Isolation And Characterization Of Stem Cells In The Rat Penile Corpus Cavernosum

Objective To explore whether endogenous stem cells and muscle-derived stem cells are present in the rat penile corpus cavernosum by isolating and culturing corpus cavernosum cells and detecting the expression of stem cell markers stem cell antigen-1(Sca-1),Oct4 and myoblast marker Desmin.Methods The penile corpus cavernosum were taken under sterile condition from 2-month-old Sprague-Dawley rats which were weight 200～250 gram,then digested and dissociated by typeⅠcollagenase and trypsin.Cells were collected by filtering and centrifugating and cultured in DMEM,with 20℅ fetal bovine serum.Stem cells were isolated from the primary culture using the modified Preplate technique. PP1-PP6 adhered cells were obtained and detected the expression of stem cell markers Sca-1,Oct4 and myoblast marker Desmin by flow cytometry,immunofluorescence and western blot.Moreover,the double expression of Sca-1/Oct4,Sca-1/Desmin,Oct4/ Desmin were detected by immunofluorescence.Results PP1 and PP2 cells adhered on the flask quickly.They were mostly long spindle-shaped fibroblast.The adherence of PP3-PP5 cells decreased gradually.It appeared short spindle-shaped and polygon cells.PP6 cells were smaller and round floating on the culture medium.Adhered cells could be seen and shaped round or spindly from day two to three.PP3 cells were showed Sca-1(+)0.3℅±0.2℅, Oct4(+)0.2℅±0.3℅ by flow cytometry,which had no significantly difference between the control (p﹥0.05),and Desmin(+)15.3℅±1.5℅,which had significantly difference between the control (p﹤0.05).PP6 cells were showed Sca-1(+)5.7℅±0.7℅, Oct4(+)2.6℅±0.3℅,Desmin (+)41.1℅±1.6℅,which had significantly difference between PP3 (p﹤0.05).Positive immunofluorescence outcomes were found with the stem cell markers and myoblast markers.Sca-1 or Oct4 scattered in microscope field of vision. Positive staining of Sca-1 mainly distributed in the cytoplasm,whereas Oct4 mainly distributed in the cell nucleus.The cells expressed Desmin were aggregation, the positive staining of Desmin distributed in the cytoplasm.It also showed that very few cells were double-positive staining Sca-1/Oct4,Sca-1/Desmin and Oct4/Desmin. Sca-1/Oct4 stained in the cytoplasm and cell nucleus respectively,Sca-1/Desmin in the cytoplasm simultaneously,Oct4/Desmin in the cell nucleus and cytoplasm respectively.Western blot showed that the clear presentation of Sca-1 and Oct4 protein were not detected in PP1-PP5 cells, but the presentation of both protein were detected in pp6 cells. Desmin protein was detected in pp1-pp6 cells and increased gradually.Conclusion The cells from rat penile corpus cavernosum had the cell phenotype of stem cells and muscle-derived stem cells,which showed that endogenous stem cells and muscle-derived stem cells were present in the penile corpus cavernosum.