title: Optimal freezing and thawing for the survival of peripheral nerves in severed rabbit limbs.
abstract: This study aimed to investigate the optimal freezing and thawing procedures for the survival of peripheral nerves in severed rabbit limbs. Twenty New Zealand White rabbits were randomized into four groups: normal control, slow-freezing fast-thawing, slow-freezing slow-thawing, fast-freezing fast-thawing, with five animals in each group. The hind limbs of the rabbits were severed at 1 cm above the knee joint. The severed limbs were cryopreserved with various freezing and thawing procedures. The sciatic nerves were harvested and trypsinized into single nerve fibers for morphological evaluation. The cell viability of the nerve fibers was examined by staining with Calcein-AM and propidium iodide. The fluorescent intensity of the nerve fibers was measured with a laser scanning confocal microscope. The morphology of the nerve fibers in the slow-freezing fast-thawing group was very similar with that of the normal control group, with only mild demyelination. The slow-freezing fast-thawing group and slow-freezing slow-thawing group showed severely damaged nerve fibers. The fluorescent intensities of the nerve fibers was significantly different among the four groups, with a decreasing order of normal control, slow-freezing fast-thawing, slow-freezing slow-thawing, and fast-freezing fast-thawing (P < 0.05). Of the various cryopreservative procedures, slow-freezing fast thawing has the minimal effects on the survival of nerve fibers in severed rabbit limbs.
authors.affiliation: Department of Orthopaedics, General Hospital of The Second Artillery Beijing 100088, China.
authors.forename: Zexing
authors.initials: Z
authors.lastname: Zhu
authors.affiliation: Department of Orthopaedics, General Hospital of The Second Artillery Beijing 100088, China.
authors.forename: Lin
authors.initials: L
authors.lastname: Qiao
authors.affiliation: Department of Orthopaedics, Center Hospital of Linfen Linfen 041000, Shanxi Province, China.
authors.forename: Yandong
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Department of Orthopaedics, General Hospital of The Second Artillery Beijing 100088, China.
authors.forename: Shuming
authors.initials: S
authors.lastname: Zhang
mesh_terms: D000818:Animals; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods*; D005615:Freezing; D006614:Hindlimb / Q000294:innervation*; D009412:Nerve Fibers; D010525:Peripheral Nerves; D011817:Rabbits
keywords: Cryopreservation; fluorescent intensity; nerve fibers; rabbit limbs; survival