rnascope probe mm-lgr5  (Advanced Cell Diagnostics Inc)

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Co-expression of LGR5 and epithelial or mesenchymal marker in palatal areas. Transversal sections of mice heads through the nasopharyngeal duct area double stained by anti-EGFP and anti-E-cad as an epithelial marker (A,D,G,J,M) or anti-EGFP and anti-vimentin as a mesenchymal marker (B,E,H,K,N) . There is no overlap of expression of LGR5 (anti-EGFP) and epithelial marker E-cad at E11.5 (A) , E12.5 (D) , E13.5 (G) , E16.5 (J) , E18.5 (M) . Co-expression of LGR5 and mesenchymal marker vimentin is located in the dorsal mesenchyme of forming palatal shelves adjacent to the epithelium at E11.5 (B) , in mesenchyme surrounding future nasopharyngeal duct at E12.5 (E) , E13.5 (H) and in the mesenchyme surrounding the nasal septum at E16.5 (K) and E18.5 (N) . Examples of analyzed cell populations of LGR5-positive cells in the nasopharyngeal duct area and underlaying nasal septum (C,F,I,L,O,Q) . Graph displays quantification of LGR5-positive cells in nasopharyngeal duct area (P) . The highest number of positive cells was observed at E13.5. nc, nasal cavity; nd, nasopharyngeal duct; oc, oral cavity; p, palate; ps, palatal shelve; t, tongue; s, septum. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Expressing, Marker, Staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Distribution of LGR5-positive cells during palatogenesis. Hematoxylin-Eosin staining of transversal head sections at prenatal and postnatal stages with focus on the palatal area (A,C,E,G,I,K,M) . At E11.5, LGR5-positive cells are located adjacent to the epithelium in the dorsal mesenchyme of forming palatal shelves ( B and B′,B´´ in detail ) . Later at E12.5 and E13.5, LGR5-positive signal is present in the dorsal mesenchyme surrounding future nasopharyngeal duct ( D,F and D´,F´ in detail ) and in the mesenchyme of developing palatal shelves (D´´,F´´) . Next, LGR5-positive cells are situated in the mesenchyme surrounding the nasal septum ventrally from nasal cartilage at E14.5 ( H,H´ in detail ) , E16.5 ( J,J´ in detail ) , E18.5 ( L,L´ in detail ) and P4 ( N,N´ in detail) . Several LGR5-positive cells are located in the ventral palatal epithelium or in the mesenchyme of palatal shelves at E14.5 (H´´) . Weak to no expression of LGR5 is found in the epithelium and mesenchyme of palatal shelves at E16.5 (J´´) , E18.5 (L´´) and P4 (N´´) . lg, lingual groove; Mc, Meckel’s cartilage; nd, nasopharyngeal duct; ns, nasal septum; oc, oral cavity; p, palate; ps, palatal shelve; t, tongue; th, tooth. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Staining, Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Distribution of LGR5-positive cells in the vomeronasal organ area. Hematoxylin-Eosin stained sections of vomeronasal organ at several prenatal and one postnatal embryonic stage (A,C,E,G,I,K, M) . Expression of LGR5 is located in the mesenchyme underlying the lateral epithelium of the nasal cavity at E11.5 ( B,B´ in detail ) and E12.5 (D) . LGR5-positive cells are observed in the mesenchyme of developing nasal conchae at E12.5 (D´) , E13.5 (F´) and E14.5 (H´) . Signal of LGR5-positive cells is also situated in the mesenchyme of both sensory and non-sensory epithelium of VNO at E13.5 (F) and E14.5 (H) . At the next stages, LGR5-positive cells are located in the mesenchyme of non-sensory epithelium of VNO at E16.5 (J) , E18.5 (L) or P4 (N) , as well as in the lateral mesenchyme surrounding the nasal cavity, and in the mesenchyme surrounding ventral epithelium of the nasal cavity at E16.5 (J´) , E18.5 (L´) and P4 (N´) . i, incisor; nc, nasal cavity; ns, nasal septum; nse, non-sensory epithelium; oe, oral epithelium; oc, oral cavity; p, palate; ps, palatal shelves; se, sensory epithelium; t, tongue; vno, vomeronasal organ. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Staining, Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Localization of LGR5-positive cells during lingual groove development. Transversal sections of heads through the lingual groove area stained by Hematoxylin-Eosin at several prenatal and one postnatal stage (A,C,E,G,I,K,M) . LGR5-positive cells are situated in the mesenchyme of labial and lingual side of developing groove at E11.5 ( B,B´ in detail ) and E12.5 ( D,D´ in detail ) . Population of LGR5-positive cells is situated in the labial and lingual side of groove mesenchyme and also around epithelium of salivary glands at E13.5 ( F,F´ in detail ) , E14.5 ( H,H´ in detail ) , E16.5 ( J, J´ in detail ) , E18.5 ( L,L´ in detail ) and P4 ( N,N´ in detail ) . lg, lingual groove; Mc, Meckel’s cartilage; oc, oral cavity; ps, palatal shelve; sg, salivary gland; t, tongue; th, tooth. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Fate of LGR5-positive cells during palatogenesis. Timeline of tamoxifen application and collection of samples (A) . Transversal sections of lineage tracing in the developing nasopharyngeal duct and palate of Lgr5-EGFP-CreERT2 (green color) and Rosa26-tdTomato reporter mice (red color) at E18.5 after tamoxifen administration (B) . The expression of LGR5/EGFP positive cells at final time point at E18.5 in the mesenchyme beyond epithelium of nasopharyngeal duct (C, D) . LGR5/tdTomato positive cells at time point from E13.5 to E18.5 (C´,D´) and the overlap of LGR5/EGFP positive cells and LGR5/tdTomato positive cells at E18.5 (C´´,D´´) . Sagittal section of lineage tracing in the caudal region of palate (E, E´,E´´) . Higher-magnification of palatal area with LGR5/tdTomato positive cells in mesenchyme of palate (F) and an overlap of LGR5/EGFP and LGR5/tdTomato cells in the mesenchyme of nasopharyngeal duct (G) . Surprisingly, LGR5/EGFP positive cells are located in the epithelium of rugae palatinae (H´) . Moreover, several LGR/tdTomato cells are situated in the similar area of rugae palatinae, thus there is an overlap of LGR5/EGFP and LGR5/tdTomato cells (H´´) . nc, nasal cavity; nd, nasopharyngeal duct; ns, nasal septum; oc, oral cavity; p, palate; rp, rugae palatinae; t, tongue. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: (A,B) Colocalization analysis of LGR5/EGFP and LGR5/tdTomato positive cells in 3D. Quantitative measurements of colocalization between LGR5/EGFP positive cells (green color) and LGR5/tdTomato positive cells (red color) in three distinct morphological structures of the upper jaw region. Nuclei counterstained with DAPI are in blue. In the upper part of the panel, analyzed structures are graphically dissected in three dimensions of the Z-stack image: 1) anterior palate (AP), 2) area of vomeronasal organ (VNO), 3) roof of nasopharyngeal cavity (RNC). (C,D,E) In the lower part of the panel, there are bar charts displaying measured tdTomato/EGFP overlap in individual structures in Mander’s coefficient and both variations of Pearson’s coefficients. For the sake of figure compactness, analyzed structures are designated in short within the bar charts.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Colocalization analysis of LGR5/EGFP and LGR5/tdTomato positive cells on tissue sections. Quantitative analyses of colocalization between LGR5/EGFP positive cells (green color) and LGR5/tdTomato positive cells (red color) on immunoassayed tissue sections. Nuclei counterstained with DAPI are in blue. In left panel, there are representative images displaying overlayed channels red, green and blue (A,B,C) . Images in right panel (A´,B´,C´) are corresponding to binary masks images from the left panel demonstrating classified pixels above the threshold, which were used for colocalization coefficient calculation. Pixels for LGR5/EGFP channel are in green, LGR/tdTomato channel in red and co-localized pixels are labeled with turquoise color. (D) Boxplots summarizing colocalization analysis from all images of each group: nd, nasopharyngeal duct; vno, vomeronasal organ; lg, lingual groove. Boxplots filled with red color representing colocalization coefficient for red channel, and boxplots filled with green color representing colocalization coefficient for green channel. nd displays the highest level of colocalization for both channels. On the other side, sample lingual groove exhibit the lowest level of colocalization in both channels. vno exhibit the higher level of colocalization in green channel, whereas red channel displays lower level of colocalized pixels. lg, lingual groove; nd, nasopharyngeal duct; nc, nasal cavity; nse, non-sensory epithelium; s, septum; se, sensory epithelium; sg, salivary glands; t, tongue; vno, vomeronasal organ.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Labeling

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Lineage tracing of LGR5-positive cells during development of vomeronasal organ. Timeline of tamoxifen application and collection of embryos (A) . Low-magnification image of VNO after tamoxifen induction and embryos collection at E18.5. LGR5/EGFP positive cells are represented by green color, LGR5/tdTomato positive cells are represented by red color and the overlap of LGR5/EGFP positive cells and LGR5/tdTomato positive cells is represented yellow color (B) . Higher-magnification of lineage tracing in the VNO area (C-D´´) . The LGR5/EGFP positive cells are situated in the mesenchyme underlying the lateral non-sensory epithelium, several cells are localized also around sensory epithelium of VNO (C,C´´) and in the mesenchyme underlying lateral epithelium nasal cavity. They are also spread through the mesenchyme in the ventral part of nasal septum (D,D´´) . Similar distribution like LGR5/EGFP positive cells is observed in LGR5/tdTomato positive cells. These cells are situated in mesenchyme of non-sensory epithelium (C´,C´´) , in the mesenchyme underlying lateral epithelium nasal cavity (D´,D´´) where is their overlap detected (C´´,D´´) . In contrast with LGR5/EFP positive cells, the LGR5/tdTomato cells are not observed in the mesenchyme of sensory epithelium of VNO (C´,C´´) . These observations are confirmed also by sagittal sections of vomeronasal organ in low-magnification pictures (E, E´, E´´ and in detail F,G) . nc, nasal cavity; nd, nasopharyngeal duct; ns, septum; nse, non-sensory epithelium; oc, oral cavity; se, sensory epithelium; vno, vomeronasal organ. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Fate of LGR5-positive cells during development of lingual groove. Timeline of tamoxifen induction and collection of embryos (A) . Low-magnification image of the area of lingual groove after lineage tracing analysis. The overlap of LGR5/EGFP positive cells (green color) and LGR5/tdTomato positive cells (red color) represented by yellow color (B) . Higher-magnification of fate mapping of LGR5-positive cells (C-D´´) . The LGR5/EGFP positive cells are situated in the mesenchyme of labial side of the lingual groove, some of them are localized into the lingual side of groove (C,C´´) or they are spread up to the outlets of salivary glands (D,D´´) . The LGR5/tdTomato positive cells are distributed in the similar way, in the labial and lingual mesenchyme of the groove (C´,C´´) and in the mesenchyme around salivary glands (D´,D´´) . There is observed an overlap of LGR5/EGFP positive cells and LGR5/tdTomato positive cells in above mentioned regions (C´´,D´´) . The sagittal sections of lingual groove confirm distribution pattern of LGR5-positive cell in the area of lingual groove (E,E´, E´´ and in detail F, G) . lg, lingual groove; Mc, Meckel´s cartilage; oc, oral cavity; sg, salivary gland; t, tongue; th, tooth. Scale bars = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Craniofacial abnormalities of Lgr5 −/− mice at E16.5. Lgr5 -deficient mice exhibit several distinct craniofacial defects (A-L) as shown on transversal histological sections of Lgr5 -deficient mice stained by Hematoxylin-Eosin. Low magnification of transversal section of rostral area in Lgr5 +/+ (A) and Lgr5 −/− (B) mice. High magnification of tongue area in WT embryo (A´) and Lgr5 -deficient embryo with distinct ankyloglossia (B´) . No craniofacial defects can be observed in the area of lingual groove at WT embryo (C,E,E´) . Branching of epithelium (D) and multiplication of outlets of salivary glands (F,F´) was observed in Lgr5 -deficient mice. Low-magnification image of palate area of Lgr5 +/+ (G) and Lgr5 −/− (H) mice. High magnification of developing palate with correctly formed nasopharyngeal duct (G´) . Abnormal morphology of nasopharyngeal duct in mutant embryo (H´) . Histological section of middle area of nasopharyngeal duct of Lgr5 +/+ (I) and the atypically formed nasopharyngeal duct of Lgr5 −/− embryos (J) . The caudal area of the nasopharyngeal duct in WT embryo (K) and mutant embryo (L) . (M–N) Sagittal sections through the head display tongue fusion with mouth floor (M´) or dysregulation of salivary gland development (N´) . The expression of E-cadherin is not changed in Lgr5 +/+ (O) and Lgr5 −/− (P) . Expression of β-catenin is decreased in developing nasopharyngeal duct area in Lgr5 −/− in contrast to WT animal (Q, R) . Nuclei are counterstained by DAPI (Blue). e, eye; lg, lingual groove; Mc, Meckel’s cartilage; nd, nasopharyngeal duct; oc, oral cavity; p, palate; sg, salivary gland; t, tongue; vno, vomeronasal organ. Scale bars for A, B. G, H, M, N μ 1 mm. Scale bars for others = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Staining, Mutagenesis, Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Craniofacial abnormalities of Lgr5 −/− mice at E18.5. Lgr5 −/− mice exhibit several craniofacial abnormalities also later in development at E18.5 (A-L) . Low-magnification images of transversal section through rostral area of the head in Lgr5 +/+ (A) and Lgr5 −/− (B) mice stained by Hematoxylin-Eosin. Higher-magnification images of tongue area in WT embryo (A´) without abnormalities and Lgr5 -deficient embryo with ankyloglossia (B´) . While there is not any defect in the area of lingual groove of WT embryo (C,E) , Lgr5 -deficient mice exhibit noticeable epithelial branching and folding in the area of lingual groove (D) . Salivary glands are located in more caudal part in Lgr5 −/− animals (F) in comparison to Lgr5 +/+ embryos (E) . Low magnification (G) and high magnification (G´) image of developing palate. In Lgr5 -deficient mice, there is the cleft palate in low-magnification (H) as well as high-magnification (H´) images. (J) The histological sections of Lgr5 +/+ nasopharyngeal duct (I) and Lgr5 −/− nasopharyngeal duct with distinct midline epithelial bridge. Caudal area of nasopharyngeal duct of WT embryo (K) and mutant embryo with distinct abnormalities such as an epithelial fusion of nasopharyngeal duct (L) . There is downregulation of E-cadherin expression mutant mice (N) in comparison to WT (M) . Also β-catenin expression is decreased in Lgr5 +/+ (O) and Lgr5 −/− (P) mice. i, incisor; lg, lingual groove; m, molar; Mc, Meckel’s cartilage; nc, nasal cavity; nd, nasopharyngeal duct; oc, oral cavity; p, palate; s, septum; sg, salivary gland, t. tongue; vno, vomeronasal organ. Scale bars for A, B. G, H μ 1 mm. Scale bars for others = 100 μm.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Staining, Comparison, Mutagenesis, Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Wnt signaling in Lgr5 -deficient mice. RNA expression of three LGR5 ligands— Rspo1, Rspo2 and Rspo3 and Wnt target gene Axin2 in Lgr5 +/+ and Lgr5 −/− mice at E16.5. The expression of genes is labelled by red color. Erythrocytes, which are naturally autofluorescent, are labelled by green color. RNA expression of Rspo1 is observed in the mesenchyme surrounding nasal septum and nasopharyngeal duct. The significant expression of Rspo1 is situated in palate (A,A´) of Lgr5 +/+ mice. The similar expression of Rspo1 is observed in Lgr5 −/− mice (B,B´) . Rspo2 is expressed also in the mesenchyme surrounding nasal septum in both WT embryo (C,C´) and Lgr5 -deficient mice (D,D´) . The expression of Rspo3 is situated in the mesenchyme around nasopharyngeal duct and surrounding nasal septum in Lgr5 +/+ mice (E,E´) and similar expression pattern of Rspo3 is found in Lgr5 −/− mice (F,F´) , but it seems to be more disperse in the mesenchyme of Lgr5 −/− embryo than Lgr5 +/+ embryo. Axin2 expression is situated in the mesenchyme surrounding nasopharyngeal duct and nasal septum in both WT (G,G´) and mutant embryos (H,H´) . (I-J) qPCR analyses of selected Wnt genes in WT and KO mice embryos and in different craniofacial tissues. (I) The expression of Axin2, Ndk1 and Sfrp2 in the palate. The expression of all genes is higher in Lgr5 +/− palatal cells in comparison to Lgr5 −/− cells in both WT and mutant embryos. (J) In the area of nasopharyngeal duct, the expression of Axin2, Ndk1 and Sfrp2 is also increased in Lgr5 +/− in comparison to cells collected from Lgr5 −/− mice. (K) In the lingual groove, the expression of all genes is increased in Lgr5 +/− cells in comparison to cells collected from Lgr5 −/− animals. nd, nasophraryngeal duct; oc, oral cavity; p, palate; s, septum. Scale bars = 100 μm. The graphs indicate all values and average and they were compared to corresponding controls by t -test (unpaired, 2-tails) * p < 0.05; ** 0.05 < p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: RNA Expression, Expressing, Mutagenesis, Comparison

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Gene expression profile of LGR5-positive and LGR5-negative cells by RNA sequencing. (A) Roof of nasopharyngeal area (rectangle) was collected from animals Lgr5 +/− mice at E16.5 as shown of transversal section through the head stained by Hematoxylin-Eosin. (B) LGR5/EGFP-positive and LGR5/EGFP-negative cells were sorted and analyzed separately. Four samples were used for each cell populations and these samples exhibit high similarity as shown on heat map. 2362 genes exhibited statistically significance between groups. (C) Volcano plot of significantly expressed genes. (D) Differences in expression of Lgr5 in LGR5/EGFP-positive and LGR5/EGFP-negative cells. (E) Heat map of feature genes with focus on epithelial and mesenchymal markers as well as Wnt signaling pathway members. The graphs values indicate average ±SD and p value when data were compared with the corresponding control group.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Gene Expression, RNA Sequencing, Staining, Expressing, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Role of LGR5-positive mesenchymal cells in craniofacial development

doi: 10.3389/fcell.2022.810527

Figure Lengend Snippet: Gene expression of molecules associated with basal membrane. (A–F) Differences in gene expression of Lama1, Lamb1, Lamc3, Flnc and Fblim1 in LGR5/EGFP-positive and LGR5/EGFP-negative cells. Laminin expression in WT (G-H´´) and Lgr5 -deficient animals (I-J´´) detected by immunohistochemistry displays weaker signal of basal membrane marker in mutant animals nd, nasopharyngeal duct; oc, oral cavity; p, palate; s, septum. Scale bars = 100 μm. The graphs values indicate average ±SD and p value when data were compared with the corresponding control group.

Article Snippet: An LGR5 probe (RNAscope ® Probe Mm-LGR5, cat. No. 312171, Advanced Cell Diagnostics, United States) was used to detect transcripts, and DAPI (cat. No. 323 108, Advanced Cell Diagnostics, United States) was used for nucleus staining.

Techniques: Gene Expression, Membrane, Expressing, Immunohistochemistry, Marker, Mutagenesis, Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: BMAL1 Regulates the Daily Timing of Colitis

doi: 10.3389/fcimb.2022.773413

Figure Lengend Snippet: Regenerative pathways are disrupted and decreased in Bmal1 -/- mutant mice. (A) Experiment schematic: Bmal1 +/+ control and Bmal1 -/- mutant mice were collected before DSS treatment (Day 0), after 6 days of 4% DSS treatment to induce colitis (Day 6) and following 10 days of recovery after DSS removal (Day 16). (B) Representative images of sections probed for transcriptional expression of the fetal-like regenerative precursor marker Hopx (in pink), and the ISC marker Lgr5 (in blue, highlighted with blue arrow heads) in untreated (Day 0) Bmal1 +/+ control and Bmal1 -/- mutant mice. Bmal1 -/- mutants show decreased levels of both (C) Hopx and (D) Lgr5 markers in untreated mice. (Unpaired t- test, Hopx *p= 0.0091, Lgr5 *p= 0.0062). (E) Representative images of crypts probed for transcriptional expression of Hopx , and Lgr5 in DSS-colitis treated (Day 6) Bmal1 +/+ control and Bmal1 -/- mutant mice. (F) Upon colitis induction at Day 6, Bmal1 -/- mutant mice exhibit a decrease in Hopx in crypts. (Unpaired t- test, Hopx *p=0.0491). (G) Bmal1 -/- mutant mice exhibit a decrease in Lgr5 in crypts. (Unpaired t- test, Lgr5 *p=0.0301). (H) Representative images of lesions probed for transcriptional expression of Hopx , and Lgr5 in DSS-colitis treated (Day 6) Bmal1 +/+ control and Bmal1 -/- mutant mice. (I) Lesions of Bmal1 +/+ control mice also show decreased Hopx expression. (Unpaired t- test, Hopx *p=0.0489). (J) Lesions of Bmal1 +/+ control and Bmal1 -/- mutant mice show low but similar Lgr5 expression. (Unpaired t- test, p=0.2521). (K) Representative images of crypts probed for transcriptional expression of Hopx , and Lgr5 following 10 days of recovery (Day 16) Bmal1 +/+ control and Bmal1 -/- mutant mice. (L) After 10 days of recovery, Bmal1 -/- mutant mice exhibit similar Hopx expression in crypts. (Unpaired t- test, p=0.66226). (M) Bmal1 -/- mutant mice exhibit a decrease in Lgr5 in crypts. (Unpaired t- test, *p= 0.0468). (N) Representative images of Bmal1 -/- mutant lesion sections probed for transcriptional expression of Hopx , and Lgr5 following 10 days of recovery (Day 16). Sections labeled not/applicable (n/a) indicate that lesions were not present in Bmal1 +/+ control mice. (O) Quantification of Hopx expression in Bmal1 -/- mutant lesions following 10 days recovery. (P) Quantification of Lgr5 expression in Bmal1 -/- mutant lesions following 10 days recovery. (All error bars represent SEM. Scale bar = 50µm).

Article Snippet: The probes used were RNAscope Probe- Mm-Lgr5 ACDBio Cat# 312171 and RNAscope Probe- Mm-Hopx-C2 ACDBio Cat# 405161-C2.

Techniques: Mutagenesis, Expressing, Marker, Labeling