Journal:

Article Title: Essential requirement for Wnt signaling in proliferation of adult small intestine and colon revealed by adenoviral expression of Dickkopf-1

doi: 10.1073/pnas.2536800100

Figure Lengend Snippet: Expression analysis of Wnt/β-catenin target genes, CD44 and EphB2, in the gastrointestinal tract of Ad Dkk1- or Ad Fc-treated adult C57BL/6 mice (12–16 weeks old). Organs were harvested 2 days after Ad Dkk1 i.v injection (109 pfu). (Left) Ad Dkk1 repression of CD44 expression in proliferative zones of all levels of the gastrointestinal epithelium. Arrowheads indicate the absence of CD44 immunoreactivity in proliferative compartments of the intestinal epithelium in Ad Dkk1 animals. *, residual CD44 staining in nonepithelial lamina propria. (Right) Ad Dkk1 repression of EphB2 in small intestine and colon. Repression was weaker in ascending colon and no repression was observed in stomach. EphB2 immunofluorescence was performed with Alexa 488 detection of EphB2 immunoreactivity (green) and Hoechst 33342 nuclear counterstain (blue). Stomach (st), duodenum (du), jejunum (je), ileum (il), cecum (ce), ascending colon (ac), and descending colon (dc) are shown.

Article Snippet: The following antibodies were used: Rat anti-mouse CD44 (1:100; BD Pharmingen), rat anti-mouse Ki67 (1:100; DAKO), goat anti-mouse EphB2 (1:100; R & D Systems), rabbit anti-rat FABP (1:100; Novus Biologicals, Littleton, CO), and rabbit anti-human lysozyme (1:100; DAKO).

Techniques: Expressing, Injection, Staining, Immunofluorescence

Journal: Cancers

Article Title: Spatial EGFR Dynamics and Metastatic Phenotypes Modulated by Upregulated EphB2 and Src Pathways in Advanced Prostate Cancer

doi: 10.3390/cancers11121910

Figure Lengend Snippet: Single-cell RNA-seq showing upregulation of Src and Eph-ephrin signaling in advanced prostate cancer cells. ( A ) Dot plot of differentially expressed (DE) genes (n = 171) in Src signaling. LNCaP, LNCaP-Abl, and PC3 are clusters distinctly separated from the other clusters. ( B ) Dot plot of DE gene expression (n = 20) in Eph-Ephrin signaling. Unpaired t -test, ** p < 0.01, *** p < 0.0001. ( C ) The expression of 16 representative genes from EPH Transcripts of 16 genes in four cell lines were validated using bulk RNA qRT-PCR. ( D ) mRNA levels of EPHB2 and SRC predict poor prognosis in The Cancer Genome Atlas (TCGA) prostate cancer patient cohort using Kaplan–Myer estimation analysis. The red line represents the patients with a high expression of the genes with more than 2 SD as compared to the patients presented in the blue line.

Article Snippet: Antibodies against SRC1 (2109, Cell Signaling Biotechnology, Danvers, MA, USA), EPHB2 (AF467-SP, R&D Systems), EGFR (MS-311, Thermo Fisher Scientific), and secondary antibodies with FITC and Alexa 530 (712-585-153, Jackson ImmunoResearch Laboratories, West Grove, PA, USA) were purchased commercially.

Techniques: RNA Sequencing, Gene Expression, Expressing, Quantitative RT-PCR

Journal: Cancers

Article Title: Spatial EGFR Dynamics and Metastatic Phenotypes Modulated by Upregulated EphB2 and Src Pathways in Advanced Prostate Cancer

doi: 10.3390/cancers11121910

Figure Lengend Snippet: Dasatinib inhibits proliferation, migration, invasion, and EGFR diffusivity in advanced prostate cancer cells. ( A ) Src is highly upregulated in PC3 (2.5x) and DU145 (1.6x) as compared to LNCaP shown in Western blots. EphB2 is overexpressed in both PC3 and DU145. Both proteins are almost not expressed in LNCaP-Abl. ( B ) Src is present in these cell lines. There is an intense level of Src on the PC3 cell membrane. ( C ) Immunostaining of EphB2 protein is present in plasma and membrane. ( D ) Image-based IncuCyte assays allow us to conduct the time-lapse analysis of cell proliferation, migration, and invasion of the cells treated with or without dasatinib. The dasatinib significantly inhibits the proliferation, migration, and invasion of DU145 and PC3 cells but reduces the EGFR diffusivity in only PC3 cells. The mean value of each bar was measured at the end time of each assay or at the 48th hour. All statistical analyses were performed using the unpaired t -test. The asterisk represents the level of statistical significance for t -test: *** p < 0.001, ** p < 0.01, * p < 0.05. The error bar represents the standard error of the mean. The RWD stands for relative wound density.

Article Snippet: Antibodies against SRC1 (2109, Cell Signaling Biotechnology, Danvers, MA, USA), EPHB2 (AF467-SP, R&D Systems), EGFR (MS-311, Thermo Fisher Scientific), and secondary antibodies with FITC and Alexa 530 (712-585-153, Jackson ImmunoResearch Laboratories, West Grove, PA, USA) were purchased commercially.

Techniques: Migration, Western Blot, Membrane, Immunostaining, Clinical Proteomics

Journal: Cancers

Article Title: Spatial EGFR Dynamics and Metastatic Phenotypes Modulated by Upregulated EphB2 and Src Pathways in Advanced Prostate Cancer

doi: 10.3390/cancers11121910

Figure Lengend Snippet: Disruption of EphB2/Src pathways leads to attenuated cell motility, invasion, and EGFR diffusion in advanced prostate cancer cells. ( A ) Effective gene knockdowns in siRNA-treated DU145 and PC3. ( B ) Structured Illumination Microscopy (SIM) images of siRNA treated cells. Maximum intensity projection on the xy plane and orthogonal cross-sections (xz and yz) of DU145 and PC3 siRNA treated cells. ( C ) Quantification of cortical actin based on fluorescence intensities of xz and yz orthogonal projections along the apical plasma membrane. The number of projections analyzed is labeled on each bar. ( D ) EGFR diffusivities of the siRNAs treated cells. The error bar represents the standard error of the mean. ( E , F ) The image-based assays allow us to conduct the time-lapse analysis of cell migration and invasion on the siRNA-treated cells. The error bar represents the standard deviation. All statistical analyses were performed using the unpaired t -test. The asterisk represents the level of statistical significance for t -test: *** p < 0.001, ** p < 0.01, * p < 0.05. ( G ) Schematic shows the effects of EMT-induced actin reorganization on EGFR dynamics and the Src/EphB2 induced signaling from the plasma membrane that controls cell behavior.

Article Snippet: Antibodies against SRC1 (2109, Cell Signaling Biotechnology, Danvers, MA, USA), EPHB2 (AF467-SP, R&D Systems), EGFR (MS-311, Thermo Fisher Scientific), and secondary antibodies with FITC and Alexa 530 (712-585-153, Jackson ImmunoResearch Laboratories, West Grove, PA, USA) were purchased commercially.

Techniques: Disruption, Diffusion-based Assay, Microscopy, Fluorescence, Clinical Proteomics, Membrane, Labeling, Migration, Standard Deviation

Journal: Scientific reports

Article Title: Tofacitinib and budesonide treatment affect stemness and chemokine release in IBD patient-derived colonoids.

doi: 10.1038/s41598-025-86314-2

Figure Lengend Snippet: Fig. 5. LGR5 and EphB2 protein expression in differentiated colonoids in response to tofacitinib or budesonide treatment. (a) Immunoblots showing LGR5 and EphB2 protein expression in colonoids from N = 6 donors (4HC and 2UC) cultured at 2% oxygen. Original full blot images are presented in the Supplementary Figure S1. (b,c) Quantification of LGR5 and EphB2 at group level in response to different treatments (1–6) (d,e) Paired quantification of LGR5 and EphB2 protein expression between selected treatments. The colonoids were pre-treated with tofacitinib or budesonide for 3 days alone or before TNF + Poly(I:C) (see Fig. 1). The x-axis shows the treatment, and the y-axis shows fold induction. Fold expression is generated by normalizing to vehicle control (DMSO 0.033%) for each donor and further normalized to GAPDH expression. Differences were evaluated using paired t-test. *P < 0.05.

Article Snippet: The following antibodies were used: LGR5 Mouse mAb (1:1000, #MA5-25644, Invitrogen), EPH Receptor B2 (EphB2) Goat mAb (1:1000 dilution, #AF467, R&D Systems), Phospho-MLKL (Ser358) (E7G7P) Rabbit mAb (1:1000 dilution, #18640, Cell Signaling Technology), MLKL (E7V4W) Mouse mAb (1:1000 dilution, #26539, Cell Signaling Technology), Caspase-3/cleaved caspase-3 (D3R6Y) Rabbit mAb (1:1000 dilution, #14220, Cell Signaling Technology), and GAPDH (D16H11) XP rabbit monoclonal antibody (1:5000 dilution, #5174, Cell Signaling Technology).

Techniques: Expressing, Western Blot, Cell Culture, Generated, Control

Journal: Neural Development

Article Title: EphB2 signaling regulates lesion-induced axon sprouting but not critical period length in the postnatal auditory brainstem

doi: 10.1186/1749-8104-8-2

Figure Lengend Snippet: EphB2 expression at P7. EphB2 is expressed in the VCN (asterisk, A ) and in the MNTB ( B ) at P7. Immunohistochemistry results ( A - B ) were confirmed with X-gal histochemistry on tissue from P7 EphB2 lacZ/lacZ mice ( C - F ). X-gal staining is present in VCN (asterisk, C ) and in axons and calyces in MNTB ( D and E ). No axonal staining is present at the midline (arrow, E ), where VCN axons cross to reach the contralateral MNTB. VCN axons in the ventral acoustic stria (arrowheads, F and G ), a population of proximal uncrossed VCN axons, also lack staining (black arrowheads, F ). Unilateral VCN NeuroVue dye labeling is shown in panel ( G ) to demonstrate the location of the ventral acoustic stria (white arrowheads) in panel ( F ) in relation to VCN (asterisks, F and G ) and lateral superior olive (LSO). Scale bars in panels B (also applies to D ) and E represent 100 μm3

Article Snippet: Slides were transferred to tris-buffered saline (TBS) and 0.05% Triton X-100 (TBST), incubated in blocking solution containing 3% normal rabbit serum (Vector Laboratories, Burlingame, CA) in TBST for one hour, and incubated overnight with goat anti-EphB2 polyclonal antibody (R&D Systems, Minneapolis, MN) diluted to 10 μg/mL in 1% normal rabbit serum in TBST.

Techniques: Expressing, Immunohistochemistry, Staining, Labeling

Journal: Neural Development

Article Title: EphB2 signaling regulates lesion-induced axon sprouting but not critical period length in the postnatal auditory brainstem

doi: 10.1186/1749-8104-8-2

Figure Lengend Snippet: EphB2 expression at P10. At P10, EphB2 is expressed in the VCN (asterisk, A ) and MNTB ( B ). Immunohistochemistry results ( A - B ) were confirmed with X-gal histochemistry on tissue from P10 EphB2 lacZ/lacZ mice ( C - F ). Similar to X-gal staining at P7, staining is present in VCN (asterisk, C ) and in axons and calyces in MNTB ( D and E ). No axonal staining is present at the midline (arrow, E ), where VCN axons cross to reach the contralateral MNTB. VCN axons in the ventral acoustic stria also lack staining (arrowheads, F ). Scale bar in panel ( A ) represents 200 μm and also applies to panels ( C and F ). Scale bars in panels B (also applies to D ) and E represent 100 μm.

Article Snippet: Slides were transferred to tris-buffered saline (TBS) and 0.05% Triton X-100 (TBST), incubated in blocking solution containing 3% normal rabbit serum (Vector Laboratories, Burlingame, CA) in TBST for one hour, and incubated overnight with goat anti-EphB2 polyclonal antibody (R&D Systems, Minneapolis, MN) diluted to 10 μg/mL in 1% normal rabbit serum in TBST.

Techniques: Expressing, Immunohistochemistry, Staining

Journal: Neural Development

Article Title: EphB2 signaling regulates lesion-induced axon sprouting but not critical period length in the postnatal auditory brainstem

doi: 10.1186/1749-8104-8-2

Figure Lengend Snippet: Unilateral VCN removal at P7 induces ipsilateral projections to the MNTB. After unilaterally removing the VCN at P7, labeled calyces are seen in the MNTB contralateral (MNTBc) to the intact, dye-filled VCN, as well as in the ipsilateral MNTB (MNTBi, arrows) of wild type ( A - D ) and EphB2 −/− ( E - H ) mice. The mean ipsilateral to contralateral (I/C) ratio from EphB2 −/− mice was greater than the mean I/C ratio from wild type mice (I, P < 0.01). Scale bar in panel ( A ) represents 200 μm and also applies to panels ( B - H ).

Article Snippet: Slides were transferred to tris-buffered saline (TBS) and 0.05% Triton X-100 (TBST), incubated in blocking solution containing 3% normal rabbit serum (Vector Laboratories, Burlingame, CA) in TBST for one hour, and incubated overnight with goat anti-EphB2 polyclonal antibody (R&D Systems, Minneapolis, MN) diluted to 10 μg/mL in 1% normal rabbit serum in TBST.

Techniques: Labeling

Journal: Neural Development

Article Title: EphB2 signaling regulates lesion-induced axon sprouting but not critical period length in the postnatal auditory brainstem

doi: 10.1186/1749-8104-8-2

Figure Lengend Snippet: Numbers of labeled calyces after cochlear nucleus removal

Article Snippet: Slides were transferred to tris-buffered saline (TBS) and 0.05% Triton X-100 (TBST), incubated in blocking solution containing 3% normal rabbit serum (Vector Laboratories, Burlingame, CA) in TBST for one hour, and incubated overnight with goat anti-EphB2 polyclonal antibody (R&D Systems, Minneapolis, MN) diluted to 10 μg/mL in 1% normal rabbit serum in TBST.

Techniques: Labeling

Journal: Neural Development

Article Title: EphB2 signaling regulates lesion-induced axon sprouting but not critical period length in the postnatal auditory brainstem

doi: 10.1186/1749-8104-8-2

Figure Lengend Snippet: Unilateral VCN removal at P10 does not induce projections to the ipsilateral MNTB. Labeled calyces are seen in the contralateral MNTB (MNTBc) of wild type (A and C) and EphB2 −/− mice ( E and G) after removing the VCN at P10, but not in the ipsilateral MNTB (MNTBi) of either mouse (B, D, F, H) . The mean ipsilateral/contralateral (I/C) ratios did not differ after unilateral VCN removal at P10 ( I, P > 0.05). Taken together, deafferentation-induced innervation of the ipsilateral MNTB decreases postnatally in wild type and EphB2 −/− mice (J) . EphB2 −/− mice have more ipsilateral sprouting after unilateral deafferentation at P2 and P7, but not at P10, when the critical period for this process has ended. Y-axis values in panel ( J ) are I/C ratios represented as a percentage of the I/C ratio obtained after cochlea removal in wild type mice at P2. P2 data is from Nakamura et al ., 2012. Scale bar in panel ( A ) represents 200 μm and also applies to panels ( B - H ).

Article Snippet: Slides were transferred to tris-buffered saline (TBS) and 0.05% Triton X-100 (TBST), incubated in blocking solution containing 3% normal rabbit serum (Vector Laboratories, Burlingame, CA) in TBST for one hour, and incubated overnight with goat anti-EphB2 polyclonal antibody (R&D Systems, Minneapolis, MN) diluted to 10 μg/mL in 1% normal rabbit serum in TBST.

Techniques: Labeling