Journal: Cancers

Article Title: CYP26A1 Links WNT and Retinoic Acid Signaling: A Target to Differentiate ALDH+ Stem Cells in APC -Mutant CRC

doi: 10.3390/cancers16020264

Figure Lengend Snippet: Inducing wt-APC enhances differentiation of neuroendocrine cells: GLP2R+ cells increase and ALDH+ cells decrease. ( A ) GLP2R+ and GLP2R− cells were sorted from parent HT29 cells (mt- APC ) and cells induced to express wt-APC . The percentage of GLP2R+ subpopulation is shown. ( B ) Cells were stained for GLP2R, and relative number was calculated by normalizing to untreated mt- APC cells. The relative number of GLP2R+ cells quadrupled when wt-APC was induced (black bars) and increased by 6-fold with the combination of wt-APC and ATRA (comparing mt- APC black bar and wt-APC gray bar). ( C ) The percentage of ALDH+ SCs cells decreased by ~20% with ATRA alone and up to ~60% when HT29 cells were induced to express wt - APC . ( D ) Results from NanoString profiling show a similar decrease in ALDH1A1 mRNA in response to ATRA and wt-APC . Experiments were performed 3 times and statistical significance was determined by two-way ANOVA with multiple comparisons.

Article Snippet: NEC and ALDH+ (ALDEFLUOR+) cells were quantified and/or isolated using GLP2R antibody conjugated to Alexafluor 594 (Novus Biologicals, Centennial, CO, USA) with IgG control (R&D) or ALDEFLUOR Assay (Stem Cell Technologies, Vancouver, BC, USA) according to the manufacturer’s instructions.

Techniques: Staining

Journal: Cancers

Article Title: CYP26A1 Links WNT and Retinoic Acid Signaling: A Target to Differentiate ALDH+ Stem Cells in APC -Mutant CRC

doi: 10.3390/cancers16020264

Figure Lengend Snippet: Protein expression of neuroendocrine markers is increased with induction of wt-APC . HT29 cells were induced to express wt-APC . Western blotting ( A ) and densitometry ( B ) were performed to analyze the protein expression of neuroendocrine markers. Expression of full-length APC is shown and verified in the top right panel of ( A ). Chromogranin A (CHGA), GLP2R, and NSE/ enolase were increased with wt-APC, but SSTR1 was increased to a lesser extent. * The lower CHGA band was analyzed for densitometry, higher bands may indicate post-translational modification such as glycosylation of the protein. The experiment ( n = 3) was performed with averages plotted in ( B ) and a representative blot is shown in ( A ). Statistical significance was determined by multiple unpaired t -test. The original whole blot of A is included in - Western Blots.

Article Snippet: NEC and ALDH+ (ALDEFLUOR+) cells were quantified and/or isolated using GLP2R antibody conjugated to Alexafluor 594 (Novus Biologicals, Centennial, CO, USA) with IgG control (R&D) or ALDEFLUOR Assay (Stem Cell Technologies, Vancouver, BC, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Western Blot, Modification, Glycoproteomics

Journal: Nature Communications

Article Title: Age-impaired remyelination is associated with dysregulated microglial transitions

doi: 10.1038/s41467-025-64906-w

Figure Lengend Snippet: a Representative scatterplot showing Ly6g + neutrophils, Ly6c + monocytes and Ly6c - Ly6g - microglia from Cd45 + Mrc1 - Cd11b + cells. b Unsupervised clustering of 47,871 microglia and the ( c ) proportional contribution of each cluster isolated from d – f young, 2-month-old and g – i middle-aged, 10-month-old male mice from naïve (n young = 10, n middle-aged = 10), 7 DPI (n young = 8, n middle-aged = 12) and 21 DPI (n young = 10, n middle-aged = 12) conditions, respectively. j UMAP depicting the expression of Idh2—a protein involved in the oxidative phosphorylation pathway. k Proportions of each sample (naïve (n young = 10, n middle-aged = 10), 7 DPI (n young = 8, n middle-aged = 12) and 21 DPI (n young = 10, n middle-aged = 12)) present in neutrophil and monocyte populations. Two-way analysis of variance (ANOVA) with Sidak’s post-hoc test. Each point represents two animals combined, and all data are presented as mean ± SEM. Metabolic pathway activity across ( l ) microglial states identified in (Fig. ), throughout remyelination in ( m ) young mice and ( n ) young combined with middle-aged mice as a function of the genes found in the KEGG metabolic pathways. Source data are provided as a source data file.

Article Snippet: The pellet was resuspended for 40 min in 200 μL of intracellular stain cocktail containing HBSS, 10 μL of BD Brilliant Stain Buffer (BD Biosciences, Cat. No. 566385) and the following antibodies at a 1:100 dilution: Alexa Fluor 647 GAPDH Antibody (#3907, Cell Signaling Technology, Cat. No. 3907S), Alexa Fluor 700 Ki-67 Antibody (Clone 16A8, BioLegend, Cat. No. 652420), Alexa Fluor 488 IRF7 Antibody (Bioss, Cat. No. BS-3196R-A488), APC-eFluor 780 CD68 Antibody, (FA-11, eBioscience, Cat. No. 47-0681-82), DyLight 350 IGF-I/IGF-1 Antibody (Novus Biologicals, Cat. No. NBP2-48001UV), DyLight 594 IDH2 Antibody (Novus Biologicals, Cat. No. NBP2-22166DL594), Brilliant Violet 711TM CD206 Antibody (Clone C068C2, BioLegend, Cat. No. 141727), PE Osteopontin Antibody (R&D Systems, Cat. No. IC808P), PE/Cyanine7 Phospho STAT1 Antibody (Clone A15158B, BioLegend, Cat. No. 686408), PerCP-eFluorTM 710 Ccl3 Antibody (eBioscience, Cat. No. 46-7532-82) and centrifuged (600 × g , 5 min, 4 °C).

Techniques: Isolation, Expressing, Phospho-proteomics, Activity Assay