Regulation Of Transcription Factor Pea3 On Proliferation And Differentiation Of Mesenchymal Stem Cells

Background and purposeRotator cuff tear is a common disease in sports medicine,and the incidence rate is extremely high in the middle-aged and elderly population.Pain and dysfunction caused by rotator cuff injury seriously affect the normal life and work of the patient.Most of the current rotator cuff injuries can be repaired by different surgical methods.However the re-tear rate of rotator cuff still remains high.Mesenchymal stem cells are branches of stem cells that also have the potential for self-replication and multi-directional differentiation.At present,it is found that rational regulate l the proliferation and differentiation of mesenchymal stem cells into tendon can promote the healing of the bone-tendon junction,improve the clinical function and reduce the re-tear rate.However,the biomechanical properties of the rotator cuff after healing have not yet reached normal levels,so further research is needed on the regulation mechanism of mesenchymal stem cells.Pea3 is a member of the E26 transcription factor family and plays an important role in regulating a variety of cell`s proliferation,migration and differentiation.During the development of the chicken body,FGF secreted by the sarcomere can induce the secretion of Scx by the tendon progenitor cells through the activating of Pea3,which can regulate the growth and development of tendon.Pea3 is also a transcription factor downstream of FGF,an important pathway for tendon development,and Pea3 can enhance the function of TGF-? signaling pathway by binding to the Smad protein downstream of TGF-?.Therefore,it is believed that Pea3 may be an important factor in regulating mesenchymal stem cell proliferation and myogenesis.However,the specific role and mechanism of Pea3 in the proliferation and differentiation of mesenchymal stem cells has not been studied in depth.In this project,we overexpressing Pea3 in C3H10T1/2 cells by lentivirus and studied the changes in their proliferation and differentiation ability.Methods and resultsWe first verified the surface markers of C3H10T1/2 cells according to the characteristics of mesenchymal stem cells by flow cytometry,and then overexpressing Pea3 in C3H10T1/2 cells by using lentiviral transfection system,and confirm the transfection effectiveness using q RT-PCR and Western blot.The correct expression of Pea3 in C3H10T1/2 cells was verified.Overexpression of Pea3 was found to increase the proliferation of C3H10T1/2 cells testing by CCK8 and Edu.Then,the expression of the tendon-related factors was analyzed by q RT-PCR.The results showed that overexpression of Pea3 promoted the expression of Col I,Col III,Scx,Tnmd,?-SMA and Vimentin m RNA.Then,the ERK inhibitor SCH772984 was added to the Lv-Pea3 cells to inhibit the activation of Pea3,and Western Blot was used to verify the regulation of Pea3 on the above-mentioned tendon-related factor proteins.It was found that Pea3 up-regulated Col I,Col III and Scx.However,the expression of Pea3 has no obvious effect on the expression of Tnmd protein.Finally,we used FGF2 growth factor to treat C3H10T1/2 cells.QRT-PCR and western blot showed that FGF2 could up-regulate the expression of Pea3 and Scx.ConclusionThrough the above studies,we found that the overexpression of the transcription factor Pea3 in C3H10T1/2 cells can promote its proliferation;up-regulate the expression of the tendon-related factors Col I,Col III,Scx,Tnmd,and thus promote the differentiation of C3H10T1/2 cells into tendon.At the same time,we found that Pea3 may be one of the mechanisms by which FGF regulates the proliferation and differentiation of mesenchymal stem cells.