Journal: International Journal of Stem Cells

Article Title: Notch Is Not Involved in Physioxia-Mediated Stem Cell Maintenance in Midbrain Neural Stem Cells

doi: 10.15283/ijsc22168

Figure Lengend Snippet: Effects of physiological hypoxia (physioxia) on proliferation, spontaneous differentiation and cell cycle phase distribution of fetal mNSCs. (A) Representative bright-field images of E13.5 fetal mNSCs cultured for 6 or 13 days in 3% or 21% O 2 . Scale bar is 25 μm. (B) Representative images of mNSCs stained for Tuj1, Map2, GFAP, Nestin and Hoechst. A total of 10 5 cells were seeded on a coverslip and grown for three days under physioxic or normoxic conditions. Scale bar is 50 μm. (C) Comparison of a marker selection of midbrain NSC development (from left to right side) showed additional reduction of late neuronal markers such as Lmo3 or Sox6 . (D) Quantitative analysis of immunoreactivity of mNSCs grown in 3% or 21% O 2 for Tuj1, Map2, GFAP and Nestin normalised to the total number of cells (Hoechst + ) in percent. (E) Quantitative analysis of the fraction of mNSCs in the different cell cycle phases of mNSCs grown in 3 and 21% O 2 . Plots show the relative distribution of cells grown under 3% or 21% O 2 across the different cell cycle phases G0-G1, S or G2-M in percent. (F, G) Comparison of cell cycle markers (F) and senescence/quiescence markers (G) of mNSCs by microarray analysis. Heat-maps represent the fold change of the relative mRNA expression levels of mNSCs cultured under physioxia for 48 h or 13 d in comparison to those cultured under normoxia. Colour bar displays the colour contrast level of the heat-map with red and green indicating high and low expression levels for (C, F, G). *p<0.05, **p<0.01, ***p<0.001 in respect to 21% O 2 (unpaired two-sided t-test with Bonferroni correction).

Article Snippet: Microarray chips were then immediately scanned using an Agilent microarray confocal laser scanner.

Techniques: Cell Culture, Staining, Comparison, Marker, Selection, Microarray, Expressing

Journal: International Journal of Stem Cells

Article Title: Notch Is Not Involved in Physioxia-Mediated Stem Cell Maintenance in Midbrain Neural Stem Cells

doi: 10.15283/ijsc22168

Figure Lengend Snippet: The Notch pathway is active in mNSCs. (A) Representative images of mNSCs stained for NICD and Hoechst. A total of 10 5 cells were seeded on a coverslip and cultured for three days under hypoxic or normoxic conditions. Scale bar is 50 μm. (B) Comparison of mRNA expression of mNSCs by microarray analysis. Heat-map represents the log of the fold change of the relative mRNA expression levels of mNSCs cultured under physioxia for 48 h or 13 d in comparison to those cultured under normoxia. Colour bar displays the colour contrast level of the heat-map with red and green indicating high and low expression levels. (C) Relative mRNA levels of Hes1, 3, 5, Id1, Hey1 and Hif-1α target genes Vegf, and Pgk1 in respect to Hmbs levels as housekeeping gene of mNSCs grown in 3% or 21% O 2 . (D) Relative mRNA levels of Hes1, 3, 5, Hey1, and Notch 1 and Hif-1α target genes Vegf, and Pgk1 in respect to Hmbs, normalised to the respective control condition (Hif lox/lox ) of mNSCs Hif-1α CKO grown in 3% or 21% O 2 . Red dotted line indicates threshold for relevant changed genes. *p<0.05, **p<0.01 and ***p<0.001 in respect to normoxia (B, C) or control condition Hif lox/lox (D; unpaired two-sided t-test with Bonferroni correction).

Article Snippet: Microarray chips were then immediately scanned using an Agilent microarray confocal laser scanner.

Techniques: Staining, Cell Culture, Comparison, Expressing, Microarray

Journal: BMC Bioinformatics

Article Title: Oliz, a suite of Perl scripts that assist in the design of microarrays using 50mer oligonucleotides from the 3' untranslated region

doi: 10.1186/1471-2105-3-27

Figure Lengend Snippet: Verification of specificity . Microarrays were printed with 1816 oligonucleotides in duplicate. Each sub-grid has 13 columns and 8 rows. (A) SYTO59 DNA staining demonstrating the layout of one sub-grid. The spot in the upper right corner and the spot four columns to the left were printed with Cy3-labeled oligonucleotides for orientation purposes. Some positions near the top of each sub-grid were printed with buffer only, and are thus not visible on SYTO59 staining. (B)-(F): cDNAs contain the complementary strand of one of the 50mers on the array were amplified by PCR and labeled with Cy3. Hybridization of these cDNAs to the microarray shows that only the complementary spot had fluorescent signal in most cases. cDNA for Rn.2401 also hybridized to another spot in the same sub-grid, but with lower intensity.

Article Snippet: The arrays were spun dry and were scanned using a confocal microarray scanner (BioRad, Hercules, CA).

Techniques: Staining, Labeling, Amplification, Hybridization, Microarray

Journal: iScience

Article Title: Charge Characteristics of Agouti-Related Protein Implicate Potent Involvement of Heparan Sulfate Proteoglycans in Metabolic Function

doi: 10.1016/j.isci.2019.10.061

Figure Lengend Snippet: Glycan Array Analysis of AgRP Binding to Heparan Sulfate Oligosaccharides (A) Fluorescent image of the glass slide glycan arrays showing fluorescence signals (green dots) of AgRP binding to 52 immobilized heparan sulfate oligosaccharides (low-molecular-weight heparan sulfate, LMHS). (B) Bar graph showing the relative fluorescence intensity of AgRP binding to the heparan sulfate oligosaccharides arrays. Heparan sulfate oligosaccharides 22 and 41 show the highest intensity. (C) The structures of the different heparan sulfate oligosaccharides on the slide microarray in A. Data are represented as mean ± SEM.

Article Snippet: The washed arrays were dried by centrifugation and immediately scanned for green fluorescence using an InnoScan confocal microarray scanner (Innopsys, Carbonne, France).

Techniques: Binding Assay, Fluorescence, Molecular Weight, Microarray