The effect of exercise on mobilization of hematopoietic progenitor cells involved in the repair of sciatic nerve crush injury [RETRACTED]

Laboratory investigation

Restricted access

Object

Mobilization of hematopoietic progenitor cells (HPCs) from bone marrow involved in the process of peripheral nerve regeneration occurs mostly through deposits of CD34+ cells. Treadmill exercise, with either differing intensity or duration, has been shown to increase axon regeneration and sprouting, but the effect of mobilization of HPCs on peripheral nerve regeneration due to treadmill exercise has not yet been elucidated.

Methods

Peripheral nerve injury was induced in Sprague-Dawley rats by crushing the left sciatic nerve using a vessel clamp. The animals were categorized into 2 groups: those with and without treadmill exercise (20 m/min for 60 minutes per day for 7 days). Cytospin and flow cytometry were used to determine bone marrow progenitor cell density and distribution. Neurobehavioral analysis, electrophysiological study, and regeneration marker expression were investigated at 1 and 3 weeks after exercise. The accumulation of HPCs, immune cells, and angiogenesis factors in injured nerves was determined. A separate chimeric mice study was conducted to assess CD34+ cell distribution according to treadmill exercise group.

Results

Treadmill exercise significantly promoted nerve regeneration. Increased Schwann cell proliferation, increased neurofilament expression, and decreased Schwann cell apoptosis were observed 7 days after treadmill exercise. Elevated expression of S100 and Luxol fast blue, as well as decreased numbers of vacuoles, were identified in the crushed nerve 3 weeks after treadmill exercise. Significantly increased numbers of mononuclear cells, particularly CD34+ cells, were induced in bone marrow after treadmill exercise. The deposition of CD34+ cells was abolished by bone marrow irradiation. In addition, deposits of CD34+ cells in crushed nerves paralleled the elevated expressions of von Willebrand factor, isolectin B4, and vascular endothelial growth factor.

Conclusions

Bone marrow HPCs, especially CD34+ cells, were able to be mobilized by low-intensity treadmill exercise, and this effect paralleled the significant expression of angiogenesis factors. Treadmill exercise stimulation of HPC mobilization during peripheral nerve regeneration could be used as a therapy in human beings.

Abbreviations used in this paper:BrdU = bromodeoxyuridine; CD = cluster of differentiation; CMAP = compound muscle action potential; GAP = growth-associated protein; GAPDH = glyceraldehyde-3-phosphate-dehydrogenase; G-CSF = granulocyte colonystimulating factor; GFP = green fluorescent protein; HPC = hematopoietic progenitor cell; PBS = phosphate-buffered saline; PMN = polymorphonuclear neutrophil; SFI = sciatic function index; VEGF = vascular endothelial growth factor.

Article Information

Address correspondence to: Hung-Chuan Pan, M.D., Ph.D., Department of Neurosurgery, Taichung Veterans General Hospital, No. 160, Taichung Port Road, Sec. 3, Taichung 407, Taiwan. email: hcpan2003@yahoo.com.tw.

Please include this information when citing this paper: published online November 23, 2012; DOI: 10.3171/2012.8.JNS111580.

© AANS, except where prohibited by US copyright law.

Headings

Figures

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    Evaluation of SFI, electrophysiological function (nerve conduction latency and CMAP), and muscle weight (MW) after treadmill exercise. Upper: Comparison of SFI scores after crush injury and after treadmill exercise on different days. Lower: Bar graph comparing the results of crush injury and treadmill exercise for the variables CMAP, nerve conduction latency, and muscle weight. *p < 0.05, **p < 0.01, ***p < 0.001. Lt/Rt = left/right.

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    CatWalk analysis of crush injury and treadmill exercise at different time points for the variables mean intensity (A), print area (B), stance duration (C), and swing duration (D). L/R = left/right.

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    Neurofilament (NF) expression and results of TUNEL and BrdU assay in crushed nerve 7 days after treadmill exercise. A: Representative photomicrographs of results of staining for neurofilament, TUNEL, and BrdU assay in crushed nerves. Bar = 50 μm. Insets: Merging of TUNEL and BrdU staining with S100 (red). B–D: Quantitative analyses of neurofilament expression level (B), TUNEL results (C), and BrdU staining (D).

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    Vacuole counts and S100 and Luxol fast blue staining 3 weeks after treadmill exercise. A: Representative photomicrographs illustrating vacuole, S100, and Luxol fast blue staining in each group. Bar = 50 μm. B–D: Quantitative analyses of vacuole counts (B), Luxol fast blue staining (C), and S100 staining (D).

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    Cytospin analysis of bone marrow (BM) aspirants 7 days after treadmill exercise. A: Representative examples of H & E, CD34+, and CD68+ staining. Bar = 50 μm. B: Quantitative analyses of mononuclear cell counts in each group. C: Quantitative analysis of CD34+ and CD68+ cell density in each group.

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    Mobilization and ablation of CD34+ cells in chimeric mice by treadmill exercise. Photomicrographs show CD34+ cells in crushed nerve (A, D, and G), GFP-positive cells in crushed nerve (B, E, and H), and merged images (C, F, and I). Bar = 50 μm. J: Quantitative analysis of CD34+ counts in the crushed nerve, treadmill, and treadmill+irradiation groups.

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    Deposits of immune cells and expression of angiogenesis factors in injured nerves 7 days after exercise. A: Photomicrographs of CD34+ and CD68+ cell deposits, as well as von Willebrand factor (VW) and isolectin B4 staining. Bar = 50 μm. B: Quantitative analyses of immune cell deposits in the crush and treadmill groups. C: Quantitative analyses of angiogenesis factors in the crush and treadmill groups.

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    Western blot analysis in the retrieved nerve 7 days after treadmill exercise. Left: Western blot for CD68+, CD34+, GAP43, and VEGF in the 2 groups. C = left crushed sciatic nerve; R = right sciatic nerve as a control. Right: Graph showing the quantitative results of Western blot analysis with respect to GAPDH.

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