All data were normalized to the expression of the gene whose expression does not change under the experimental conditions

All data were normalized to the expression of the gene whose expression does not change under the experimental conditions. hESCs positive for the transcription element Nanog, Oct4, Sox2, SSEA-4, and TRA-1-60. Insets symbolize DAPI staining.(7.33 MB BMS-986020 sodium TIF) pone.0010246.s003.tif (6.9M) GUID:?C0489A32-FBF5-43CE-A70F-26E48A89CCB4 Number S2: Karyograms of hESC lines. A G-banding karyograms showing normal karyotypes of hESC lines, Regea 06/015 at passage 35, Regea 07/046 at passage 36, Regea 08/013 at passage 25 and Regea 06/040 at passage 71.(9.41 MB TIF) pone.0010246.s004.tif (8.9M) GUID:?153D07E6-AC89-4FAD-A352-5357FE998411 Number S3: Histology of teratomas from cell line Regea 06/015 at passage 35. A) An overview of a teratoma. B) Endoderm: fine detail from the right part of the overview showing a lumen defined by high cylindrical cells (*). The cells are compatible with endodermal differentiation of intestinal or respiratory type. C) Ectoderm: a cluster of neurons assembled into a ganglion like structure (arrow). Materials tracts emanates from the cluster. D) Ectoderm: a squadmous epithelial island composed of fairly vacuolated cells (arrow). E) Mesoderm: lower part an island composed of hypertrophic chondrocytes (arrow).(7.14 MB TIF) pone.0010246.s005.tif (6.8M) GUID:?838AC514-F130-47B2-AA7C-3B1B99128F11 Number S4: Histology of teratomas from cell line Regea 07/046 at passage 17. A) This image shows tubular constructions lined by cuboidal to cylindrical epithelium and aggregates consisting of smooth muscle mass cells (sm) are seen. B) Endoderm: some PASD+ cells (arrow) can be seen in the image, which are compatible with Goblet cells indicating the endodermal differentiation. C) Endoderm and mesoderm: a single tubule (*) lined by cuboidal to cylindrical epithelium. Note that it is inlayed inside a loosely arranged connective cells – primitive mesenchyme. An interpretation would be that this represents endodermal (epithelial tubules) and mesodermal parts. D) Endoderm or ectoderm: this image shows an epithelium lined tubule (*) inlayed within a very loosely arranged mesenchyme. The epithelium can be interpreted as bilayered potentially representing a squamous variant. It is impossible to state if it is endodermal BMS-986020 sodium or ectodermal in source.(8.57 MB TIF) pone.0010246.s006.tif (8.1M) GUID:?D5004A52-6665-44A2-98C6-1EF472CBD72F Number S5: Histology of teratomas from cell line Regea 08/013 at passage 86. A) Ectoderm: Large part of NCAM-positive, neuronal cells (arrow). B) Mesoderm: Bundles of muscle mass cells are stained positive for Desmin (arrow). C) Endoderm: Pseudostratified ciliated columnar epithelium stained positive for HNF3 (arrow).(6.05 MB TIF) pone.0010246.s007.tif (5.7M) GUID:?2D02F43C-E6FE-482E-843D-3126B084A72E Number S6: Histology of teratomas from cell line Regea 06/040 at passage 34. A) and B) An overview of teratomas. Endoderm and mesoderm: notice cartilage (c) and tubular structure (*) defined by cylindrical cells suggesting mesodermal differentiation and varying amounts of Goblet cells suggesting Rabbit Polyclonal to ALPK1 endodermal differentiation. C) Endoderm: a high power look at of an area from B is definitely shown. Note clean muscle mass like cells (sm) and cylindrical cells with spread mucous generating Goblet cells. D) Ectoderm: a strong nuclear manifestation of MITF (arrow) specific for retinal pigment epithelial cells is seen indicating ectodermal differentiation.(5.84 MB TIF) pone.0010246.s008.tif (5.5M) GUID:?9A03BD8B-96F2-4B13-AA55-F6508D889EAF Abstract Background The growth of stem cells in conditions requires ideal balance between signs mediating cell survival, proliferation, and self-renewal. For medical software of stem cells, the use of completely defined conditions and removal of all animal-derived materials from your establishment, tradition, and differentiation processes is desirable. Strategy/Principal Findings Here, we report the development of a fully defined xeno-free medium (RegES), capable of assisting the development of human being embryonic stem cells (hESC), induced pluripotent stem cells (iPSC) and adipose stem cells (ASC). We describe the use of the xeno-free medium in the derivation and long-term ( 80 passages) tradition of three pluripotent karyotypically normal hESC lines: Regea 06/015, Regea 07/046, and Regea 08/013. Cardiomyocytes and neural cells differentiated from these cells show features characteristic to these cell types. The same formulation of the xeno-free medium is capable of assisting the undifferentiated growth of iPSCs on human being feeder cells. The characteristics of the pluripotent hESC and iPSC lines are comparable to lines derived and cultured in standard undefined culture conditions. In the tradition of ASCs, the xeno-free medium provided significantly higher proliferation rates than ASCs cultured in medium containing allogeneic human being serum (HS), while keeping the differentiation potential and characteristic surface marker manifestation profile of ASCs, although significant variations in the surface marker manifestation of ASCs cultured in HS and RegES press were exposed. Summary/Significance Our results demonstrate that human being ESCs, iPSCs and ASCs can BMS-986020 sodium be managed in the same defined xeno-free medium formulation for a prolonged period of time while keeping their characteristics, demonstrating the applicability of the simplified xeno-free medium formulation for the production of clinical-grade stem cells. The basic xeno-free formulation explained herein has the potential to be further optimized for specific applications relating to establishment, development and differentiation of various stem cell.