Journal: International journal of molecular sciences

Article Title: The Role of Type VI Collagen in Alveolar Bone.

doi: 10.3390/ijms232214347

Figure Lengend Snippet: Figure 1. The expression of Type VI collagen in oral tissues. (A) H&E staining of mandibles from 3-month-old mice. Dashed line shows periodontal ligament. (B) Representative immunofluorescent staining image of Type VI collagen in mouse mandibles of 3-month-old mice (n = 3). Magnified images of boxed areas in panel B are shown in (a’,b’,c’,d’). Nuclei were stained with DAPI (blue). Type VI collagen expression (red) in the periodontal ligament (PDL) (yellow arrowheads), bone marrow (BM) (green arrowheads), gingiva (Gn (blue arrowheads). (C) The abundance of alpha chains of Type VI collagen in murine alveolar bone from 6-week-old mice. M: molar, AB: alveolar bone, DP: dental pulp, Gn: gingiva, BM: bone marrow, PDL: periodontal ligament, Ep: epithelium, CT: connective tissue. Scale bars: (A,B): 500 µm, (a’–d’): 50 µm.

Article Snippet: Primary antibody specific to Type VI collagen (Fitzgerald, North Acton, MA, USA) was applied to samples at a 1:50 dilution.

Techniques: Expressing, Staining

Journal: Cell reports

Article Title: PTK7 is a positive allosteric modulator of GPR133 signaling in glioblastoma

doi: 10.1016/j.celrep.2023.112679

Figure Lengend Snippet: (A) Confocal microscopic image from “sandwich cultures” shows layered cells expressing either GPR133 (green) or PTK7 (red). Top view from a central confocal slice (center panel: x and y dimension) and orthogonal views (upper panel: z and x; right panel: z and y) depicted on the same scale. Scale bars, 10 μM. (B) HTRF assays in “sandwich cultures” demonstrate that PTK7 in neighboring cells increased cAMP levels in cells expressing wild-type (WT) cleaved GPR133, but not the uncleavable H543R mutant. A secreted PTK7 with no transmembrane domain (no TM, aa 1–703) did not influence GPR133 signaling. Two-way ANOVA: middle layer F 2, 96 = 4.62, not significant; outer layer F 2, 96 = 3.93, not significant; interaction of inner and outer layer F 4, 96 = 7.29, p < 0.0001. Tukey’s multiple comparisons: GPR133-WT-expressing cells with empty vector vs. PTK7 co-culture, p < 0.0001; GPR133-WT-expressing cells with full-length PTK7 vs. secreted PTK7 (no TM) co-culture, p < 0.0001. All other comparisons not significant; n = 4–22 independent experiments. (C) Pre-coating wells with PTK7 NTF (aa 1–703) significantly increased cAMP levels in WT GPR133-expressing, but not H543R uncleavable mutant-expressing cells. COL6 (native purified human collagen VI) had no effect on signaling. Two-way ANOVA: GPR133 variant expressed F 2, 80 = 11.65, p < 0.0001; protein coating F 2, 80 = 12.90, p < 0.0001; interaction of factors F 4, 80 = 9.24, p < 0.0001. Tukey’s multiple comparisons: GPR133-WT-expressing cells on PTK7 NTF-coated vs. uncoated dishes, p < 0.0001; GPR133-WT-expressing cells on PTK7 NTF-coated vs. COL6-coated dishes, p < 0.0001). All other comparisons not significant; n = 5–15 independent experiments. (D) Combinatorial effect of PTK7 binding and p13 Stachel peptide treatment on GPR133 signaling. HEK293T cells expressing either WT GPR133 or empty vector control were seeded onto PTK7 NTF-coated (1.17 μg/cm 2 ) or control-coated wells. Cells were then treated with synthetic p13 Stachel peptide (500 μM), inactive control peptide (500 μM), or solvent controls, and cAMP levels were measured by HTRF. The combination of PTK7 NTF binding and Stachel peptide elicited an additive response in GPR133 signaling compared with the individual treatments. Two-way ANOVA, GPR133 expression effect F 1, 48 = 365.1, p < 0.0001; treatment effect F 7, 48 = 13.2, p < 0.0001; interaction of GPR133 expression and treatment F 7, 48 = 13.2, p < 0.0001. Tukey’s multiple comparisons: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. n = 4 independent experiments. Data are depicted as mean ± SEM. See also and .

Article Snippet: Human Collagen Type VI , Rockland , 009-001-108.

Techniques: Expressing, Mutagenesis, Plasmid Preparation, Co-Culture Assay, Purification, Variant Assay, Binding Assay, Control, Solvent