| Objective: 1)To examine the temporal and spatial expression levels of BDNF and its receptor TrkB during Xenopus tropicalis early embryogenesis,metamorphosis and in the adult frog;2)To establish the heart injury and repair model in adult Xenopus tropicalis;3)To study the expression levels of BDNF and its receptor TrkB during the heart repair processes in adult Xenopus tropicalis after apical resection.Methods: 1)Use RT-PCR and Whole-mount in situ hybridization to examine the temporal and spatial expression pattern of BDNF and TrkB during Xenopus tropicalis early embryogenesis;2)Use immunohistochemistry and RT-PCR to check the expression of BDNF and TrkB during tadpole metamorphosis and in the adult Xenopus tropicalis;3)Establish the heart injury and repair model in adult diploid Xenopus tropicalis by apical resection;4)Use Quantitative Real-time PCR to detect the expression levels of BDNF and its receptor TrkB during the heart repair processes in adult Xenopus tropicalis after apical resection.Results: 1)RT-PCR analysis revealed that the expression of BDNF and TrkB started from the oocyte and constitutively expressed with an increasing trend throughout the whole early embryonic development.The relative expression level of TrkB was always higher than BDNF at same stage,and the expression levels of both genes showed a positive correlation;2)WISH analysis revealed that the expression of BDNF was correlated with TrkB.Both genes expressed in the animal pole of the blastomere in the cleavage stage,expressed mainly near the foramenand neural axis in the late gastrula stage,expressed at a high level in the neural plate and the paraxial mesoderm at early neural fold stage,expressed widely in the brain,eyes,ears,branchial arch as well as the spinal cord during the tail bud stage and started to express in the heart at St36;3)Immunohistochemical results showed that,during Xenopus tropicalis metamorphosis,BDNF persistently expressed in the melencephalon,pharynx and heart.4)RT-PCR analysis revealed that,in the adult tissues,BDNF expressed highly in the brain,heart and liver while low expression was found in the kidney and muscle of the hindlimb.However,BDNF expression was not detected in the lungs;TrkB expressed highly in the brain,heart,lung,kidney and muscle of the hindlimb while its expression was not detected in the liver.5)An apical resection model of Xenopus tropicalis was constructed for heart injury and repair.The model was able to replicate heart injury and was successful in repairing a nearly scar-free heart within approximately 30 days.The entire surgical processes could be finished in approximately10 min and the survival rate was over 90%.6)The qPCR results showed that,during the repair processes of the heart in adult Xenopus tropicalis after apical resection,the expressions of BDNF and TrkB in the heart tissues dramatically decreased within a short period of 4 days after surgery.Then the BDNF expression level increased notablely after 30 days of the surgery(around three times the expression level in the normal heart)(p<0.05).However,TrkB expression level always returned to normal levels at 14 days after surgery.Conclusions: 1)BDNF and TrkB,both are among the maternal factors,begin to express in the oocyte.These two genes express throughout the early stages of embryogenesis and show a trend of increasing,and mainly express in the brain,eyes,ears,branchial arch and the notochord.This suggests that BDNF-TrkB pathway may play a role in the process of early embryonic development,and it may mainly function on the development of the nerves and the nerve-related tissues.2)BDNF and TrkB are detected in the heart of the Xenopus tropicalis embyro in St36,in which the heart starts to ring and to beat.Therefore,the BDNF-TrkB pathway may play a role during the development and maturation of the heart.3)During Xenopus tropicalis metamorphosis,the expression level of BDNF in the brain,pharynx and heart have no obvious changes.This suggests that the BDNF-TrkB pathway may play a stable role in the brain and the heart.4)In adult Xenopus tropicalis,the expressions of BDNF andTrkB are more widely distributed in the peripheral tissues such as the liver,lung,kidney and muscle,which indicates that the functions of BDNF-TrkB pathway are important in brain,heart and other organs.5)The apical resection model of Xenopus tropicalis for heart injury and repair could be used to test molecular interventions that stimulate nearly scar-free cardiac repair and will provide a novel way to study adult mammalian heart repair and regeneration.6)BDNF-TrkB pathway may play a role in the process of cardiac repair in the post-surgery adult Xenopus tropicalis. |
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