Journal: Cell Death & Disease

Article Title: Vitamin D opposes multilineage cell differentiation induced by Notch inhibition and BMP4 pathway activation in human colon organoids

doi: 10.1038/s41419-024-06680-z

Figure Lengend Snippet: A Light microscopy images of organoids (patient #110) cultured for 72 h in DIFF, BMP4, DBZ or B + D media in the absence (vehicle) or presence of calcitriol (100 nM). Scale bar, 500 μm. B RT-qPCR analysis of the RNA levels of enterocytic ( KRT20 , CA1 , KLF4 and FABP2 ) and mucosecretory ( AGR2 , TFF2 and TFF3 ) genes in organoids from patients #47, #86, #110, #130 for 48 h and #47, #110, #130, #159 for 72 h cultured in DIFF (red), BMP4 (blue), DBZ (orange) or B + D (green) media in the absence (vehicle) or presence of calcitriol (100 nM). * P < 0.05, ** P < 0.01, *** P < 0.001. C RT-qPCR analysis of the RNA levels of stemness genes in organoids from the same patients and in the same conditions as in B. D Western blot analysis and quantification of the effect of calcitriol on the expression of enterocytic (CA1) and mucosecretory (TFF2) marker proteins in organoids from four patients incubated in PROL or B + D media in the absence (vehicle) or presence of calcitriol (100 nM) for 48 h. The stemness PTK7 protein was used as control of differentiation and GAPDH as loading control. * P < 0.05. E Immunofluorescence analysis of the expression of enterocytic and mucosecretory protein markers in organoids (patient #158) cultured for 48 h in PROL or B + D media in the absence (vehicle) or presence of calcitriol (100 nM). Scale bars, 30 µm. Box-plots show the quantification of fluorescence intensity (Log 2 MGV) (patients #92, #158 and #166; the number of organoids analyzed was 68 for KLF4, 104 for KRT19, 110 for AGR2 and 79 for TFF2). ** P < 0.01, *** P < 0.001. F Western blot analysis and quantification of the level of phospho(P)-SMAD1/5/8 in organoid cultures that were incubated for 24 h with calcitriol (100 nM) or vehicle before incubation in DIFF medium or BMP4 medium (50 ng/mL) during the indicated times. MGV: mean gray value.

Article Snippet: Whole-cell extracts (15–30 μg or 80 μg the case of phospho-SMAD analyses) were separated by SDS-PAGE, transferred to PVDF membranes and incubated with the following primary antibodies: rabbit polyclonal-CA1 (CUSABIO, TX, USA, #CSBPA004364GA01HU), mouse monoclonal-GAPDH (Abcam, Cambridge, UK, #ab8245), rabbit monoclonal-PTK7 (Cell Signalling, MA, USA, #25618), rabbit polyclonal-SMAD1/5/8 (Santa Cruz, TX, USA, #sc6031-R), rabbit monclonal-Phospho-SMAD1/5/8 (Cell Signalling, #9511 S) and rabbit polyclonal-TFF2 (Proteintech, IL, USA, #13681-1-AP).

Techniques: Light Microscopy, Cell Culture, Quantitative RT-PCR, Western Blot, Expressing, Marker, Incubation, Control, Immunofluorescence, Fluorescence

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Establishment of CA1 KI ApoE [−/−] C57BL/6 mice via the CRISPR/Cas9 gene re-editing technique. A. Diagram of the establishment of KI mice with CA1 cDNA inserts. An expression cassette with a CAG promoter and CA1-encoding gene was inserted into intron 1 of the Rosa26 locus in the C57BL/6 mouse chromosome via the CRISPR/Cas9 system. B. Representative PCR images showing the successful establishment of CA1-overexpressing KI mice. a. PCR with primer set 1 yielded one band of 296 bp in KI mice. b. PCR with primer set 2 yielded one band of 335 bp in KI mice. c. PCR with primer set 3 yielded one band of 245 bp in the KI mice. KI: knock-in mice; WT: wild type; M: molecular size marker of DNA.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: CRISPR, Expressing, Knock-In, Marker

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Changes in the body weights of the AS model mice. CA1-overexpressing ApoE [−/−] mice and ordinary ApoE [−/−] mice were induced to AS with high-fat food. These patients were divided into the following groups: normal, AS model, AS model with MTZ treatment and AS model with MTZ preventive-treatment. Each group included 15 mice. Mice with CA1 overexpression generally had greater body weights than did those without CA1 overexpression. The weight of the mice with AS was generally greater than that of the mice without AS, and the weight of the mice with AS was reduced following MTZ treatment and MTZ-preventive treatment.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Over Expression

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Sudan IV staining of mouse cardiac aorta tissues. The number and extent of accumulated lipids in the aorta were semiquantitatively analyzed by calculating lipid accumulation in the whole aorta. The lipid accumulation in the CA1-overexpressing mice was much greater than that in the mice without CA1 overexpression. MTZ treatment significantly decreased the quantity and volume of lipid accumulation. ∗∗ indicates P < 0.01, ∗∗∗ indicates P < 0.001, and ∗∗∗∗ indicates P < 0.0001.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Staining, Over Expression

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: HE staining of mouse cardiac aorta tissues. The extent and composition of the aortic lesions were semiquantified by calculating the surface plaque area across the entire aortic area. The number of aortic plaques and wall thickness in CA1-overexpressing mice were greater than those in mice without CA1 overexpression. ∗ indicates P < 0.05, ∗∗ indicates P < 0.01, ∗∗∗ indicates P < 0.001 and ∗∗∗∗ indicates P < 0.0001.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Staining, Over Expression

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Oil Red O staining of mouse cardiac aorta tissues. The number and size of fat deposits were semiquantitatively analyzed. Fat deposition in cardiac aorta tissues was greater in CA1-overexpressing mice with AS than in mice with AS without CA1 overexpression. MTZ treatment significantly decreased the volume and number of fat deposits. ∗ indicates P < 0.05, ∗∗ indicates P < 0.01, ∗∗∗ indicates P < 0.001 and ∗∗∗∗ indicates P < 0.0001.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Staining, Over Expression

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Biochemical examination of mouse peripheral blood. Compared with those in healthy controls, HDL levels were significantly lower, and AI, LDL, TC and TG levels were elevated in AS mice. The levels of these indices were restored after MTZ treatment. Moreover, the HDL level was significantly lower and the levels of AI, LDL, TC and TG were greater in CA1-overexpressing mice than in ApoE [−/−] mice with normal CA1 expression, regardless of whether these mice were induced to develop AS or treated with MTZ. ∗ indicates P < 0.05, ∗∗ indicates P < 0.01, ∗∗∗ indicates P < 0.001 and ∗∗∗∗ indicates P < 0.0001.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Expressing

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Immunostaining of CA1 expression in mouse aortic tissues. Immunohistochemistry revealed CA1 expression (brown) in the aortic plaques of the animals with AS (↓). CA1 was also weakly expressed in aortic VSMCs (↓↓). Immunofluorescent immunohistochemistry revealed that CA1 levels were increased in the cardiac aorta tissues of CA1-overexpressing ApoE [−/−] mice, regardless of whether the KI mice had induced AS, compared with those of ordinary ApoE [−/−] mice. The quantified signal is normalized to the total cellularized area. ∗∗∗ indicates P < 0.001 and ∗∗∗∗ indicates P < 0.0001.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Immunostaining, Expressing, Immunohistochemistry

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Calcification of mouse cardiac aorta tissues via Von Kossa staining. Extensive calcium deposition was detected in cardiac aorta tissues from CA1-overexpressing AS and AS mice following MTZ treatment. Little calcification was observed in the aortic tissues of ordinary ApoE [−/−] mice with induced AS.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Staining

Journal: Atherosclerosis Plus

Article Title: ApoE [−/−] CA1-overexpressing knock-in mice aggravated atherosclerosis by increasing M1 macrophages

doi: 10.1016/j.athplu.2025.03.003

Figure Lengend Snippet: Immunofluorescence analysis of macrophages in mouse cardiac aorta tissues. The aortic tissues were stained with DAPI (blue). CD86 (green) represents the M1 macrophage subtype, and CD163 (yellow) represents the M2 macrophage subtype. The expression levels of CD86 were semiquantitatively analyzed. Higher CD86 expression was detected in the aortic tissues of CA1-overexpressing mice than in those of ordinary Apoe [−/−] mice. Increased CD86 expression was detected in the aortic tissues of both CA1-overexpressing mice and ordinary ApoE [−/−] mice when they were induced to AS. ∗ indicates P < 0.05, ∗∗ indicates P < 0.01, ∗∗∗ indicates P < 0.001 and ∗∗∗∗ indicates P < 0.0001.

Article Snippet: Rabbit anti-mouse CA1 antibody was obtained from Servicebio (China).

Techniques: Immunofluorescence, Staining, Expressing

Journal: Journal of Cellular Physiology

Article Title: Voltage‐dependent activation of Rac1 by Na v 1.5 channels promotes cell migration

doi: 10.1002/jcp.29290

Figure Lengend Snippet: The large conductance Ca 2+ ‐activated K + channel K Ca 1.1 regulates the membrane potential but not intracellular Na + . (a) MDA‐MB‐231 cells labeled with K Ca 1.1 antibody (green), phalloidin to label the actin cytoskeleton (red), and DAPI to label the nucleus (blue). (b) Western blot of K Ca 1.1 in control MDA‐MB‐231 cells and cells in which Na v 1.5 has been knocked down with shRNA. Positive control = rat brain lysate. Loading control = α‐tubulin. (c) Representative perforated patch clamp recording showing activation of outward current using the K Ca 1.1 activator (NS‐1619; 1 µM) and inhibition with iberiotoxin (100 nM). The cell was held at −120 mV for 250 ms before depolarization to + 60 mV for 300 ms. (d) Current–voltage relationship of the K Ca 1.1 current. Cells were held at −120 mV for 250 ms before depolarization to voltages ranging from −60 to +90 mV in 10 mV steps for 300 ms ( n = 5). Data are fitted with single exponential functions. (e) Dose‐dependent effect of NS‐1619 on the steady‐state V m ( n ≥ 6). Data are fitted to a sigmoidal logistic function. (f) Effect of NS‐1619 (1 µM) on steady‐state V m ( n = 12). (g) Effect of NS‐1619 (1 µM, 5 min) on [Na + ] i ( n = 22). (h) V m recorded using intracellular solution with free [Ca 2+ ] buffered to 5.7 nM versus 100 nM ( n ≥ 10). Data are mean and SEM. ** p < .01; Student's paired t test. DAPI, 4',6‐diamidino‐2‐phenylindole; shRNA, short hairpin RNA

Article Snippet: The following primary antibodies were used: rabbit anti‐Na v 1.5 (1:100; Alomone); rabbit anti‐K Ca 1.1 (1:100; Neuromab); mouse anti‐Rac1‐GTP (1:500; NewEast Biosciences); rabbit anti‐total Rac1 (1:10; NewEast Biosciences); and mouse anti‐CD44 (1:100; AbD Serotec).

Techniques: Labeling, Western Blot, shRNA, Positive Control, Patch Clamp, Activation Assay, Inhibition

Journal: eLife

Article Title: Effects of microcompartmentation on flux distribution and metabolic pools in Chlamydomonas reinhardtii chloroplasts

doi: 10.7554/eLife.37960

Figure Lengend Snippet:

Article Snippet: Antibody , rabbit Anti- Beta-CA1 , AgriSera , Cat# AS11 1737; RRID: AB_10752086 , (1:7500).

Techniques: Software