ABSTRACT
Objective:To observe the denervation and reinnervation of the neuromuscular junction (NMJ) and muscle spindle after sciatic nerve crush in mice.Methods:From January, 2019 to October, 2019, 18 C57BL/6 mice were randomly and evenly divided into injured groups (sciatic nerve crush group, 12 mice) and control groups (sham group, 6 mice). Mice in the injured group were sacrificed at day-1, day-2, day-3 and 4 weeks after the sciatic nerve injury. Mice in the control group were sacrificed at day-3 and 4 weeks from the day of experiment. The tibialis anterior muscles with crushed nerve were collected for immunohistochemistry staining by neurofilament (NF), synaptophysin (Syn) and α-bungarotoxin (α-BTX). Data were analysed by unpaired t-test. P<0.05 was considered significant difference. Results:The completely denervation of NMJ reached (92.4±8.85)% at day-3 after the nerve crush, compared with (5.19±1.32)% in the control group ( P<0.05). However, the muscle spindle lost γ-motor neuron innervation at day-2 after the nerve crush and completely denervated at day-3 after the surgery. For the reinnervation, no difference of no reinnervation of NMJ [(3.02±0.78)% vs. (4.22±2.08)%], partial reinnervation of NMJ [(6.44±1.91)% vs. (7.94±2.12)%] and completely reinnervation of NMJ [(90.54±10.44)% vs. (87.84±13.89)%] were observed between the control group and the injured group 4 weeks after the injury. While, for the muscle spindle, only the acetylcholine receptors at each end were partly reinnervated. The innervation of the middle part of the muscle spindle was thin and discontinued without typical spiral winding structure. Conclusion:The denervation speed of NMJ and muscle spindle was comparable after the peripheral nerve injury. The γ-motor neuron lost innervation ahead of the sensory neuron in the muscle spindle. The reinnervation of NMJ is earlier than that of the muscle spindle, and the re-innervation of the γ-motor neuron is prior to the sensory neuron in the muscle spindle.
ABSTRACT
Objective:To explore the expressions of Disabled-1 (Dab1 )in human breast epithelial cells and breast cancer cells,and to clarify its role in cell cycle.Methods:Real-time PCR was used to analyze the Dab1 mRNA expressions in breast epithelial cells MCF-10A and breast cancer cells MCF-7,BT-549,and MDA-MB-231. The Dab1 protein expressions in those cells were tested by Western blotting method. The BT-549 cells at logarithmic growth phase were divided into control,pKH3,and pKH3-Dab1 groups;the cell cycle was investigated by flow cytometry.Results:The Real-time PCR results showed that the Dab1 mRNA expression levels in MCF-7 cells (0.504 ± 0.037),BT-549 cells (0.302 ± 0.027),and MDA-MB-231 cells (0.330 ± 0.031 )were reduced compared with MCF-10A cells (0.998±0.020)(P <0.05).The Western blotting results showed that the Dab1 protein expression levels in breast cancer cells MCF-7 (0.134±0.014),BT-549 (0.076±0.01),and MDA-MB-231 (0.074±0.005)were reduced compared with MCF-10A cells (0.227±0.021)(P <0.05).Compared with control group and pKH3 group,the cell cycle in pKH3-Dab1 group was inhibited at G1 phase detected by flow cytometry analysis. Conclusion:The expression of Dab1 is down-regulated in breast cancer cells,and the over-expression of Dab1 can inhibit the cell cycle at G1 phase.