OBJECTIVE: To define how to preserve the severed limbs to prolong the period of replantation. METHODS: The original articles about preservation of severed limbs in recent years were reviewed, it was suggested that the period of replantation was determined by the injury of skeletal muscle. RESULTS: When the environment of severed limbs was changed, the injures of skeletal muscle could be decreased. CONCLUSION: After the severed limbs are reasonably preserved, the period of replantation may be prolonged.
Objective To explore the in vitrodifferentiation of the rat mesenchymal stem cells (MSCs ) into the skeletal muscle cells induced by the myoblast differentiation factor (MyoD) and 5-azacytidine. Methods The MSCs were taken from the rat bone marrow and the suspension of MSCs was made and cultured in the homeothermia incubator which contained 5% CO2at 37℃. The cells were observed under the inverted phase contrast microscope daily. The cells spreading all the bottom of the culture bottle were defined as onepassage. The differentiation of the 3rd passage of MSCs was induced by the combination of 5-azacytidine, MyoD, transforming growth factor β1, and the insulin like growth factor 1. Nine days after the induction, the induced MSCs were collected, which were analyzed with the MTT chromatometry, theflow cytometry, and the immunohistochemistry. Results The primarily cultured MSCs grew as a colony on the walls of the culture bottle; after the culture for 5-7 days, the cells were shaped like the fibroblasts, the big flat polygonal cells, the medium sized polygonal cells, and the small triangle cells; after the culture for 12 days, the cells were found to be fused, spreadingall over the bottle bottom, but MSCs were unchanged too much in shape. After the induction by 5-azacytidine, some of the cells died, and the cells grew slowly. However, after the culture for 7 days, the cells grew remarkably, the cell volume increased gradually in a form of ellipse, fusiform or irregularity. After theculture for 14 days, the proliferated fusiform cells began to increase in a great amount. After the culture for 18-22 days, the myotubes increased in number and volume, with the nucleus increased in number, and the newly formed myotubes and the fusiform myoblst grew parallelly and separately. The immunohistochemistry for MSCs revealed that CD44 was positive in reaction, with the cytoplasm ina form of brown granules. And the nucleus had an obvious border，and CD34 was negative. The induced MSCs were found to be positive for desmin and specific myoglobulin of the skeletal muscle. The flow cytometry showed that most of the MSCs and the induced MSCs were in the stages of G0/G1，accounting for 79.4％ and 62.9％，respectively; however, the cells in the stages of G2/S accounted for 20.6％ and 36.1％. The growth curve was drawn based on MTT，which showed that MSCs weregreater in the growth speed than the induced MSCs. The two kinds of cells did not reach the platform stage，having a tendency to continuously proliferate.ConclusionIn vitro,the rat MSCs can be differentiated into the skeletal muscle cells with an induction by MyoD and 5-azacytidine, with a positive reaction for the desmin and the myoglobulin of the skeletal muscle. After the induction, the proliferation stage of MSCs can be increased, with a higher degree of the differentiation into the skeletal muscle.
Objective To review the current researches of scaffold materials for skeletal muscle tissue engineering, to predict the development trend of scaffold materials in skeletal muscle tissue engineering in future. Methods The related l iterature on skeletal muscle tissue engineering, involving categories and properties of scaffold materials, preparative techniqueand biocompatibil ity, was summarized and analyzed. Results Various scaffold materials were used in skeletal muscle tissue engineering, including inorganic biomaterials, biodegradable polymers, natural biomaterial, and biomedical composites. According to different needs of the research, various scaffolds were prepared due to different biomaterials, preparative techniques, and surface modifications. Conclusion The development trend and perspective of skeletal muscle tissue engineering are the use of composite materials, and the preparation of composite scaffolds and surface modification according to the specific functions of scaffolds.
The human skeletal muscle drives skeletal movement through contraction. Embedding its functional information into the human morphological framework and constructing a digital twin of skeletal muscle for simulating physical and physiological functions of skeletal muscle are of great significance for the study of "virtual physiological humans". Based on relevant literature both domestically and internationally, this paper firstly summarizes the technical framework for constructing skeletal muscle digital twins, and then provides a review from five aspects including skeletal muscle digital twins modeling technology, skeletal muscle data collection technology, simulation analysis technology, simulation platform and human medical image database. On this basis, it is pointed out that further research is needed in areas such as skeletal muscle model generalization, accuracy improvement, and model coupling. The methods and means of constructing skeletal muscle digital twins summarized in the paper are expected to provide reference for researchers in this field, and the development direction pointed out can serve as the next focus of research.
Objective To review researches of the role of inhibitorof differentiation 2(Id2) in skeletal muscle regeneration. Methods The latest original literature concerning Id2 and its role in skeletal muscle regeneration was extensively reviewed. Results Id2 could form heterodimers by combining with E protein to prevent myogenic regulatory factors (MRFs) forming heterodimers by combining with E protein, to inhibit the transcription activity of MRFs anddifferentiation of skeletal muscle cell. Conclusion Id2 plays an important role in skeletal muscle regeneration.
OBJECTIVE: To observe the changes of heme oxygenase-1 (HO-1) expression in the skeletal muscle after ischemia-reperfusion of hind limb in rats. METHODS: A model of hind limb ischemia was made by clamping femoral artery with a microvascular clip. Soleus muscle was obtained from the animals received sham operation, 4 h ischemia without reperfusion and 2 h, 4 h, 8 h, 16 h, 24 h reperfusion after 4 h ischemia. Soleus histology and malondialdehyde (MDA) content were measured. The levels of HO-1 mRNA and protein were measured in different time by Northern blotting, Western blotting and immunohistochemistry technique. RESULTS: After ischemia-reperfusion of limb, HO-1 mRNA increased at the 2nd hour, reached a peak at the 8th hour, and returned toward baseline at the 24th hour. The change of protein level was essentially in agreement with that of mRNA. Immunohistochemical results showed that HO-1 expressed primarily in skeletal muscle cytoplasma. There were no positive signals of mRNA and protein in sham group and in ischemia group. After limb reperfusion, MDA contents in the soleus muscle increased significantly when compared with that in the sham group (P lt; 0.05). MDA content of the 8th after reperfusion decreased significantly when compared with that of the 4 h after reperfusion (P lt; 0.05). CONCLUSION: Ischemia-reperfusion can induce HO-1 expression in skeletal muscle in rats, which may provide protection for injured tissue.
OBJECTIVE To observe the ultrastructural changes and number of satellite cells in different muscles with different denervation interval and investigate the mechanism of denervation atrophy. METHODS Muscles of different denervation interval were harvested, which were 6 biceps brachii and 6 abductor digiti minimi. The ultrastructure of the samples were observed under transmission electron microscope. The number of nucleus and satellite cells were counted to calculate the percentage content of satellite cells. RESULTS In early stage of denervation, the myofilament and sarcomere of the majority were well oriented. The nucleoli of some muscle cell nucleus were enlarged and pale. Vacuolarization was also seen in some mitochondria. There was no obvious proliferation of collagen fiber around myofibers. After denervation of half a year, rupture and disorientation of myofilament was seen. The nucleus became smaller, dark stained, and some of them were condensed. There was proliferation of fibroblasts, adipose cells and collagen fibers around myofibers. Motor endplate was not recognized one year after denervation. In the early stage of denervation, satellite cell percentage of the two muscles was relatively high. It then declined with time. One year after denervation, satellite cells were scarcely detected. Comparison of the curves for satellite cell declination in two muscles revealed that the declination of the abductor digiti minimi was faster than that of biceps brachii. Decrease of the former started 3 months after denervation, while the latter started after 6 months. CONCLUSION Disappearing of motor endplate and proliferation of collagen fibers are main factors that affect the treatment outcome in late cases. Decrease of satellite cell number is another cause. The correlation of less satellite cell in abductor digiti minimi and poorer recovery of hand intrinsic muscles indicates that increment of satellite cells in long-term denervated muscles may be one of the effective measures to improve treatment outcome.
Objective To investigate the influence of clenbuterol on the expression of nerve growth factor (NGF) in denervated red and white muscles and the neurotrophism of the denervated muscles.Methods Sixty-four Wister rats, weighed 200-250 g, were divided into 8 groups(8 rats per group), including 4 experimental groups and 4 control groups. The denervated model was made in rats by dissection of sciatic nerves. Clenbuterol was given at a dose of 200 μg/kg per day in the experimental group, saline in the control group. The expression of NGF was measured with immunohistochemistry after 1, 3, 7 and 14 days of injury. The culture methods of dorsal root ganglions of the chick embryos were used to measure the neurotrophism of extracts of the muscles. Results Compared with the control groups, the NGF content of gastrocnemious(GAS) increased on the 1st day (Plt;0.05) and the NGF content of soleus(SOL) increased greatly on the 1st, 3rd and 7th dayafter injury in the experimental groups (Plt;0.01). In the experimental groups, the NGF amount of GAS reached the highest value on the 1st day after injury(Plt;0.01) and then decreased gradually. And the NGF amount of SOL had slight difference between different time. The NGF content of the SOL was higher than that of GASon the 7th day (Plt;0.05). The sensory neurotrophism of the extracts was similar between SOL and GAS.Conclusion Clenbuterol can change the expression of NGF in denervated muscles, but the change was different in SOL and GAS. The sensory neurotrophism of the denervated muscles were determined by all of the neurotrophic factors in them.
In order to study the influence of reperfusion following ischemia on microvesseles and microcirculation of skeletal muscle, unilateral hindlimbs of 16 rabbits were subjected to normothermic ischemia for 2 and 5 hours by tourniquet. After release of the tourniquet, microcirculation of the peritenon on dorsum of the foot was observed for 1 hours by intravital microscope. At 1 hour and 72 hours following reperfusion, the anterior tibia muscle biopsiy were taken and the specimens were subjected to light and electron microscopic examinations. It was found that after release of the tourniquet, in the limbs undergone 2 hours ischemia, there was immediate and well distributed reflow in the microvesseles of peritenon though a few aggregates of red cells and increase in the number of adherent leukocytes occured in some venules, and the microvesseles of the skeletal muscle only showed signs of minimal injury, the muscle fibers could survive in the limbs undergone 5 hours of ischemia, however, there was serious disturbance of microcirculation in theperitenon, which was characterized by "no reflow" in most area and there was signi ficant increase in the number of leukocytes adherent to venular endothelium, and the microvesseles of the skeletal muscle showed signs of severe injury, including remarkable swelling of the endothelial cell, disruption of the basement membrane and interstitial edema, and finally, most of the muscle fibers had necrosis occured. The results demonstrated that reperfusion following ischimia might result in microvascular injury and microcirculation disorder in the ischemic area. The degree of the injury and disorder depended on the duration of ischemic period, and was an important factor which determined the fate of the parenchymal cell.
Objective To study the effect of motor nerve implantation after ectopic transplantation of skeletal muscle on nerve regeneration in rat. Methods Sixty Sprague-Dewley male 8 monthold rats were randomly divided into 3 groups: control group,in situ implantation group and ectopic transplantation group. In control group, obturator nerve controlling right gracilis was cut off. In in situ implantation group, the right gracilis was cut off and replanted to its original site, and the obturator nerve was implanted to the muscle. In ectopic transplantation group, the right gracilis was cut off and transplanted to the muscle of the left leg, and the obturator nerve was implanted to the muscle. After 25 weeks, the neurophysiological information was collected through electromyography and the weight of the muscle was measured. Results The potentialwithout control of the nerve existed in control group. There were no significant differences in latency, amplitude and conduct velocity betweenin situ implantation group and ectopic transplantation group(Pgt;0.05).The atrophy of gracilis was dominant incontrol group, the weight of the muscle was 158.0±19.3 mg. The weights of the muscle were 509.6±14.5 mg in ectopic transplantation group and 516.8±12.7 mg in in situ mplantation group, showing no significant difference (P＞0.05). The weights of the muscle in in situ implantation and ectopic transplantation group were larger than that in control group, showing significant difference(P＜0.05). Conclusion Motor nerve implantation after ectopic transplantation of skeletal muscle could prevent the atrophy of the muscle and resume partial function of nerve.