15 research outputs found
Differentiation of KSP-positive cells into each segment of renal tubular cells.
<p>(a) PCR showing the expression of segment-specific genes of renal tubular cells. The samples were KSP-positive cells examined right after cell purification with flow cytometry, KSP-positive cells forming tubular structures co-cultured with NIH3T3-Wnt4, and NIH3T3-Wnt4 alone. The bands were quantified using ImageJ, and graphs normalized against GAPDH were shown. After sorting: a sample of cells collected right after cell sorting of KSP-positive cells. Tubular structures: a sample of KSP-positive cells that form tubular structures and feeder cells of NIH3T3-Wnt4. Feeder alone: a sample of feeder cells, NIH3T3-Wnt4. (b) Immunofluorescence showing Megalin, AQP2 and Podocalyxin in tubular structures formed by KSP-positive cells co-cultured with NIH3T3-Wnt4. A micron scale is shown at the lower right corner.</p
Wnt4 enhanced the tubular formation of KSP-positive cells.
<p>(a) Fluorescence microscopy of tubular structures formed by KSP-positive cells derived from CAG-GFP EB3. The upper lane shows pictures of tubular structures of KSP-positive cells co-cultured with NIH3T3-Wnt4, and the lower lane shows those co-cultured with NIH3T3-Mock. Scale bar, 200 µm. (b) Bar graph indicating the numbers of tubular structures under the indicated conditions. One tubular structure from a branching to a branching is counted as one. (c) Immunofluorescence of KSP-positive or KSP-negative cells showing KSP and human-specific mitochondria as a negative control. Scale bar, 400 µm. (d) PCR showing the expression of Osr1, KSP and GAPDH of KSP-positive and -negative cells after three days of 3D culture in Matrigel with NIH3T3-Wnt4.</p
KSP-positive cells formed tubular structures in Matrigel.
<p>(a) Light microscopy of KSP-positive and KSP-negative cells under the conditions indicated in the upper left corner of the pictures on the day after flow cytometry. A micron scale is shown at the lower right corner of the pictures. (b) Electron microscopy of a cross section of the tubular structures formed by KSP-positive cells. The asterisks indicate the lumens, the arrowheads point to microvilli-like structures, and an arrow indicates a tight junction. Magnification is shown at the lower right corner.</p
KSP-positive cells derived from mouse ES cells showed the characteristics of developing kidneys.
<p>(a) Flow cytometry for KSP in CAG-GFP EB3 on day 18 of the differentiation. Cells were gated using side scatter and forward scatter, and dead cells were excluded using propidium iodide. Negative control: Cells incubated with no antibody. 2nd antibody: Cells incubated with only streptavidin-Alexa Fluor 647. 1st/2nd antibody: Cells incubated with anti-KSP antibody and streptavidin-Alexa Fluor 647. (b) Heat map of kidney development (GO:0001822) for KSP-positive and KSP-negative cells derived from CAG-GFP EB3. The microarray data was deposited in the Gene Expression Omnibus (GSE40694, <a href="http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE40694" target="_blank">http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE40694</a>).</p
HGF and IGF-1 enhanced the expression of KSP combined with Activin.
<p>(a) Induction method for evaluating the effects of HGF and IGF-1 on mouse ES cells. (b) Expression of KSP in conditions with HGF or IGF-1 determined using real-time PCR at day 18. The values were normalized against GAPDH. The asterisk indicates <i>P</i><0.05 versus Activin (10 ng/mL) alone. (c) Flow cytometry for KSP in CAG-GFP EB3 at day 18 of the differentiation with Activin and IGF-1.</p
KSP-positive cells were incorporated with renal tubular cells of mouse embryonic kidneys.
<p>KSP-positive and KSP-negative cells were injected into mouse embryonic kidneys. An immunofluorescent analysis was performed on day 3 after the injection. A micron scale is shown at the lower right corner. (a) KSP-positive cells. The arrows indicate the ES cells that incorporated with the renal tubular cells, and a high magnification picture is also shown. (b) KSP-negative cells.</p
Smooth muscle specific antibodies used for analysis
<p>Smooth muscle specific antibodies used for analysis</p
Fluorescence-conjugated monoclonal antibodies used for FACS analysis
<p>Fluorescence-conjugated monoclonal antibodies used for FACS analysis</p
Characterization of peripheral blood and umbilical cord-derived EPC (pEPC and uEPC, respectively) by flow cytometer.
<p>a) Representative forward and side scatter profile of cultured pEPC. b-d) Flow cytometric analysis of ulex-lectin binding/acLDL uptake (b, c) and various cell surface markers (d) in pEPC. e) Flow cytometric analysis of cell surface markers in uEPC.</p
Characterization of transplanted human VPC-derived vascular cells.
<p>a) Flow cytometric analysis of cell surface markers on expanded human VPC-derived VEGF-R2<sup>+</sup>VE-cadherin<sup>+</sup> cells ( = EC). b) Immunofluorescence image of CD31 (green) and αSMA (red) with nuclear staining (blue) in expanded EC. Scale bar: 100 µm. c) Immunostaining of mural cell markers (brown) with hematoxyline counter-staining of expanded VPC-derived VEGF-R2<sup>+</sup>VE<sup>−</sup>cadherin- cells ( = MC). Scale bar: 100 µm. d, e) RT-PCR analysis of mural cell (d) and skeletal/cardiac specific (e) markers in human VPC-derived vascular cells.</p