expressed. At 2 days, immunoreactive thin processes extended into the distal region over the ligation crush site and strong IR was observed after 3 days. SyI was accumulated in the proximal region at the early phases after release. These results suggest that SyI may be related to the translocation of vesicles to the elongated membranes by a fast axonal transport in the regenerating sprouts.
Animals ; Axonal Transport ; Axons/*physiology/ultrastructure ; Immunohistochemistry ; Male ; Nerve Crush ; Nerve Regeneration/*physiology ; Protein Transport ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve/physiology ; Synapsins/*metabolism ; Time Factors

OBJECTIVETo culture interleukin-1beta (IL-1beta)-activated Schwann cells (SCs) with human hair keratins (HHKs) for artificial nerve bridge construction.

METHODSSCs purified by primary culture with or without IL-1beta activation were cultured with HHKs decorated by extracellular matrix (ECM), and the artificial nerve bridge was implanted into the defect of rat sciatic nerve. The morphology of the SCs cultured with HHKs was monitored by inverted microscope, scanning electron microscope and evaluated by immunocytochemical staining, and the expression of nerve growth factor (NGF) in the sciatic nerve was observed by in situ hybridization.

RESULTSActivated SCs showed better ability to adhere to the HHKs and grew well. The HHKs component in the artificial nerve bridge underwent degradation in the sciatic nerve defect after 3 to 4 weeks, and IL-1beta activation resulted in enhanced NGF expression in the SCs.

CONCLUSIONThe constructed artificial nerve bridge by three-dimensional culture of IL-1beta-activiated SCs with HHKs decorated by ECM promotes the repair of sciatic nerve defects and accelerates sciatic nerve regeneration.

Animals ; Animals, Newborn ; Axons ; physiology ; Cell Culture Techniques ; Cell Movement ; physiology ; Cells, Cultured ; Hair ; chemistry ; Humans ; Interleukin-1beta ; pharmacology ; Keratins ; pharmacology ; Microscopy, Electron, Scanning ; Nerve Growth Factor ; biosynthesis ; Nerve Regeneration ; drug effects ; Rats ; Rats, Sprague-Dawley ; Schwann Cells ; drug effects ; metabolism ; ultrastructure ; Sciatic Nerve ; injuries ; physiopathology ; surgery ; Tissue Engineering ; methods