Journal: Human Molecular Genetics

Article Title: Deficiency of Patched 1-induced Gli1 signal transduction results in astrogenesis in Swedish mutated APP transgenic mice

doi: 10.1093/hmg/ddu370

Figure Lengend Snippet: Shh signal components are altered in the hippocampus of both APP23 mice and AD patients. (A) Ptc1 expression (green) was visualized by immunostaining in the dentate gyrus with an increased Ptc1 expression in 12-month-old APP23 mice, especially in the cells of hilus region. Neurons were stained with red. Scale bar: 50 µm. (B) Western blot showed the protein expressions of Shh signaling components in the hippocampus of 3-month-old APP23 mice. (C) Statistical analysis showed that the Smo expression level is increased (n = 10, Student's t-test, **P < 0.01) in APP23 mice of 3 months old of age, whereas the levels of Ptc1, Ptc2, Gli1, Gli2 and Gli3 expressions were still not significantly changed. (D) Western blot showed the protein expressions of Shh signaling components in the hippocampus of 12-month-old APP23 mice. (E) Statistical analysis showed that the expression levels of Ptc1, Ptc2, Smo and Gli1 were increased (n = 10, Student's t-test, *P < 0.05, **P < 0.01 versus corresponding WT) in APP23 mice of 12 months old of age, whereas the levels of Gli2 and Gli3 expressions were not significantly changed. (F) Western blot showed the protein expressions of Shh signaling components in the hippocampus of 24-month-old APP23 mice. (G) Statistical analysis showed that the expression levels of Smo, Gli1 and Gli2 are increased (n = 10, Student's t-test, *P < 0.01 versus corresponding WT) in APP23 mice of 24 months old of age, whereas the levels of Ptc1, Ptc2 and Gli3 expressions were significantly reduced (n = 10, Student's t-test, *P < 0.05, **P < 0.01 versus corresponding WT). (H) Western blot showed that Shh signaling components were expressed in the hippocampus of isolated from human brains of HC and AD patients. (I) The protein levels of Shh, Smo, Gli1 and Gli2 expression were significantly increased, whereas the expression levels of Ptc1, Ptc2 and Gli3 were significantly reduced in comparison with corresponding health controls without dementia (n = 5, Student's t-test, *P < 0.05, **P < 0.01 versus HC).

Article Snippet: The blot was probed by rat anti-ShhN terminal protein (1:1000, MAB4641, R&D), rabbit anti-patched 1 (Ptc1, 1:2000, sc-9016, Santa Cruz Biotechnology), rabbit anti-patched 2 (Ptc2, PA1-46223, 1:1000, Thermo-Scientific), rabbit anti-smoothened (Smo, 1:2000, sc-13943, clone:488-787, Santa Cruz Biotechnology), rabbit anti-Gli1 (1:1000, AB3444, Millipore Bioscience Research Reagents), rabbit anti-Gli2 (Chip grade, ab26056, 1:1000, Abcam) and rabbit anti-Gli3 (ab69838, 1:1000, Abcam).

Techniques: Expressing, Immunostaining, Staining, Western Blot, Isolation

Journal: Human Molecular Genetics

Article Title: Deficiency of Patched 1-induced Gli1 signal transduction results in astrogenesis in Swedish mutated APP transgenic mice

doi: 10.1093/hmg/ddu370

Figure Lengend Snippet: Aβ1–42 not Aβ1–40 increases the expressions of Shh signaling in hippocampal GPCs derived from WT mice. The suspended cells were incubated with the presence of 0, 0.1, 1, 10 or 100 µm of Aβ1–42 or Aβ1–40 for 3 days. (A) Shh expression with different molecular weights was shown by western blot from the cultured medium and the cells treated with Aβ1–42. (B) Shh levels were significantly increased in the conditioned medium (normalized to corresponding cell lysate) and cultured cells in a dose-dependent manner with Aβ1–42 treatment (ANOVA test, *P < 0.05, **P < 0.01). (C) The expression of Shh signaling components was shown in the cells with Aβ1–42 treatment. (D) The levels of Ptc1, Ptc2, Smo, Gli1 and Gli2 were significant elevated (ANOVA test, *P < 0.05, **P < 0.01) but not significant changes of Gli3 levels with the exposure of Aβ1–42 (ANOVA test, P > 0.05). (E) Shh expression was shown from the conditioned medium and GPCs treated with Aβ1–40. (F) Shh levels were not significantly increased in the conditioned medium (normalized to corresponding cell lysate) and the cultured GPCs with the Aβ1–40 treatment (ANOVA test, P > 0.05). (G) The expression of Shh signaling component molecules was shown in the GPC lysate with the Aβ1–40 treatment. (H) The levels of the components were not significantly changed with the Aβ1–40 treatment. (I) Western blot showed Ptc1 expression with the exogenous Shh application. (J) A statistical analysis showed a significant promotion of Ptc1 expression levels in WT hippocampus progenitor cells for 3 days of Shh incubation (ANOVA test, **P < 0.01). Experiments were repeated three times per condition.

Article Snippet: The blot was probed by rat anti-ShhN terminal protein (1:1000, MAB4641, R&D), rabbit anti-patched 1 (Ptc1, 1:2000, sc-9016, Santa Cruz Biotechnology), rabbit anti-patched 2 (Ptc2, PA1-46223, 1:1000, Thermo-Scientific), rabbit anti-smoothened (Smo, 1:2000, sc-13943, clone:488-787, Santa Cruz Biotechnology), rabbit anti-Gli1 (1:1000, AB3444, Millipore Bioscience Research Reagents), rabbit anti-Gli2 (Chip grade, ab26056, 1:1000, Abcam) and rabbit anti-Gli3 (ab69838, 1:1000, Abcam).

Techniques: Derivative Assay, Incubation, Expressing, Western Blot, Cell Culture

Journal: Human Molecular Genetics

Article Title: Deficiency of Patched 1-induced Gli1 signal transduction results in astrogenesis in Swedish mutated APP transgenic mice

doi: 10.1093/hmg/ddu370

Figure Lengend Snippet: The schematic presentation shows Shh-Ptc1-Gli1 signaling pathway in normal (Left) condition and disease (right) conditions. In general, Shh binds Ptc, which activates and promotes the effecter Gli1. Gli1 enters the nucleus and binds to Ptc promoter and proliferative genes. Excessive Aβ in AD brains promotes and activates GSK-3β, which inhibits Gli1 and, in turn, decreased the transcription of Ptc and proliferative genes.

Article Snippet: The blot was probed by rat anti-ShhN terminal protein (1:1000, MAB4641, R&D), rabbit anti-patched 1 (Ptc1, 1:2000, sc-9016, Santa Cruz Biotechnology), rabbit anti-patched 2 (Ptc2, PA1-46223, 1:1000, Thermo-Scientific), rabbit anti-smoothened (Smo, 1:2000, sc-13943, clone:488-787, Santa Cruz Biotechnology), rabbit anti-Gli1 (1:1000, AB3444, Millipore Bioscience Research Reagents), rabbit anti-Gli2 (Chip grade, ab26056, 1:1000, Abcam) and rabbit anti-Gli3 (ab69838, 1:1000, Abcam).

Techniques:

Journal: Pharmaceutics

Article Title: Persistent Properties of a Subpopulation of Cancer Cells Overexpressing the Hedgehog Receptor Patched

doi: 10.3390/pharmaceutics14050988

Figure Lengend Snippet: A small population of ACC cells H295R overexpresses Ptch1 at the plasma membrane . ( A ) H295R were labeled with an anti-Ptch1 antibody directed against the extracellular loop and cells presenting Ptch1 at their plasma membrane (H295R-PM-Ptc+ AF594+ cells) were sorted. AF594+ in blue represents the percentage of cells with Ptch1 at the cell surface (H295R-PM-Ptc+ cells). ( B ) Surface labeling of Ptch1 using anti-Ptch1 antibody directed against the extracellular loop of Ptch1 (Alexa 594 in red) on nonpermeabilized parental H295R and H295R-PM-Ptc+ cells. Nuclei were stained with DAPI (in blue). The histogram represents the mean ± SEM of Alexa 594 fluorescence intensity per cell (****: p -value < 0.00005 ( p -value = 2 × 10 −36 )).

Article Snippet: Nitrocellulose membranes were then incubated overnight at 4 °C with rat anti-Ptch1 antibody (MAB41051 R&D Systems; 1 μg/mL), rabbit anti-ADCY2 (Abnova) (1/1000), rabbit anti-Gli2 (Abcam) (1/1000), rabbit anti-ABCG2 (Genetex) (1/500), rabbit anti-GRK5 (Genetex) (1/1000), rabbit anti-SLUG (SNAI2) (Genetex) (1/2500), rabbit anti-SOX5 (Genetex) (1/1000), rabbit anti-Nanog (Cell Signaling Technology) (1/1000), and mouse anti-tubulin (Sigma) (1/1000) or rabbit anti-GAPDH (Abcam ab37168) (1/10,000).

Techniques: Clinical Proteomics, Membrane, Labeling, Staining, Fluorescence

Journal: Pharmaceutics

Article Title: Persistent Properties of a Subpopulation of Cancer Cells Overexpressing the Hedgehog Receptor Patched

doi: 10.3390/pharmaceutics14050988

Figure Lengend Snippet: H295R-PM-Ptc+ cells are more resistant to chemotherapy than parental cells . ( A ) Doxorubicin (dxr) cytotoxicity. H295R and H295R-PM-Ptc+ cells were treated for 48 h with increasing concentrations of dxr before cell viability measure. ( B ) Doxorubicin IC50 of H295R-PM-Ptc+ and H295R parental cells in the absence or the presence of 10 μM of the Ptch1 efflux inhibitor methiothepin. ( C ) H295R-PM-Ptc+ cells accumulate less doxorubicin than parental H295R cells. Cells on coverslips were incubated with 2 μM dxr for 15, 30, 60, 180 and 240 min and immediately fixed with PFA. Dxr fluorescence was acquired using a filter for Alexa 594 and quantified using ImageJ software. About 100 cells (from three wells) were scored per condition per experiment. All data presented are the mean ± SEM of at least 3 independent experiments. Significance is attained at p -value < 0.05 (*), (**** p < 0.00005).

Article Snippet: Nitrocellulose membranes were then incubated overnight at 4 °C with rat anti-Ptch1 antibody (MAB41051 R&D Systems; 1 μg/mL), rabbit anti-ADCY2 (Abnova) (1/1000), rabbit anti-Gli2 (Abcam) (1/1000), rabbit anti-ABCG2 (Genetex) (1/500), rabbit anti-GRK5 (Genetex) (1/1000), rabbit anti-SLUG (SNAI2) (Genetex) (1/2500), rabbit anti-SOX5 (Genetex) (1/1000), rabbit anti-Nanog (Cell Signaling Technology) (1/1000), and mouse anti-tubulin (Sigma) (1/1000) or rabbit anti-GAPDH (Abcam ab37168) (1/10,000).

Techniques: Incubation, Fluorescence, Software

Journal: Pharmaceutics

Article Title: Persistent Properties of a Subpopulation of Cancer Cells Overexpressing the Hedgehog Receptor Patched

doi: 10.3390/pharmaceutics14050988

Figure Lengend Snippet: Differentially expressed genes (DEG) between H295R-PM-Ptc+ and parental H295R cells selected for their role in cancer. Genes overexpressed are indicated in red and genes underexpressed are in blue.

Article Snippet: Nitrocellulose membranes were then incubated overnight at 4 °C with rat anti-Ptch1 antibody (MAB41051 R&D Systems; 1 μg/mL), rabbit anti-ADCY2 (Abnova) (1/1000), rabbit anti-Gli2 (Abcam) (1/1000), rabbit anti-ABCG2 (Genetex) (1/500), rabbit anti-GRK5 (Genetex) (1/1000), rabbit anti-SLUG (SNAI2) (Genetex) (1/2500), rabbit anti-SOX5 (Genetex) (1/1000), rabbit anti-Nanog (Cell Signaling Technology) (1/1000), and mouse anti-tubulin (Sigma) (1/1000) or rabbit anti-GAPDH (Abcam ab37168) (1/10,000).

Techniques: Activation Assay, Expressing, Inhibition, Gene Expression, Migration, Marker, Biomarker Discovery

Journal: Pharmaceutics

Article Title: Persistent Properties of a Subpopulation of Cancer Cells Overexpressing the Hedgehog Receptor Patched

doi: 10.3390/pharmaceutics14050988

Figure Lengend Snippet: Composition of active modules containing one or more of the identified genes of interest listed in Table 1 (in bold) with genes upregulated in red and genes downregulated in blue, representative enrichment and role of differentially expressed genes (DEGs) in cancers.

Article Snippet: Nitrocellulose membranes were then incubated overnight at 4 °C with rat anti-Ptch1 antibody (MAB41051 R&D Systems; 1 μg/mL), rabbit anti-ADCY2 (Abnova) (1/1000), rabbit anti-Gli2 (Abcam) (1/1000), rabbit anti-ABCG2 (Genetex) (1/500), rabbit anti-GRK5 (Genetex) (1/1000), rabbit anti-SLUG (SNAI2) (Genetex) (1/2500), rabbit anti-SOX5 (Genetex) (1/1000), rabbit anti-Nanog (Cell Signaling Technology) (1/1000), and mouse anti-tubulin (Sigma) (1/1000) or rabbit anti-GAPDH (Abcam ab37168) (1/10,000).

Techniques: Migration, Cell Differentiation, Activation Assay, Membrane, Activity Assay

Journal: bioRxiv

Article Title: Inhibition of PTCH1 drug efflux activity enhances chemotherapy efficacy against triple negative breast cancer

doi: 10.1101/2025.09.03.673759

Figure Lengend Snippet: A . Relative PTCH1 RNA expression in TNBC (red), Her2 (blue), luminal B (LB, green), and luminal A (LA, orange) in the TCGA cohort is illustrated by box plots (log 2 transformed). Outliers are shown within each population (open circles). Student’s t test was used to compare RNA levels between two groups. The p values are indicated (*P < 0.05; **P < 0.01; ***P < 0.001; ns P > 0.05). B . Normalized PTCH1 mRNA expression according to TNBC (IHC) status from all DNA microarray data from bc-GenExMiner v4.5 illustrated by box plots (log 2 transformed) (Jezequel et al 2021). C . Boxplots showing PTCH1 mRNA expression (normalized counts from RNA seq data) in circulating tumor cells (CTC) isolated from metastasis breast cancer patient peripheral blood (Wurth R et al 2025). D . High PTCH1 mRNA expression is associated with a poorer prognosis in breast cancer. Distant Metastasis Free Survival (DMFS) and disease-free survival (DFS) data based on PTCH1 mRNA expression were obtained using the exhaustive prognostic analysis on all status breast cancers on the bc-GenExMiner v4.5 web portal, and illustrated by Kaplan–Meier (KM) curves for all ER and PR breast cancers with positive nodules. The obtained Hazard Ratio (HR) with 95% confidence interval and log-rank p-values are shown.

Article Snippet: After 1 h at room temperature in blocking buffer (20 mmol/L Tris-HCl pH 7.5, 45 mmol/L NaCl, 0.1 % Tween-20, and 5 % non-fat milk), nitrocellulose membranes were incubated overnight at 4 °C with the monoclonal rat anti-PTCH1 antibody from R&D system biotechne clone 413220 (1/1000), the monoclonal mouse anti-P-gp antibody from abcam (ab3366, 1/1000), the rabbit anti-ABCG2 antibody from GeneTex (GTX50793, 1/500) or the rabbit anti-GAPDH antibody from Elabscience (1/20000).

Techniques: RNA Expression, Transformation Assay, Expressing, Microarray, RNA Sequencing, Isolation

Journal: bioRxiv

Article Title: Inhibition of PTCH1 drug efflux activity enhances chemotherapy efficacy against triple negative breast cancer

doi: 10.1101/2025.09.03.673759

Figure Lengend Snippet: Distant Metastasis Free Survival (DMFS), disease-free survival (DFS) and overall survival (OS) data based on PTCH1 mRNA expression were obtained from the intrinsic molecular subtypes’ prognostic analysis on ER+/HER2-high proliferative breast cancers performed on bc-GenExMiner v4.5 web portal and illustrated by Kaplan–Meier curves. The obtained Hazard Ratio (HR) with 95% confidence interval and log-rank P-values are shown.

Article Snippet: After 1 h at room temperature in blocking buffer (20 mmol/L Tris-HCl pH 7.5, 45 mmol/L NaCl, 0.1 % Tween-20, and 5 % non-fat milk), nitrocellulose membranes were incubated overnight at 4 °C with the monoclonal rat anti-PTCH1 antibody from R&D system biotechne clone 413220 (1/1000), the monoclonal mouse anti-P-gp antibody from abcam (ab3366, 1/1000), the rabbit anti-ABCG2 antibody from GeneTex (GTX50793, 1/500) or the rabbit anti-GAPDH antibody from Elabscience (1/20000).

Techniques: Expressing

Journal: bioRxiv

Article Title: Inhibition of PTCH1 drug efflux activity enhances chemotherapy efficacy against triple negative breast cancer

doi: 10.1101/2025.09.03.673759

Figure Lengend Snippet: A . PTCH1 mRNA expression (log 2 transformed) in various luminal and HER2 breast cancer cell lines (dark green) and in TNBC cell lines (other colors). TNBC cell lines are depicted according to the “Lehmann TNBC subtype” nomenclature : basal-like 1 (yellow), basal-like 2 (pale green), immunomodulatory (brown), luminal androgen receptor (dark pink), mesenchymal (pale pink) and mesenchymal stem-like (pink). B . PTCH1 protein expression in three TNBC cell lines. Western-blot was performed on 50 µg extracts from TNBC cell lines (MDA-MB-231, HCC-38 and MDA-MB-468) with antibodies directed against PTCH1. PTCH1 and GAPDH signals were quantified using ImageJ software. Data presented are the mean ± SEM of at least 3 independent experiments. Significance is calculated using Oneway ANOVA Turkey’s multiple comparisons test and attained at P < 0.05 (*).

Article Snippet: After 1 h at room temperature in blocking buffer (20 mmol/L Tris-HCl pH 7.5, 45 mmol/L NaCl, 0.1 % Tween-20, and 5 % non-fat milk), nitrocellulose membranes were incubated overnight at 4 °C with the monoclonal rat anti-PTCH1 antibody from R&D system biotechne clone 413220 (1/1000), the monoclonal mouse anti-P-gp antibody from abcam (ab3366, 1/1000), the rabbit anti-ABCG2 antibody from GeneTex (GTX50793, 1/500) or the rabbit anti-GAPDH antibody from Elabscience (1/20000).

Techniques: Expressing, Transformation Assay, Western Blot, Software

Journal: Molecular Cancer

Article Title: Co-activation of hedgehog and AKT pathways promote tumorigenesis in zebrafish

doi: 10.1186/1476-4598-8-40

Figure Lengend Snippet: Elevated phospho-AKT1 and Patched 1 levels in zebrafish tumors . A, B, a 3-week-old transgenic fish with trunk tumor (B) and its age-matched tumor-free fish (A). C, D, a 12-week-old fish with an eye tumor (D) and its age-matched tumor free fish (C). Immunofluorescence was done on cryosections for the above fish. E-H, an astrocytoma from a double transgenic fish showed elevated levels of both Patched 1 (F) and phosphorylated AKT (H). Immunofluoresence was done on paraffin sections for this tumor. Negative controls for Patched 1 (E) and phosphorylated AKT (G) were treated the same way except no primary antibodies were added. N, Notochord; AKT-P, phosphorylated AKT. Scale bars, 100 μm for A-D, 50 μm for E-H.

Article Snippet: A rabbit anti-GFP polyclonal antibody (Invitrogen, 1:500), a rabbit anti-phospho-AKT (Thr308) antibody (C31E5, Cell Signalling Technology, 1:200), and a goat anti-Patched 1 (zebrafish) antibody (Everest Biotech, 1:200) were used for immunofluorescence studies of the tumors.

Techniques: Transgenic Assay, Immunofluorescence