Expression of transporter-associated genes found in corneal endothelium, cultured corneal endothelial cells and corneal stroma.

<p>Transporters expressed in CECs that are at least 2 times higher in young CEC-DM than in stroma, and less than 2 fold difference in expression between young and old CEC-DM. Genes are ranked in descending order of RPM values of young CEC-DM. RPM values are rounded off to the nearest integer, and fold change values are rounded off to the nearest decimal place.</p

Identification of culture conditions for inducing neuroectoderm and mesendoderm differentiation in CDM-BSA.

<p>H9 hESCs were grown for different periods in CDM-BSA supplemented with different growth factors (Activin, FGF2 or BMP4) or chemical inhibitors [SB431542 (SB) for Activin/Nodal, SU5402 (SU) for FGF or BIO for GSK3β/Wnt]. Then, immunostaining analyses were used to determine the fraction of cells expressing the pluripotency marker Oct-4, the neuroectoderm marker Sox2, the mesoderm marker Brachyury, and the endoderm marker Sox17. The levels of marker-expressing cells were divided into five arbitrary categories: 0 for absence of expression, Very Low (VL) for <1% expressing cells, Low (L) for <5% expressing cells, Moderate (M) for >10% expressing cells, High (H) for >50% expressing cells. Indicated in blue are conditions allowing the generation of neuroectoderm (expressing Sox2) and in red those inducing mesendoderm (cells expressing Brachyury or Sox17).</p

Expression of SMADs in corneal endothelium, cultured corneal endothelial cells and corneal stroma.

<p>Genes are ranked in descending order of RPM values of young CEC-DM. RPM values are rounded off to the nearest integer.</p

Genes not expressed in the corneal stroma.

<p>Gene list is filtered for RPM>10 for CEC Young, CEC Old and CEC Culture, and RPM<1 for corneal stroma. Genes are ranked in descending order of RPM values of CEC Young. All values are rounded off to 2 decimal places.</p

The human corneal tissue, the isolated endothelium and the cultured corneal endothelial cells.

<p>A. Corneal stroma with an intact Descemet’s membrane (DM), arrowed (left) and a corneal stromal without the DM layer (right). B. Peeled CEC-DM complex in a typical DM-roll, with the endothelium on the outside. C. Confluent culture of human CECs at the second passage. Scale bars: 100 µm.</p

Pathway analysis of transcription factors via DAVID Functional Annotation.

<p>Transcription factors expressed in young CEC-DM, old CEC-DM and CEC culture (RPM>10) that are more than two-fold higher in young CEC-DM versus stroma, were analyzed in DAVID. Pathways that are over represented are ranked in descending order of percentage of genes representing this category.</p

Identification of culture conditions for inducing neuroectoderm and mesendoderm differentiation in CDM-PVA.

<p>H9 hESCs were grown for different periods in CDM-PVA supplemented with different growth factors (Activin, FGF2 or BMP4) or chemical inhibitors SU5402 (SU) for FGF. Then, immunostaining analyses were used to determine the fraction of cells expressing the pluripotency marker Oct-4, the neuroectoderm marker Sox2, the mesoderm marker Brachyury, and the endoderm marker Sox17. The levels of marker-expressing cells were divided into five arbitrary categories: 0 for absence of expression, Very Low (VL) for <1% expressing cells, Low (L) for <5% expressing cells, Moderate (M) for >10% expressing cells, High (H) for >50% expressing cells. Indicated in blue are conditions allowing the generation of neuroectoderm (expressing Sox2) and in red those inducing mesendoderm (cells expressing Brachyury or Sox17).</p

Gene expression of transcription factors found in corneal endothelium, cultured corneal endothelial cells and corneal stroma.

<p>Transcription factors expressed in CECs that are at least 2 times higher in young CEC-DM than in stroma, and less than 2 fold difference in expression between young and old CECs. Genes are ranked in descending order of RPM values of young CEC-DM. RPM values are rounded off to the nearest integer, and fold change values are rounded off to the nearest decimal place.</p

High-throughput QPCR analysis of genes lowly or not expressed in corneal stroma.

<p>BM: bone marrow; IVS: interventricular septum; SG: salivary gland; SM: skeletal muscle; SI: small intestine; SC: spinal cord; U/C: uterus/cervix; FB: fetal brain; FL: fetal liver; SF: stromal fibroblast; CC: CEC culture; CEC-DM-y: CEC-DM young; CEC-DM-o: CEC-DM-old.</p

Donor information.

<p>A total of 27 donor corneas consisting of 13 single donor corneas and 7 paired donor corneas were used in this study. Donor age ranged from 19–76.</p><p>A: pooled CEC-DM from five young donors;</p><p>B: pooled CEC-DM from five old donors;</p><p>C: Isolated for cell culture;</p><p>D: young CEC-DME control;</p><p>E: old CEC-DME control.</p><p>OS: oculus sinister.</p><p>OD: oculus dexter.</p