During development, the Scx cells expressed Gli1 found to be prominent populations, driving the cells toward enthesis [2-3]. Throughout lifetime, the TGFb signaling controls crucial cellular processes of the proliferation and differentiation [7]. The potential novel precision therapy from the regeneration of the enthesis is proposed in this study using endogenous enthesis stem cells (eESC). The eESC is hypothesized to express the pluripotency combined with their renewability that potentially improves the repair process by the regeneration of the supraspinatus enthesis. The first objective of this study aims to trace and characterize the eESC in the supraspinatus enthesis. The cell sub-population in the enthesis is characterized using flow cytometric analysis of the immunofluorescent Scx, Gli1, CD146 and CD90 positive cells. The timed mice are studied here to understand whether there should be the age dependent eESC. To understand the enthesis regeneration, the investigation of the crosstalk between the TGFb and hedgehog pathways is proposed to support the eESC capacity of the …show more content…
eESC-/-TGFbRII and eESC-/-Smo, respectively) to analyze the interaction between TGFb and hedgehog pathways. To test whether the eESC-/-TGFbRII and eESC-/-Smo are knocked out, the phosphorylated Smad2/3 and Gli1/2 gene expression levels are monitored. The activation of the pathway with recombinant TGFb and Ihh (i.e. rh-TGFb, and rh-Ihh) is performed on both eESC-/-TGFbRII and eESC-/-Smo and compared with the wildtype. Further, the ongoing experiments of the single cell analysis and the eESC in vivo model would provide details in regeneration. The single cell analysis is developed to perform the RNA sequencing and gene expression of the eESC from the different timed mice and cell types. The RNA sequencing of different cell types is studied between the Scx+Gli1-CD146- and Scx+Gli1+CD146+. The punched injury of the mouse model is used to observe the eESC during repair and regeneration in vivo. The plasmid luciferase eESC reporter is constructed and transplanted at the injury site. To examine the effective regeneration of the eESC at the supraspinatus enthesis, the live image by two-photon confocal microscope accompanied with the post-mortal histology of the immunofluorescent eESC would be