【Abstract】 Objective To investigate the effect of massage on quadriceps femoris repair and the expressions of Desmin and α-Actin in rabbits so as to explore the possible molecular mechanisms of massage in repair of muscle injury. Methods Twenty-seven New Zealand white rabbits, weighing (2.0 ± 0.5) kg, were randomly divided into 3 groups: groups A (n=3), B (n=12), and C (n=12). In group A, the rabbits were not treated as controls; in groups B and C, the rabbit models of quadriceps femoris injury were prepared by self-made beater. In group B, no massage therapy was given as nature recovery controls; in group C, RT-N2 intelligent massage device was used for massage therapy at 8 days after injury, at 3 000-3 100 r/min for 15 minutes, every day for 7 days or for 14 days. The quadriceps femoris specimens were taken from 6 rabbits of groups B and C at 14 days and 21 days, respectively. HE staining was employed to detect the histomorphological change. Immunohistochemistry staining and Western blot were used to detect Desmin and α-Actin expressions. The massage therapy effect was evaluated by the histomorphological change and Desmin and α-Actin expressions. Results All rabbits survived to the end of experiment in groups B and C. No histological change was found with regular order of muscle fibers and no connective tissue in group A; obvious tissue necrosis was seen with broken muscular fibers, muscle atrophy, and irregular order in group B; and in group C, the skeletal muscle morphology and muscle atrophy were obviously improved with regenerated muscle fibers when compared with group B. Immunohistochemistry staining showed that the Desmin and α-Actin expressions obviously reduced in groups B and C, which were significantly weaker than that in group A (P lt; 0.05); the Desmin and α-Actin expressions were significantly ber in group C than in group B (P lt; 0.05), and at 21 days than at 14 days in group C (P lt; 0.05). Western blot results showed that the Desmin and α-Actin expressions were significantly higher in group A than in groups B and C (P lt; 0.05), and the expressions were lowest at 14 days in group B. Conclusion The histomorphology and cytoskeletal structure can be significantly improved after massage, which may help to repair muscle injury by up-regulation of Desmin and α-Actin expressions.
Objective To study whether human amniotic fluid colony derived stem cells (hAFCSCs) are involved in regeneration of injured muscles in mice and to investigate the method and feasibil ity of hAFCSCs-based cytotherapy in the treatment of injured muscles. Methods Human second-trimester amniotic fluid was collected through ultrasound-guided amniocentesis, hAFCSCs were isolated from second-trimester amniotic fluid and cultured, and the cells at 6th-8th passages were spared. The mRNA was extracted to identify the stem cell related genes by RT-PCR. The muscular injury model of bilateral tibial is anterior muscle was establ ished by cardiotoxin and X-ray irradiation in 16 Nod/Scid mice (aged 6-8 weeks, and weighing 20-24 g). The hAFCSCs (3.3 × 107/mL, 30 μL) were injected into the right injured tibial is anterior muscles as the experimental group, while the same volume of complete medium (α-MEM containing 15%FBS, 18%Chang B, 2%Chang C, 1% penicill instreptomycin, and 1% L-glutamine) was injected into the left injured tibial is anterior muscles as the control group. At 2 and 4 weeks after cell transplantation, the immunofluorescence staining of tibial is anterior muscles was performed to detect hepatocyte growth factor receptor (c-Met), myogenic regulatory factor (Myf-5), Laminin, Desmin, and human specific nuclear mitotic apparatus protein (NuMa). Results The clone formation was observed at 5-7 days of primary hAFCSCs culture; after 8-10 days, the clones with homogeneous morphology were selected for subculture. Adequate stem cells were available after 6th-8th subculture. RT-PCR analysis showed that hAFCSCs expressed mRNA of the stem cell related genes. The immunofluorescence double-staining showed that NuMa expressed in tibial is anterior muscles of the experimental group and no myogenic phenotype expressed at 2 weeks after cell transplantation, and that single cell co-expressed NuMa and c-Met or Myf-5 at 4 weeks after cell transplantation. In some myofibers, NuMa and Laminin or Desmin were also co-expressed. No NuMa positive hAFCSCs were detected in the control group at 2 and 4 weeks after cell transplantation. Conclusion hAFCSCs can participate in the regeneration of injured mouse muscle.