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Objective To study the effects of semaphorin 3A (Sema3A) on myoblast function and its transcriptional regulation mechanism. Methods Myoblasts were treated with 1.0 µg/ml, 0.1 µg/ml, 0.01 µg/ml recombinant Sema3A protein, and the control group was set up. The proliferation and migration of myoblasts were observed by the IncuCyte system and scratch test. The expression levels of myogenic differentiation factors were detected by RT-qPCR. Transcriptome sequencing was performed for the myoblasts intervened by 1.0 µg/ml recombinant Sema3A protein and those in control group. The differential expressed genes were screened out for GO enrichment analysis. Part of differential expressed genes were verified of transcription level by RT-qPCR. Results IncuCyte cell imaging count showed that Sema3A promoted proliferation of myoblasts, the cell count increased about 17.36% in 1.0 µg/ml Sema3A group than in control group (P<0.001). Scratch test showed the enhanced migration ability of myoblasts by Sema3A, the cell scratch healing rate increased by 69.66% in 1.0 µg/ml Sema3A group than in control group (P<0.001).RT-qRCR showed that during myoblast differentiation, 1.0 µg/ml Sema3A up-regulated the expression of Myf5 by 37.47%±11.67%(P<0.01), and up-regulated the expression of MyoG by 26.57%±11.31% (P<0.05). Bioinformatics analysis showed that Sema3A up-regulated the functional pathway associated with the development and maturation of muscle cells and muscle fibers, bone and chondrocyte proliferation, osteogenic differentiation, and bone mineralization; down-regulated the functional pathway associated with the lipoprotein metabolism and chylomicrons. RT-qPCR verified the partial differentially expressed genes in enrichment pathway, among which the expressions of Prox1 and Myh11 increased by 22.48%±14.42% (P<0.05) and 21.42%±5.81% (P<0.01),respectively; and of LipC and ApoC2 decreased by 23.08%±15.38% (P<0.05) and 55.97%±26.51% (P<0.05), respectively. Conclusion Exogenous Sema3A protein can up-regulate the expression of myoblast differentiation related genes Myf5 and MyoG, and may promote the proliferation and migration of myoblasts by transcriptional regulation of skeletal muscle development,maturation and lipoprotein metabolism.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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