Tetrodotoxin(TTX)is a potent neurotoxin firstly discovered in the ovary of pufferfish.The genetic basis of voltage-gated sodium channel resistance to TTX has been widely studied,but it remains unclear in the evolution...Tetrodotoxin(TTX)is a potent neurotoxin firstly discovered in the ovary of pufferfish.The genetic basis of voltage-gated sodium channel resistance to TTX has been widely studied,but it remains unclear in the evolution history of voltage-gated sodium channel resistance to TTX in pufferfish with different TTX concentrations.In this study,six scn4aa coding sequences of pufferfish were firstly cloned and sequenced,then used to investigate the adaptive evolution of scn4aa associated with TTX concentration and reconstruct ancestral sequences with seven scn4aa of other fishes.The result of CODEML(codon substitution model)program from the PAML(phylogenetic analysis by maximum likelihood)package shows only in the genus of Takifugu,which contains TTX highly in the liver,under positive selection.The result also indicates that three of four positively selected sites are located in the intracellular regions,which may compensate for normal function.The ancestral sequence reconstruction may suggest that the replacements providing weak toxin resistance might have appeared first in scn4aa,then the genus Takifugu evolved stronger resistance to TTX later.These results contribute to the explanation of the evolutional history of voltage-gated sodium channel resistance to TTX in pufferfish.展开更多
Objective The present study is to observe in vitro the proliferation ability of the muscle cells from permanent myopathy （PM） patients of nomokalaemic periodic paralysis （normKPP）, which is caused by mutations of ...Objective The present study is to observe in vitro the proliferation ability of the muscle cells from permanent myopathy （PM） patients of nomokalaemic periodic paralysis （normKPP）, which is caused by mutations of Metl592Val in the skeletal muscle voltage gated sodium channel （SCN4A） gene on chromosome 17q23.1. We also evaluate the possible effect of the foreign basic fibroblast growth factor （bFGF） in preventing and curing PM. Methods The gastrocnemius muscle cells were taken from two male patients with PM of the same Chinese family with Metl592Val mutation of SCN4A, determined by gene screening. Four male patients suffering from the skeletal injury without PM were taken as control. All preparations were protogenerationally cultured in vitro. Proliferation of the cultured preparations was measured by MTT. Activities of the lactic dehydrogenase （LDH）, creatine kinase （CK）, and protein content in these cells were also detected. The effects of bFGF with different doses （10 ng/mL, 20 ng/mL, 40 ng/mL, 80 ng/mL, 120 ng/mL and 160 ng/mL） on the above mentioned parameters were also evaluated. Results Cells from both PM and control subjects were successfully cultured in vitro. The cultivation of the muscle cells from PM patients in vitro was not yet seen. Results indicated the obvious stimulation of bFGF on cell proliferation, activities of LDH and CK, protein synthesis, in a dose dependent manner. The optimal dose of bFGF was 120 ng/mL （P〈0.05）, beyond which greater dose caused a less effect. The effect of bFGF on 160 ng/mL was stronger than that on 80 ng/mL, but there was no significant difference （P〉0.05）. Conclusion Myoblastic cells from patients with PM had a weaker ability of developing into the myotubules, thus they were unable to perform effective regeneration, which resulted in a progressive necrosis. The exogenous bFGF could promote the division and proliferation of the muscle cells in vitro. These results shield a light on bFGF＇s potential role in preventing and treating PM.展开更多
基金supported by National Natural Science Foundation of China (41176108)Key Innovation Project of Shanghai Education Commission (14zz145).
文摘Tetrodotoxin(TTX)is a potent neurotoxin firstly discovered in the ovary of pufferfish.The genetic basis of voltage-gated sodium channel resistance to TTX has been widely studied,but it remains unclear in the evolution history of voltage-gated sodium channel resistance to TTX in pufferfish with different TTX concentrations.In this study,six scn4aa coding sequences of pufferfish were firstly cloned and sequenced,then used to investigate the adaptive evolution of scn4aa associated with TTX concentration and reconstruct ancestral sequences with seven scn4aa of other fishes.The result of CODEML(codon substitution model)program from the PAML(phylogenetic analysis by maximum likelihood)package shows only in the genus of Takifugu,which contains TTX highly in the liver,under positive selection.The result also indicates that three of four positively selected sites are located in the intracellular regions,which may compensate for normal function.The ancestral sequence reconstruction may suggest that the replacements providing weak toxin resistance might have appeared first in scn4aa,then the genus Takifugu evolved stronger resistance to TTX later.These results contribute to the explanation of the evolutional history of voltage-gated sodium channel resistance to TTX in pufferfish.
文摘Objective The present study is to observe in vitro the proliferation ability of the muscle cells from permanent myopathy （PM） patients of nomokalaemic periodic paralysis （normKPP）, which is caused by mutations of Metl592Val in the skeletal muscle voltage gated sodium channel （SCN4A） gene on chromosome 17q23.1. We also evaluate the possible effect of the foreign basic fibroblast growth factor （bFGF） in preventing and curing PM. Methods The gastrocnemius muscle cells were taken from two male patients with PM of the same Chinese family with Metl592Val mutation of SCN4A, determined by gene screening. Four male patients suffering from the skeletal injury without PM were taken as control. All preparations were protogenerationally cultured in vitro. Proliferation of the cultured preparations was measured by MTT. Activities of the lactic dehydrogenase （LDH）, creatine kinase （CK）, and protein content in these cells were also detected. The effects of bFGF with different doses （10 ng/mL, 20 ng/mL, 40 ng/mL, 80 ng/mL, 120 ng/mL and 160 ng/mL） on the above mentioned parameters were also evaluated. Results Cells from both PM and control subjects were successfully cultured in vitro. The cultivation of the muscle cells from PM patients in vitro was not yet seen. Results indicated the obvious stimulation of bFGF on cell proliferation, activities of LDH and CK, protein synthesis, in a dose dependent manner. The optimal dose of bFGF was 120 ng/mL （P〈0.05）, beyond which greater dose caused a less effect. The effect of bFGF on 160 ng/mL was stronger than that on 80 ng/mL, but there was no significant difference （P〉0.05）. Conclusion Myoblastic cells from patients with PM had a weaker ability of developing into the myotubules, thus they were unable to perform effective regeneration, which resulted in a progressive necrosis. The exogenous bFGF could promote the division and proliferation of the muscle cells in vitro. These results shield a light on bFGF＇s potential role in preventing and treating PM.
