Relationship between Changes of Pericyte Number and Wound Age during Repair of Skeletal Muscle Contusion in Mice / 法医学杂志

Objective To observe the change pattern of pericyte number at different time periods after mice skeletal muscle contusion and discuss its role in wound age estimation. Methods A mice gastrocnemius muscle contusion model was established. The form and number changes of pericytes at 1, 3, 5, 7, 9, 14, and 28 d post-injury were detected by multiple immunofluorescence staining. Results Compared with the slender shape of pericytes in normal skeletal muscles, pericytes in the contusion area had increased volume, rounder form and a round nuclei. Part of pericytes were found to express satellite cell markers paired-box transcription factor （Pax7） or myoblast determination 1 （MyoD1）. The changes of pericyte number in skeletal muscles after contusion were time-dependant, and showed unimodal distribution with the extension of wound age. In the central contusion area, the number of pericytes peaked at 5 d post-injury while in the peripheral contusion area, the number of pericytes peaked at 5 d and 7 d post-injury. Conclusion The number of pericytes in contusion area varies time-dependently after skeletal muscle contusion in mice and might be a reference index for muscle wound age estimation, and is involved in the repair and regeneration of skeletal muscle injury.

Comparison of the efficacy of losartan, hydrocortisone and acetylsalicylic acid (ASA) in preventing the development of fibrous scar tissue in skeletal muscle / Comparação da eficácia de losartana, hidrocortisona e ácido acetilsalicílico (AAS) na prevenção do desenvolvimento de tecido cicatricial fibroso nos músculos esqueléticos / Comparación de la eficacia de losartán, hidrocortisona y ácido acetilsalicílico (AAS) en la prevención del desarrollo de tejido cicatricial fibroso en los músculos esqueléticos

OBJECTIVE: To analyze fibrous scar tissue inhibition capacity with the use of losartan, hydrocortisone and acetylsalicylic acid. METHOD: The sample consisted of 120 male heterogeneic Wistar rats with a muscle laceration model. The rats were divided into four groups of 30 animals each: control group, losartan group, ASA group and hydrocortisone group. The animals were anesthetized and a 2.5 cm longitudinal incision was made in the left thoracolumbar paravertebral region. The muscles were subjected to a Grade III lesion caused by applying Kelly hemostatic forceps for 60 seconds, followed by sectioning with scissors. The skin was sutured with 3-0 nylon monofilament thread. The animals were placed in individual cages with plenty of food and water. The losartan group received losartan diluted in water at a dose of 0.1 mg/mL (10 mg/kg/day), the ASA Group received a 3 mg/mL ASA solution (300 mg/kg/day), and the hydrocortisone group received a 0.2 mg/mL hydrocortisone solution (20 mg/kg/day). RESULTS: The control, losartan, hydrocortisone and aspirin groups had a fibrotic area of 0.95 ± 0.35 mm, 0.55 ± 0.34 mm, 0.93 ± 0.33 mm, and 0.66 ± 0.36 mm, respectively. We observed a significantly smaller fibrotic area in the losartan group compared to the control (p=0.01) and hydrocortisone (p=0.01) groups. There were no significant differences among the other groups. CONCLUSION: The healing of striated skeletal muscle produced less fibrous scar tissue when exposed to losartan in comparison to the control group or the hydrocortisone group. Level of Evidence I; Randomized double-blind placebo-controlled study.

OBJETIVO: Analisar a capacidade de inibição de formação de tecido cicatricial fibroso com losartana, hidrocortisona e AAS. MÉTODOS: A amostra consistiu em 120 ratos Wistar heterogênicos machos com modelo de laceração muscular. Os ratos foram distribuídos em quatro grupos de 30 animais: grupo controle, grupo losartana, grupo AAS e grupo hidrocortisona. Os animais foram anestesiados e submetidos a uma incisão em sentido longitudinal de 2,5 cm de extensão na região paravertebral toracolombar esquerda, e os músculos sofreram uma lesão grau III com pinça hemostática de Kelly durante 60 segundos e posterior secção com tesoura. A pele foi suturada com nylon monofilamentar 3-0. Os animais foram colocados em gaiolas individuais, com água e alimento à vontade. O grupo losartana recebeu losartana diluída em água na dose de 0,1 mg/ml (10 mg/kg/dia), o grupo AAS recebeu solução de AAS 3 mg/ml (300 mg/kg/dia), o grupo hidrocortisona recebeu solução de hidrocortisona 0,2 mg/ml (20 mg/kg/ dia). RESULTADOS: Os grupos controle, losartana, hidrocortisona e AAS apresentaram área fibrótica de0,95 ± 0,35 mm, 0,55 ± 0,34 mm, 0,93 ± 0,33 mm, 0,66 ± 0,36 mm, respectivamente. Observou-se área fibrótica significativamente menor do grupo losartana em comparação com o grupo controle (p = 0,01) e hidrocortisona (p = 0,01). Nos demais grupos não houve diferença significativa. CONCLUSÃO: A cicatrização do músculo estriado esquelético produziu menos tecido cicatricial fibroso quando exposto à losartana do que quando comparado com o grupo controle ou o grupo hidrocortisona. Nível de Evidência I; Estudo duplo-cego randomizado controlado por placebo.

OBJETIVO: Analizar la capacidad de inhibición de formación de tejido cicatricial fibroso con losartán, hidrocortisona y AAS (ácido acetilsalicílico). MÉTODOS: La muestra consistió en 120 ratas Wistar heterogéneas machos con modelo de laceración muscular. Las ratas fueron distribuidas en cuatro grupos de 30 animales: grupo control; grupo losartán; grupo AAS y grupo hidrocortisona. Los animales fueron anestesiados y sometidos a una incisión longitudinal de 2,5 cm de extensión en la región paravertebral toracolumbar izquierda y los músculos sufrieron una lesión de grado III con pinza hemostática de Kelly durante 60 segundos y posterior sección con tijera. La piel se suturó con monofilamento de nylon 3-0. Los animales fueron dispuestos en jaulas individuales con abundante comida y agua. El grupo losartán recibió losartán diluido en agua a una dosis de 0,1 mg/ml (10 mg/kg/día), el grupo AAS recibió solución de AAS de 3 mg/ml (dosis 300 mg/kg/día), el grupo hidrocortisona recibió solución hidrocortisona de 0,2 mg/ml (20 mg/kg/día). RESULTADOS: Los grupos control, losartán, hidrocortisona y AAS mostraron área fibrótica de 0,95 ± 0,35 mm, 0,55 ± 0,34 mm, 0,93 ± 0,33 mm, 0,66 ± 0,36 mm, respectivamente. Se observó área fibrótica significativamente menor del grupo losartán en comparación con el grupo control (p = 0,01) e hidrocortisona (p = 0,01). En los demás grupos no hubo diferencias significativas. CONCLUSIÓN: La cicatrización del músculo estriado esquelético produjo menos tejido cicatricial fibroso cuando fue expuesto a losartán que cuando fue comparado con el grupo control o el grupo hidrocortisona. Nivel de Evidencia I; Estudio doble ciego aleatorio controlado por placebo.

Recent Advances in Skeletal Muscle Stem Cells for Duchenne Muscular Dystrophy Treatment / 한양의대학술지

Muscle stem cells, which are known as satellite cells have heterogeneous components of committed myogenic progenitors, non-committed satellite cells, and mesenchymal stem cells. This distinguishing organization of self-renewal and differentiation capacities encourages the remarkable regenerative ability of skeletal muscles. Lately it has been proved that the satellite cell is the derivation of muscle regeneration and with the self-renew function, it roles as a true muscle stem cell. Therefore, stem cell therapy using satellite cells is considered to be ideal therapy for muscular dystrophies, which is deficient in specific muscle protein and causes muscle degeneration. Especially, Duchenne Muscular Dystrophy (DMD), which is caused by mutations at the dystrophin gene, has been targeted by much research. In this article the satellite cell characteristics, regulation of cell function, and stem cell therapy for DMD and the present progressive clinical trials will be reviewed.

Effect of erytbropoietin on apoptosis and calcium ion concentration of skeletal muscle satellite cells injured by hydrogen peroxide in rats / 中华老年医学杂志

Objective To investigate the effect of hydrogen peroxide (H_2O_2) on apoptosis and calcium ion concentration of skeletal muscle satellite cells (SMSCs) in rats, and to explore the protective effect of erythropoietin (EPO).Methods The cultured SMSCs were divided into five groups: control group,H_2O_2 group, 10, 20 and 40 U/ml EPO intervention groups.Apoptosis rates and calcium ion concentration of SMSCs were analyzed by flow cytometry, and the morphology of apoptotic cells was observed by Hoechst33258 staining.Results The apoptosis rates showed significant differences (all P<0.05) among (1.93±0.57)% in control group, (22.13±1.79)% in H_2O_2 group, (16.47±2.53)%, (4.97±0.55)% and (2.93±0.47)% in 10, 20 and 40 U/ml EPO intervention groups, respectively.And calcium ion concentrations in SMSCs were 12.67 ±0.32, 27.90±0.06 and 44.53±0.93 in 10, 20 and 40 U/ml EPO intervention groups, respectively.There was significant difference in calcium ion concentration between H_2O_2 group and control group (9.70±0.09 vs.51.37± 0.64, P< 0.05).Morphology of apoptosis was observed by Hoeehst33258 dye stains in 10, 20 U/ml EPO intervention group and H_2O_2 group, while there were less apoptotic bodies in 40 U/ml EPO intervention group and control group.Conclusions EPO might have protective effects on SMSCs injured by H_2O_2 through inhibiting apoptosis and calcium ion releasing from SMSCs.

Estudo comparativo da ação da toxina botulínica tipo A e da crotoxina sobre as células satélites da musculatura extrínseca ocular em modelo animal / A comparative study of the effects of type A botulinum toxin and crotoxin on satellite cells of extraocular muscles in rabbits

OBJETIVOS: Avaliar o efeito da toxina botulínica do tipo A e da crotoxina na ativação de células satélites das fibras musculares de retos superiores de coelhos. MÉTODOS: Os músculos retos superiores do olho direito de 29 coelhos machos albinos neozelandeses foram inoculados com toxina botulínica do tipo A ou com crotoxina, em diferentes doses. Os músculos retos superiores contralaterais de cada animal foram inoculados com solução salina em volume igual ao das toxinas. Os animais foram sacrificados 12, 18 ou 25 dias após as aplicações. Os olhos foram enucleados e cada músculo foi preparado para análise imunoistoquímica, com marcadores de células satélites. Foi realizada contagem dos núcleos corados pelos marcadores a cada cem miofibras. RESULTADOS: A aplicação de toxina botulínica e de crotoxina provocou um aumento no número de células satélites ativadas e em proliferação nos músculos retos superiores. Uma maior ativação foi observada após a aplicação de crotoxina, embora, estatisticamente, a diferença do efeito de ativação entre os grupos botoxina e crotoxina não tenha sido significativa. Nos grupos botoxina e crotoxina, não houve correlação estatisticamente significativa entre a dose e o volume aplicados e o aumento na ativação das células. O tempo de vida após a aplicação contribuiu para o aumento das células ativadas nos grupos. CONCLUSÃO: A observação de maior desorganização na estrutura muscular e de sinais de regeneração mais evidentes no grupo crotoxina parece estar correlacionada ao aumento de células satélites ativadas.

PURPOSE: To evaluate the effect of botulinum toxin A and crotoxin on satellite cell activation in the muscle fibers of superior rectus muscles of rabbits. METHODS: The superior rectus muscles in the right eyes of 29 male, albino, New Zealand rabbits were inoculated with different doses of botulinum toxin A or crotoxin. The contra-lateral superior rectus muscles in each rabbit were inoculated with the same volume of saline solution only. The animals were sacrificed either 12, 18 or 25 days after the inoculation. The eyes were enucleated and subsequently, each muscle was prepared for immunohistochemical analysis, using satellite cell markers. The positive nuclei, revealed by the markers in each 100 myofibers, were counted. RESULTS: The application of the botulinum toxin A and crotoxina triggered a more significant increase satellite cell activation and proliferation in right superior rectus muscles in rabbits when compared with a saline solution inoculation in the contralateral muscles. Greater cell activation was observed after crotoxin application, although, statistically, the difference in the effects of this activation between the botoxin and crotoxin groups was not significant. There was no statistically significant correlation between the dose and the volume applied and resulting cell activation in the botoxin and crotoxin groups. Post-application survival time contributed to the increase in activated satellite cells in all groups. CONCLUSION: The observed increase in disorganization in the muscle structure, together with more obvious signs of regeneration in the crotoxina group, suggests a correlation with the increase in satellite cell activation.

Cliulcal research of myocardium infarction therapy by transplanting satellite cells / 中国医师杂志

Objective Satellite cells(SCs) transplant was tried to treat myocardium infarction(MI) in human.Methods 10 patients who affected by coronary artery disease(CAD) and MI were selected.A piece of muscle was taken away from their quadriceps femoris 2 weeks before surgical operation.SCs were extracted,purified,cultured by modified Dorfman method,and then transplanted into the MI area by intramyocardial injection after CABG.EKG,UCG,99cTc-SPECT cardiac blood pool imaging,18F-FDG myocardial perfusion imaging were examined before operation and 3 months after operation.The cell metabolism status in the MI area were observed and LVEF were measured and analyzed by paired-samples t test with SPSS13.0 statistics software.Results After cultured in vitro for3 days,SCs were spindleshaped,and sticked to walls.SCs growth was stasis,fused into each other,and formed myotube when the cells were delayed to divide Petri dish or the concentration of fetus bovine serum(FBS) in the culture medium was reduced.Average 2.7 blood vessel bypass were established in these patients.SCs were transplanted merely in one patient whose coronary arteries were extensively sclerosed.No patients died in hospital.Heart function of these patients were recovered to NYHA I 3 months after operation.Their symptoms of angina pectoris were disap peared and quality of life was improved obviously.The cell metabolism activity was appeared again in the MI area 3 months after operatio(including the patient without CABG).LVEF were increased(9.70±4.17)% than preoperation.There is significant deviation between them(P<0.01).Conclusions Autograft of SCs can be alive in the MI area,and SCs can improve the heart function by participating the heart's systole and diastole activity directly or by altering cardiac compliance and stimulating the blood capillary proliferation.

The effect of transplantation of skeletal muscle satellite cell on retard the denervated muscles atrophy / 中华骨科杂志

Objective To observe the effect which the skeletal muscle satellite cells were transplanted on denervated skeletal muscle atrophy so as to provide the experimental data for treating denervated muscle atrophy in vivo. Methods Thirty-two Sprague-Dawley rats weighting 200-250 g were randomly divided into control group and experimental group. Each group included 16 rats, the animal model of denervated gastrocnemius muscles were formed by cutting the right sciatic nerve of rats caused nerve despair about 1 cm. Muscle satellite cells were obtained from the dorsal and lower exterenity muscle of SD rats. Before transplantation, muscle satellite cells being labeled with DAPI (4′-6-Diamidino-2-Phenylin Dole) in vitro. Muscle satellite cells and NS were implanted into denervated skeletal muscle from the two groups. Bilateral gastrocnemius muscles of each rat from the two groups were taken and weighed at second and eighth post-operative weeks respectively. The above muscles underwent anti-actin immunohistochemical and HE staining and muscle cross-sectional area of fiber and the actin content of the rats were measured by the image analysis, the data of which was handled with SPSS software. Results The satellite cells and myofiber with fluorescence were observed in transplantation site. The group of skeletal muscle satellite cells transplanted when two weeks and eight weeks denervated gastrocnemius muscle wet weight remnant rate and muscle cross-sectional areas of fiber remnant rate and muscle actin content were better than in the control group respectively(P

Primary cultivation and identification of satellite cells of mice skeletal muscles / 解放军医学杂志

Objective To explore the reliable method for primary cultivation and identification of satellite cells of mice skeletal muscles in vitro.Methods Cell suspension was obtained from posterior limb skeletal muscles of newborn mice by mixed enzymatic digestion,and the muscle satellite cells were separated by Percoll-density gradient centrifugation and purified with differential adherence method.Cells growth was observed under an inverted microscope and the growth curve was drawn.Satellite cells were identified with monoclonal antibody against desmin and ?-actin.Results The primary satellite cells were round in shape,and began on adherence one day after cultivation,and the cells distributed uniformly,most of the cells were roundish.After cultured for 48 hours,the cells began on proliferation,and then became bigger and started to division,bubble cell arrangements with 2-3 or more cells were observed.At the third or fourth day of cultivation,the cells were in exponential phase of growth,and the round cells predominated,many clusters of cell clones were found.The satellite cells formed monolayer sheet(50%-70%) 7-10 days after cultivation,the fusiform cells predominated,but there were still some round cells in this phase.The cell proliferation decreased after cultured for 10-12 days.Immunocytochemical staining showed that the obtained cells expressed myogenic marker desmin and skeletal muscle-specific protein ?-actin.Conclusions High purity muscle satellite cells could be obtained by mixed enzymatic digestion,Percoll-density gradient centrifugation and differential adherence method.Immunocytochemical staining of desmin and ?-actin is the effective method to identify muscle satellite cells.