Journal: bioRxiv

Article Title: Cell-matrix adhesion controls Golgi organization and function by regulating Arf1 activation in anchorage dependent cells

doi: 10.1101/261842

Figure Lengend Snippet: (a) Serum-starved WT-MEFs detached with Accutase (5’ SUSP) and held in suspension for 120 mins (120’ SUSP), labeled with Alexa-488 conjugated ConA, WGA, PNA and FITC-UEA lectin. Cell surface-bound lectin fluorescence was measured by FACS and median fluorescence intensity of 120’ SUSP cells (black bar) was normalized to their respective 5’ SUSP cells (equated to 100 and represented as a grey bar). The graph represents mean ± SE from 6 independent experiments. WT-MEFs expressing cherry-N1 (CNT), wild-type Arf1-Cherry-N1 (WT-Arf1) or active Arf1-Cherry-N1 (Q71L Arf1) were similarly labeled with ( b ) ConA-Alexa488 and ( c ) UEA-FITC. Surface-bound lectin fluorescence in cell population gated for cherry tagged Arf1 expression was measured by FACS and median fluorescence intensity at 120’ SUSP (black bar) normalized to their respective 5’ SUSP cells (equated to 100 and represented as a grey bar). The graph represents mean ± SE of 6 independent experiments. ( d ) Schematic represents the integrin-dependent regulation of Arf1 through BIG1/2, that allows for the recruitment of dynein which along the microtubule network regulates Golgi architecture and function.

Article Snippet: Lectin probes were purchased Invitrogen Molecular Probes, Concanavalin A-Alexa 488 (Cat. No. # C11252), PNA-Alexa-488 (Cat. No. # L21409), WGA-Alexa-488 (Cat. No. # W11261).

Techniques: Labeling, Fluorescence, Expressing

Journal: bioRxiv

Article Title: Nicotinamide riboside activates renal metabolism and protects the kidney in a model of Alport syndrome

doi: 10.1101/2024.02.26.580911

Figure Lengend Snippet: ( A ) Representative images of immunohistochemistry for p57 kip2 , followed by periodic acid–Schiff post-staining without hematoxylin counterstaining. Podocyte nuclei are stained brown, and all other glomerular structures are stained pink. ( B ) Quantification of p57 kip2 immunostaining with PodoCount, a validated algorithm to analyze p57 kip2 -stained whole slide images. Podocyte volumetric density was reduced in Alport mice and restored by NR treatment in both sexes. ( C,D ) Immunoblots for KIM-1, a tubular injury marker, in male ( C ) and female ( D ) kidney homogenate. Ponceau S, a nonspecific protein stain, was used as a loading control. ( E,F ) Quantification of immunoblots ( C,D ) shows that KIM-1 is increased in Alport mice and reduced by NR treatment in male ( E ) and female ( F ) mice. Scale bars represent 100 µm. Significance was determined by one-way ANOVA with the Holm–Šídák correction for multiple comparisons. Data are expressed as the means ± SEM. Each datum represents one glomeruli ( B ) or one mouse ( E-F ). * P < 0.05, ** P < 0.01, **** P < 0.0001. Kidney injury molecule-1, KIM-1; NR, nicotinamide riboside; PSR, picrosirius red; Veh, vehicle.

Article Snippet: Immunohistochemistry for p57 kip2 (Cat. No. ab75975, Abcam) followed by periodic acid–Schiff (Cat. No. 22-110-645, Fisher Scientific, Hampton, NH) post-staining without hematoxylin counterstaining was performed and analyzed as previously described ( ).

Techniques: Immunohistochemistry, Staining, Immunostaining, Western Blot, Marker