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Proteintech anti ca1
Anti Ca1, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/anti+ca1/CA1+Antibody/pm41673867-132-13-15
Average 93 stars, based on 14 article reviews
anti ca1 - by Bioz Stars, 2026-09
93/100 stars

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Related Articles

Membrane:

Article Title: Coptis chinensis extracellular vesicles loaded with CA1-siRNA promote endothelial repair and stent restenosis therapy by regulating the PADI2 and NF-κB pathway.
Article Snippet: .. After sealing, the membrane was incubated overnight with primary antibodies at 4 °C: anti-CA1 (13198-2-AP, Proteintech, China), anti-IL-6 (16806-1-AP, Proteintech, China), anti-IL-1β (K002095P, Solarbio, China), antiTNF-α (K002095P, Solarbio, China), anti-alpha SMA (BM0002, Boster, China), anti-MMP9 (BM4089, Boster, China), anti-IκBα (abs131168, absinthe, China), antiIκBα phosphorylation (abs172314, absinthe, China), anti-p65 (BF8005, Affinity, China), anti-p-p65 (bs-0271R, BIOSS, China), and anti-β-actin (66009-1-Ig, Proteintech, China). .. After washing, incubate the membrane with horseradish peroxidase conjugated secondary antibody for 1 h. The signals were detected using a chemiluminescence imaging system (Millipore, USA) and LumiQ universal ECL substrate (SB-WB012, Solarbio, Shanghai), and quantified using ImageJ software, normalized to β-actin levels.

Article Title: Coptis chinensis extracellular vesicles loaded with CA1-siRNA promote endothelial repair and stent restenosis therapy by regulating the PADI2 and NF-κB pathway
Article Snippet: .. After sealing, the membrane was incubated overnight with primary antibodies at 4 °C: anti-CA1 (13198-2-AP, Proteintech, China), anti-IL-6 (16806-1-AP, Proteintech, China), anti-IL-1β (K002095P, Solarbio, China), anti-TNF-α (K002095P, Solarbio, China), anti-alpha SMA (BM0002, Boster, China), anti-MMP9 (BM4089, Boster, China), anti-IκBα (abs131168, absinthe, China), anti- IκBα phosphorylation (abs172314, absinthe, China), anti-p65 (BF8005, Affinity, China), anti-p-p65 (bs-0271R, BIOSS, China), and anti-β-actin (66009-1-Ig, Proteintech, China). .. After washing, incubate the membrane with horseradish peroxidase conjugated secondary antibody for 1 h. The signals were detected using a chemiluminescence imaging system (Millipore, USA) and LumiQ universal ECL substrate (SB-WB012, Solarbio, Shanghai), and quantified using ImageJ software, normalized to β-actin levels.

Incubation:

Article Title: Coptis chinensis extracellular vesicles loaded with CA1-siRNA promote endothelial repair and stent restenosis therapy by regulating the PADI2 and NF-κB pathway.
Article Snippet: .. After sealing, the membrane was incubated overnight with primary antibodies at 4 °C: anti-CA1 (13198-2-AP, Proteintech, China), anti-IL-6 (16806-1-AP, Proteintech, China), anti-IL-1β (K002095P, Solarbio, China), antiTNF-α (K002095P, Solarbio, China), anti-alpha SMA (BM0002, Boster, China), anti-MMP9 (BM4089, Boster, China), anti-IκBα (abs131168, absinthe, China), antiIκBα phosphorylation (abs172314, absinthe, China), anti-p65 (BF8005, Affinity, China), anti-p-p65 (bs-0271R, BIOSS, China), and anti-β-actin (66009-1-Ig, Proteintech, China). .. After washing, incubate the membrane with horseradish peroxidase conjugated secondary antibody for 1 h. The signals were detected using a chemiluminescence imaging system (Millipore, USA) and LumiQ universal ECL substrate (SB-WB012, Solarbio, Shanghai), and quantified using ImageJ software, normalized to β-actin levels.

Article Title: Coptis chinensis extracellular vesicles loaded with CA1-siRNA promote endothelial repair and stent restenosis therapy by regulating the PADI2 and NF-κB pathway
Article Snippet: .. After sealing, the membrane was incubated overnight with primary antibodies at 4 °C: anti-CA1 (13198-2-AP, Proteintech, China), anti-IL-6 (16806-1-AP, Proteintech, China), anti-IL-1β (K002095P, Solarbio, China), anti-TNF-α (K002095P, Solarbio, China), anti-alpha SMA (BM0002, Boster, China), anti-MMP9 (BM4089, Boster, China), anti-IκBα (abs131168, absinthe, China), anti- IκBα phosphorylation (abs172314, absinthe, China), anti-p65 (BF8005, Affinity, China), anti-p-p65 (bs-0271R, BIOSS, China), and anti-β-actin (66009-1-Ig, Proteintech, China). .. After washing, incubate the membrane with horseradish peroxidase conjugated secondary antibody for 1 h. The signals were detected using a chemiluminescence imaging system (Millipore, USA) and LumiQ universal ECL substrate (SB-WB012, Solarbio, Shanghai), and quantified using ImageJ software, normalized to β-actin levels.

Phospho-proteomics:

Article Title: Coptis chinensis extracellular vesicles loaded with CA1-siRNA promote endothelial repair and stent restenosis therapy by regulating the PADI2 and NF-κB pathway.
Article Snippet: .. After sealing, the membrane was incubated overnight with primary antibodies at 4 °C: anti-CA1 (13198-2-AP, Proteintech, China), anti-IL-6 (16806-1-AP, Proteintech, China), anti-IL-1β (K002095P, Solarbio, China), antiTNF-α (K002095P, Solarbio, China), anti-alpha SMA (BM0002, Boster, China), anti-MMP9 (BM4089, Boster, China), anti-IκBα (abs131168, absinthe, China), antiIκBα phosphorylation (abs172314, absinthe, China), anti-p65 (BF8005, Affinity, China), anti-p-p65 (bs-0271R, BIOSS, China), and anti-β-actin (66009-1-Ig, Proteintech, China). .. After washing, incubate the membrane with horseradish peroxidase conjugated secondary antibody for 1 h. The signals were detected using a chemiluminescence imaging system (Millipore, USA) and LumiQ universal ECL substrate (SB-WB012, Solarbio, Shanghai), and quantified using ImageJ software, normalized to β-actin levels.

Article Title: Coptis chinensis extracellular vesicles loaded with CA1-siRNA promote endothelial repair and stent restenosis therapy by regulating the PADI2 and NF-κB pathway
Article Snippet: .. After sealing, the membrane was incubated overnight with primary antibodies at 4 °C: anti-CA1 (13198-2-AP, Proteintech, China), anti-IL-6 (16806-1-AP, Proteintech, China), anti-IL-1β (K002095P, Solarbio, China), anti-TNF-α (K002095P, Solarbio, China), anti-alpha SMA (BM0002, Boster, China), anti-MMP9 (BM4089, Boster, China), anti-IκBα (abs131168, absinthe, China), anti- IκBα phosphorylation (abs172314, absinthe, China), anti-p65 (BF8005, Affinity, China), anti-p-p65 (bs-0271R, BIOSS, China), and anti-β-actin (66009-1-Ig, Proteintech, China). .. After washing, incubate the membrane with horseradish peroxidase conjugated secondary antibody for 1 h. The signals were detected using a chemiluminescence imaging system (Millipore, USA) and LumiQ universal ECL substrate (SB-WB012, Solarbio, Shanghai), and quantified using ImageJ software, normalized to β-actin levels.

other:

Article Title: Quantitative Proteomic Analysis of Mouse Sciatic Nerve Reveals Post-injury Upregulation of ADP-Dependent Glucokinase Promoting Macrophage Phagocytosis
Article Snippet: The following primary antibodies were used in this study: anti-ADPGK (15639-1-AP), anti-RAC2 (10735-1-AP), anti-CA1 (13198-2-AP) and anti-MPO (22225-1-AP) from Proteintech; anti-MBP (sc-271524), anti-F4/80 (C-7) (sc-377009) and anti-c-Myc (9E10) (sc-40) from Santa Cruz; anti-NFH (Abcam, ab4680), anti-Tuj1 (Sigma-Aldrich, T2200) and anti-Actin (Cell Signaling Technology, 3700).

Article Title: M1-Type Macrophages Secrete TNF-α to Stimulate Vascular Calcification by Upregulating CA1 and CA2 Expression in VSMCs
Article Snippet: The proteins were transferred to a PVDF membrane (Millipore, Burlington, MA, USA), which was then blocked with 5% nonfat milk for 2 h. The membrane was incubated overnight at 4°C with primary antibodies against CA1, CA2 (13198-2-AP/16961-1-AP, Proteintech) or GAPDH (AF0502, Elabscience).

Article Title: M1-Type Macrophages Secrete TNF-α to Stimulate Vascular Calcification by Upregulating CA1 and CA2 Expression in VSMCs
Article Snippet: The proteins were transferred to a PVDF membrane (Millipore, Burlington, MA, USA), which was then blocked with 5% nonfat milk for 2 h. The membrane was incubated overnight at 4°C with primary antibodies against CA1, CA2 (13198-2-AP/16961-1-AP, Proteintech) or GAPDH (AF0502, Elabscience).



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Absorptive inter-crypt goblet cells are intricately linked to the microbiota. (A) Schematic representation of the experimental design illustrating the AIMD mix treatment performed in SPF mice. (B) Immunofluorescence staining of proximal colonic sections from control and AIMD mice using <t>anti-CA1</t> and UEA1 lectin for mucus and DAPI for DNA counterstaining. Quantification of CA1 + UEA1 + cells within total CA1 + cells. Scale bar, 50 µm; n = 5 per group. (C) Whole-mount proximal colon from control and AIMD mice stained with the UEA1 lectin for mucus and CellMask for the cell membrane. Representative Z-stacks from the region outlined by the white box are shown at the bottom. UEA1 + cells in inter-crypt region are indicated with arrows. Scale bars, 20 µm. (D) Staining of proximal colonic sections from control and AIMD mice with the UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in the inter-crypt region are indicated with white arrows and the epithelial surface is indicated by the white dashed line. Quantitation of these cells is shown on the right. Scale bar, 50 µm; n = 5 in each group. (E) Volcano plot showing DEGs in the absorptive populations from GF and SPF mice. Some goblet cell markers are highlighted in the figure. The Wilcoxon statistical test with a significance threshold of adjusted p < 0.05 and log2-fold change > 0.25 was used. (F) UMAP plot of the scRNA-seq data showing the Muc2 + population (red dots) within the absorptive cluster in both the GF and SPF mice, grey dots represent canonical absorptive cells (enterocytes and immature enterocytes). The right panel shows the proportion of bona fide absorptive inter-crypt goblet cells. (G) Schematic representation of the experiment with FMT from SPF to GF mice. (H) Immunofluorescence staining of proximal colonic sections from SPF colonized and GF mice with anti-CA1, UEA1 lectin for mucus, and DAPI for DNA counterstaining. UEA1 + cells in the intercrypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Quantification of CA1 + UEA1 + cells within total CA1 + cells. Scale bar, 50 µm; n = 8 per group. (I) Staining of proximal colonic sections from SPF colonized and GF mice with UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by the white dashed line. Quantitation of UEA1 + cells per crypt is shown on the right. Scale bar, 50 µm; n = 8 in each group. (J) Schematic representation of the experimental design illustrating the ABx (antibiotics cocktail) treatment performed in GF mice. (K) Proximal colonic sections from GF control and GF treated with antibiotics-cocktail mice stained with UEA1 lectin for mucus and DAPI for DNA. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by the white dashed line. Quantitation of UEA1 + cells per crypt is shown on the right. Scale bar, 50 µm; n = 4 in each group. All error bars are SEM.
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Establishment of <t>CA1</t> 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.
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Establishment of <t>CA1</t> 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.
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Establishment of <t>CA1</t> 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.
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Enrichment analysis of exclusive and more abundant protein sets. a , b Fold enrichment of functional annotations of gene ontology terms based on molecular function and biological process in PG and NP, respectively. The dot size represents the count of genes associated with each enriched term; a P < 0.05 was considered statistically significant. c UpSet diagram illustrating the identification of hub genes using different algorithms in PG. d Venn networks displaying intersecting hub proteins in PG and NP. Note that five intersected hub proteins were found in PG: <t>CA1,</t> SNCA, HBB, SLC4A1, and ANK1
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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 , <t>CA1</t> , 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.
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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 , <t>CA1</t> , 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.
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Image Search Results


Absorptive inter-crypt goblet cells are intricately linked to the microbiota. (A) Schematic representation of the experimental design illustrating the AIMD mix treatment performed in SPF mice. (B) Immunofluorescence staining of proximal colonic sections from control and AIMD mice using anti-CA1 and UEA1 lectin for mucus and DAPI for DNA counterstaining. Quantification of CA1 + UEA1 + cells within total CA1 + cells. Scale bar, 50 µm; n = 5 per group. (C) Whole-mount proximal colon from control and AIMD mice stained with the UEA1 lectin for mucus and CellMask for the cell membrane. Representative Z-stacks from the region outlined by the white box are shown at the bottom. UEA1 + cells in inter-crypt region are indicated with arrows. Scale bars, 20 µm. (D) Staining of proximal colonic sections from control and AIMD mice with the UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in the inter-crypt region are indicated with white arrows and the epithelial surface is indicated by the white dashed line. Quantitation of these cells is shown on the right. Scale bar, 50 µm; n = 5 in each group. (E) Volcano plot showing DEGs in the absorptive populations from GF and SPF mice. Some goblet cell markers are highlighted in the figure. The Wilcoxon statistical test with a significance threshold of adjusted p < 0.05 and log2-fold change > 0.25 was used. (F) UMAP plot of the scRNA-seq data showing the Muc2 + population (red dots) within the absorptive cluster in both the GF and SPF mice, grey dots represent canonical absorptive cells (enterocytes and immature enterocytes). The right panel shows the proportion of bona fide absorptive inter-crypt goblet cells. (G) Schematic representation of the experiment with FMT from SPF to GF mice. (H) Immunofluorescence staining of proximal colonic sections from SPF colonized and GF mice with anti-CA1, UEA1 lectin for mucus, and DAPI for DNA counterstaining. UEA1 + cells in the intercrypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Quantification of CA1 + UEA1 + cells within total CA1 + cells. Scale bar, 50 µm; n = 8 per group. (I) Staining of proximal colonic sections from SPF colonized and GF mice with UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by the white dashed line. Quantitation of UEA1 + cells per crypt is shown on the right. Scale bar, 50 µm; n = 8 in each group. (J) Schematic representation of the experimental design illustrating the ABx (antibiotics cocktail) treatment performed in GF mice. (K) Proximal colonic sections from GF control and GF treated with antibiotics-cocktail mice stained with UEA1 lectin for mucus and DAPI for DNA. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by the white dashed line. Quantitation of UEA1 + cells per crypt is shown on the right. Scale bar, 50 µm; n = 4 in each group. All error bars are SEM.

Journal: Gut Microbes

Article Title: Microbiota shape the colon epithelium controlling inter-crypt absorptive goblet cells via butyrate–GP R109A signalling

doi: 10.1080/19490976.2025.2573045

Figure Lengend Snippet: Absorptive inter-crypt goblet cells are intricately linked to the microbiota. (A) Schematic representation of the experimental design illustrating the AIMD mix treatment performed in SPF mice. (B) Immunofluorescence staining of proximal colonic sections from control and AIMD mice using anti-CA1 and UEA1 lectin for mucus and DAPI for DNA counterstaining. Quantification of CA1 + UEA1 + cells within total CA1 + cells. Scale bar, 50 µm; n = 5 per group. (C) Whole-mount proximal colon from control and AIMD mice stained with the UEA1 lectin for mucus and CellMask for the cell membrane. Representative Z-stacks from the region outlined by the white box are shown at the bottom. UEA1 + cells in inter-crypt region are indicated with arrows. Scale bars, 20 µm. (D) Staining of proximal colonic sections from control and AIMD mice with the UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in the inter-crypt region are indicated with white arrows and the epithelial surface is indicated by the white dashed line. Quantitation of these cells is shown on the right. Scale bar, 50 µm; n = 5 in each group. (E) Volcano plot showing DEGs in the absorptive populations from GF and SPF mice. Some goblet cell markers are highlighted in the figure. The Wilcoxon statistical test with a significance threshold of adjusted p < 0.05 and log2-fold change > 0.25 was used. (F) UMAP plot of the scRNA-seq data showing the Muc2 + population (red dots) within the absorptive cluster in both the GF and SPF mice, grey dots represent canonical absorptive cells (enterocytes and immature enterocytes). The right panel shows the proportion of bona fide absorptive inter-crypt goblet cells. (G) Schematic representation of the experiment with FMT from SPF to GF mice. (H) Immunofluorescence staining of proximal colonic sections from SPF colonized and GF mice with anti-CA1, UEA1 lectin for mucus, and DAPI for DNA counterstaining. UEA1 + cells in the intercrypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Quantification of CA1 + UEA1 + cells within total CA1 + cells. Scale bar, 50 µm; n = 8 per group. (I) Staining of proximal colonic sections from SPF colonized and GF mice with UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by the white dashed line. Quantitation of UEA1 + cells per crypt is shown on the right. Scale bar, 50 µm; n = 8 in each group. (J) Schematic representation of the experimental design illustrating the ABx (antibiotics cocktail) treatment performed in GF mice. (K) Proximal colonic sections from GF control and GF treated with antibiotics-cocktail mice stained with UEA1 lectin for mucus and DAPI for DNA. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by the white dashed line. Quantitation of UEA1 + cells per crypt is shown on the right. Scale bar, 50 µm; n = 4 in each group. All error bars are SEM.

Article Snippet: After sodium citrate antigen retrieval, primary antibodies against CHGA (ThermoFisher, #MA5-42925), CA1 (Santa Cruz Biotechnology #sc-393490), FABP2 (R&D Systems #AF1486), CLCA1 (Abcam #ab180851), LY6G (BioLegend #108408), MCT1 (Santa Cruz Biotechnology #sc365501), MCT4 (Santa Cruz Biotechnology #sc-376465), OATP (Santa Cruz Biotechnology #sc-271157), or KI67 (Abcam # ab15580) were diluted in antibody diluent (Dako #S0809) at 1:200 dilution for most antibodies (1:2000 for KI67), and the tissue sections were incubated with the antibody dilutions overnight.

Techniques: Immunofluorescence, Staining, Control, Membrane, Quantitation Assay

Absorptive inter-crypt goblet cell numbers are associated with aging (A) Staining of proximal colon sections from aged (~80 weeks old) and young (8–12 weeks old) mice with UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in the inter-crypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Scale bar 50 µm, n = 8/5 in each group. (B) Proportion of Muc2 + cells in the absorptive cluster in aged (>65 y, n = 3) and young-adult (<65 y, n = 4) human colonic samples. Analysis was performed using the Gut Cell Atlas dataset. (C) Immunofluorescence staining of young and aged human descending colonic sections using anti-CA1 and anti-MUC2 antibodies and DAPI for DNA counterstaining. Double-stained cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line (please note that MUC2 and CA1 are expressed in different compartments within a cell). Quantification of CA1 + MUC2 + cells within total CA1 + cells on the right. Scale bar, 30 µm; n = 5/8 per group. (D) Schematic representation of the experimental design for human fecal microbiota transplantation (FMT) in germ-free (GF) mice. The donors were all healthy, female (41 and 72 y, batch 1) and male (35 and 77 y, batch 2). (E) Staining of proximal colon sections from GF mice colonized with human microbiota from aged or young donors with the UEA1 lectin for mucus and DAPI for DNA counterstaining. Two independent experiments were performed using FMT from different donors for each group, as illustrated in D. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Scale bar, 50 µm; n = 5 in each group. (F) Quantification of butyrate levels in the colonic luminal content of GF mice colonized with human microbiota from aged or young donors, as described in D. Statistical significance was determined by unpaired t -test; **** p < 0.0001, *** p < 0.001, ** p < 0.01, * p < 0.05. n = 5 per group. All error bars are SEM.

Journal: Gut Microbes

Article Title: Microbiota shape the colon epithelium controlling inter-crypt absorptive goblet cells via butyrate–GP R109A signalling

doi: 10.1080/19490976.2025.2573045

Figure Lengend Snippet: Absorptive inter-crypt goblet cell numbers are associated with aging (A) Staining of proximal colon sections from aged (~80 weeks old) and young (8–12 weeks old) mice with UEA1 lectin for mucus and DAPI for DNA counterstaining. UEA1 + cells in the inter-crypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Scale bar 50 µm, n = 8/5 in each group. (B) Proportion of Muc2 + cells in the absorptive cluster in aged (>65 y, n = 3) and young-adult (<65 y, n = 4) human colonic samples. Analysis was performed using the Gut Cell Atlas dataset. (C) Immunofluorescence staining of young and aged human descending colonic sections using anti-CA1 and anti-MUC2 antibodies and DAPI for DNA counterstaining. Double-stained cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line (please note that MUC2 and CA1 are expressed in different compartments within a cell). Quantification of CA1 + MUC2 + cells within total CA1 + cells on the right. Scale bar, 30 µm; n = 5/8 per group. (D) Schematic representation of the experimental design for human fecal microbiota transplantation (FMT) in germ-free (GF) mice. The donors were all healthy, female (41 and 72 y, batch 1) and male (35 and 77 y, batch 2). (E) Staining of proximal colon sections from GF mice colonized with human microbiota from aged or young donors with the UEA1 lectin for mucus and DAPI for DNA counterstaining. Two independent experiments were performed using FMT from different donors for each group, as illustrated in D. UEA1 + cells in inter-crypt region are indicated with arrows, and the epithelial surface is indicated by a white dashed line. Scale bar, 50 µm; n = 5 in each group. (F) Quantification of butyrate levels in the colonic luminal content of GF mice colonized with human microbiota from aged or young donors, as described in D. Statistical significance was determined by unpaired t -test; **** p < 0.0001, *** p < 0.001, ** p < 0.01, * p < 0.05. n = 5 per group. All error bars are SEM.

Article Snippet: After sodium citrate antigen retrieval, primary antibodies against CHGA (ThermoFisher, #MA5-42925), CA1 (Santa Cruz Biotechnology #sc-393490), FABP2 (R&D Systems #AF1486), CLCA1 (Abcam #ab180851), LY6G (BioLegend #108408), MCT1 (Santa Cruz Biotechnology #sc365501), MCT4 (Santa Cruz Biotechnology #sc-376465), OATP (Santa Cruz Biotechnology #sc-271157), or KI67 (Abcam # ab15580) were diluted in antibody diluent (Dako #S0809) at 1:200 dilution for most antibodies (1:2000 for KI67), and the tissue sections were incubated with the antibody dilutions overnight.

Techniques: Staining, Immunofluorescence, Transplantation Assay

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.

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

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.

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

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.

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

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.

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

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.

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

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.

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

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.

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

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.

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

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.

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

Enrichment analysis of exclusive and more abundant protein sets. a , b Fold enrichment of functional annotations of gene ontology terms based on molecular function and biological process in PG and NP, respectively. The dot size represents the count of genes associated with each enriched term; a P < 0.05 was considered statistically significant. c UpSet diagram illustrating the identification of hub genes using different algorithms in PG. d Venn networks displaying intersecting hub proteins in PG and NP. Note that five intersected hub proteins were found in PG: CA1, SNCA, HBB, SLC4A1, and ANK1

Journal: International Journal of Oral Science

Article Title: A multi-platform analysis of human gingival crevicular fluid reveals ferroptosis as a relevant regulated cell death mechanism during the clinical progression of periodontitis

doi: 10.1038/s41368-024-00306-y

Figure Lengend Snippet: Enrichment analysis of exclusive and more abundant protein sets. a , b Fold enrichment of functional annotations of gene ontology terms based on molecular function and biological process in PG and NP, respectively. The dot size represents the count of genes associated with each enriched term; a P < 0.05 was considered statistically significant. c UpSet diagram illustrating the identification of hub genes using different algorithms in PG. d Venn networks displaying intersecting hub proteins in PG and NP. Note that five intersected hub proteins were found in PG: CA1, SNCA, HBB, SLC4A1, and ANK1

Article Snippet: Primary antibodies and their respective concentrations were as follows: anti-CA1 # NBP2-76980, 1:1 000 (Novus Biological), anti-CA2 # NBP2-89522, 1:1 000 (Novus Biological), anti-MMP8 # AB56303, 1:1 000 (Abcam Inc), anti-IL-6 # AB6672, 1:500 (Abcam Inc), anti-FTH1 # 858901, 1:1 000 (BIOLEGEND).

Techniques: Functional Assay

Hub proteins in the PG group

Journal: International Journal of Oral Science

Article Title: A multi-platform analysis of human gingival crevicular fluid reveals ferroptosis as a relevant regulated cell death mechanism during the clinical progression of periodontitis

doi: 10.1038/s41368-024-00306-y

Figure Lengend Snippet: Hub proteins in the PG group

Article Snippet: Primary antibodies and their respective concentrations were as follows: anti-CA1 # NBP2-76980, 1:1 000 (Novus Biological), anti-CA2 # NBP2-89522, 1:1 000 (Novus Biological), anti-MMP8 # AB56303, 1:1 000 (Abcam Inc), anti-IL-6 # AB6672, 1:500 (Abcam Inc), anti-FTH1 # 858901, 1:1 000 (BIOLEGEND).

Techniques: Functional Assay, Membrane, Transduction

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.

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