Journal:

Article Title: Renal Fibrosis

doi:

Figure Lengend Snippet: Tubular epithelial cell interactions with collagen types I and IV. MCT cells adhere preferably to type IV collagen (383 ±16.6% compared to uncoated plastic control) than to type I collagen (232.2 ± 20.5%) in cell adhesion assay (A, left). After induction of EMT with TGF-β1 and EGF, MCT cells with a fibroblast-like morphology attached increasingly to collagen type I (396.7 ± 24.3% compared to uncoated plastic control), whereas adhesion to type IV collagen was less abundant (299.5 ± 20.6%) (A, right). MCT cells grown in K1 medium adhere strongly to type IV collagen and seem to display a round-shaped, epithelial cell-like, morphology (B, left). MCT cells that were pretreated with TGF-β1 and EGF gain in capacity to attach to type I collagen and appear to have a more spindle-shaped morphology (B, right). Cultivation on type I collagen increased FSP-1 expression in MCT cells that were grown in K1 medium (140 ± 9.3% compared to uncoated plastic control), whereas cultivation on type IV collagen had no effect on FSP-1 expression in untreated cells (C, left) as was measured by ELISA of cell lysates. When EMT was induced in MCT cells with TGF-β1 and EGF, cultivation on type I collagen further increased levels of FSP-1 expression (130.6 ± 7.3% compared to uncoated plastic control), whereas coating with type IV collagen decreased levels of FSP-1 expression (58.1 ± 10.4%) and thus stabilized the epithelial phenotype (C, right). *, P < 0.05; **, P < 0.001. Original magnifications, ×400.

Article Snippet: Recombinant human TGF-β 1 , human epithelial growth factor (EGF), and the neutralizing polyclonal goat antibodies to TGF-β and EGF were purchased from R&D Systems (Minneapolis, MN).

Techniques: Control, Cell Adhesion Assay, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Nature communications

Article Title: Host interneurons mediate plasticity reactivated by embryonic inhibitory cell transplantation in mouse visual cortex.

doi: 10.1038/s41467-021-21097-4

Figure Lengend Snippet: Fig. 7 Transplant-induced ODP depends on NRG1/ErbB4 signaling. a Experimental timeline. b Intrinsic optical imaging was performed to measure ocular dominance plasticity (ODP) in adult transplant recipients around 33–35 days after transplantation (DAT). The wild-type adult recipients received either saline or soluble neuregulin 1 (NRG1) during 4 days of monocular deprivation (MD). c Examples of response amplitude before and after 4D MD in WT transplant recipients that received saline injections or were treated with NRG1 during deprivation. Scale bar = 500 µm. d Four-day MD resulted in a shift of ocular dominance index (ODI) in transplant recipients that received saline injections. Injections of exogenous NRG1 diminished transplant-induced ODI shift. Interneuron transplantation also failed to reactivate ODP in PV-cre;ErbB4flx/flx adult recipients. However, WT adults that were transplanted with cells from PV-cre;ErbB4flx/flx donor exhibited a significant shift in ODI after 4D MD. ODI is calculated as (Contra response amplitude −Ipsi response amplitude)/(Contra response amplitude + Ipsi response amplitude) (Saline n = 7 mice, red; NRG1 n = 6 mice, blue; PV-cre;ErbB4flx/flx recipients n = 6 mice, orange; PV-cre;ErbB4flx/flx donors PreMD: n = 7 mice, PostMD: n = 5 mice, yellow. Saline: **p = 0.004; NRG1: p = 0.344; PV-cre;ErbB4flx/flx

Article Snippet: To test the effect of NRG1 on the response properties of PV interneurons, some mice received subcutaneous injections of soluble NRG1 (R&D Systems, 396-HB; 0.5 μg per animal every 8 h) during MD.

Techniques: Optical Imaging, Transplantation Assay, Saline

Journal: Oncogene

Article Title: Phosphoinositide 3-kinase signaling is critical for ErbB3-driven breast cancer cell motility and metastasis

doi: 10.1038/onc.2011.275

Figure Lengend Snippet: A. Flow cytometry analysis of surface ErbB3 levels in pLXSN (solid line), ErbB3WT (dotted line), and ErbB3-Mutant (dashed line) cells. A mouse anti-ErbB3 primary followed by an APC-labeled secondary was used. The curve with solid shading represents ErbB3WT cells incubated with just the APC-conjugated secondary antibody. B . HRGβ1-induced ErbB3 tyrosine phosphorylation and association to the p85 subunit of PI3K. Serum starved MTLn3 ErbB3WT or ErbB3-Mutant cells were stimulated with buffer or HRGβ1 (0 or 0.4 HRG) for 5 minutes. ErbB3 immunoprecipitates (IP) and whole cell lysates (WCL) were resolved by SDS/PAGE, and probed for (PTyr) or p85α. Membranes were stripped and reprobed to control for ErbB3 levels. C Chemotaxis to HRGβ1 of pLXSN (light gray bars), ErbB3WT (dark gray bars) and ErbB3-Mutant (patterned bars). Data are mean and SEM of 6-27 wells in 3-6 independent experiments. D. In vitro invasion responses of ErbB3WT (black bars) and ErbB3-Mutant cells (patterned bar) into Matrigel-coated transwells stimulated by buffer or 12.5 nM HRGβ1. Data are presented as % Area invaded, and are mean and SEM of 3 independent experiments. *:p< 0.02

Article Snippet: HRGβ1 was from R & D Systems (396-HB).

Techniques: Flow Cytometry, Mutagenesis, Labeling, Incubation, Phospho-proteomics, SDS Page, Control, Chemotaxis Assay, In Vitro

Journal: Oncogene

Article Title: Phosphoinositide 3-kinase signaling is critical for ErbB3-driven breast cancer cell motility and metastasis

doi: 10.1038/onc.2011.275

Figure Lengend Snippet: A. Intravital imaging using multiphoton microscopy of primary mammary tumors of GFP-labeled tumor cells. Time-lapse z-series were acquired and the average total cell motility was determined per 40 μm z-stack (5 sections imaged at 10 μm intervals). Data are means and SEM of 24 – 33 separate z stacks from 7-8 mice per cell line. *: p<0.025, **: p<0.0000042. i/ii panels illustrate examples of (i) ErbB3WT GFP and (ii) ErbB3-Mutant GFP cell motility in vivo . Cells are green with collagen fibers detected by second harmonic scattering in purple. Frames are 6 minutes apart, arrows show migrating tumor cell; Scale bar = 25 μm. Asterisks mark a single cell in i and ii and arrows mark a moving cell in i. B . Length/width ratio comparisons. Top : representative images. Bottom: Mean and SEM of ratios of 50-100 cells from 3 tumors, 5 slices analyzed per z stack. Scale bar = 25 μm. *: p<0.002, **: p<6×10 -5 C . In vivo invasion responses. Microneedles containing buffer or 50 nM HRGβ1 were inserted into primary tumors and invasive cells were quantified. *: p<4×10 -5 , **: p< 5×10 -6 . Data are means and SEM of 9-13 measurements from 3-4 tumors per cell line (pLXSN: gray, ErbB3WT: black, ErbB3-Mutant: patterned). D. In vitro transendothelial migration assay. Cells were plated on the basal side of an endothelial monolayer either alone or mixed with macrophages (+M) and allowed to migrate for 18 hours. for comparison of ErbB3WT+M with ErbB3-Mutant+M: p<4×10 -6 . Data are represented as the fold migration over pLXSN alone, with mean and SEM of 4 independent experiments with each cell line.

Article Snippet: HRGβ1 was from R & D Systems (396-HB).

Techniques: Imaging, Microscopy, Labeling, Mutagenesis, In Vivo, In Vitro, Migration, Comparison

Journal: Oncogene

Article Title: Phosphoinositide 3-kinase signaling is critical for ErbB3-driven breast cancer cell motility and metastasis

doi: 10.1038/onc.2011.275

Figure Lengend Snippet: A . In vitro chemotaxis is inhibited by PIK-75. Migration of ErbB3WT cells in response to a gradient of either 1.25 nM HRGβ1 (black bar) or 1.25 nM HRGβ1 + 0.2 μM PIK-75 (patterned bar). *: p< 0.0001. Data are mean and SEM of measurements from 3 independent experiments. B . In vivo invasion is inhibited by PIK-75. Microneedles containing 50 nM HRGβ1or 50 nM HRGβ1 + 1 μM PIK-75 were inserted into the primary tumor and invasive cells were collected and counted. Data are mean and SEM from 6 needles for HRGβ1 and 10 needles for HRGβ1 + PIK-75 from 3 tumors for each condition. *: p< 1 × 10 -5 . C . In vivo invasion stimulated by 50 nM HRGβ1 is inhibited by systemic application of PIK-75. Mice bearing ErbB3WT tumors were injected i.p. either with vehicle control (black bar) or with 1.5 mg PIK-75 (pattern) 3 hours prior to assay. Microneedles containing 50 nM HRGβ1 were inserted into primary tumors and invasive cells were quantified as described in the methods.*: p< 0.00035. Data are means and SEM of 4-8 measurements from 4 tumors per condition. D . Cell motility in vivo using intravital imaging. Mice bearing tumors were injected i.p. either with vehicle control (black bar) or with 1.5 mg PIK-75 (pattern) 3 hours prior to imaging. Tumors were exposed by skin-flap surgery and time-lapse z-series were acquired. The average total cell motility was determined per 40 μm z-stack (5 sections imaged at 10 μm intervals). Top: Data are means and SEM of 8 (vehicle control) and 15 (PIK-75) separate z stacks from 3 mice per condition. *: p< 0.016 Bottom : Representative panels from tumors of animals injected with (i) MTLn3-ErbB3--GFP vehicle control or (ii) PIK-75 3 hours prior to imaging. Cells are green with collagen fibers detected by second harmonic scattering in purple. Frames are 6 minutes apart, arrow shows migrating tumor cell; * indicates a single cell. Scale bar = 25 μm. E . Length/width ratio comparison. Top : representative images Bottom: Mean and SEM of 40-50 cells from 3 tumors per condition, 5 slices analyzed per field. Scale bar = 25 μm.*: p<0.002

Article Snippet: HRGβ1 was from R & D Systems (396-HB).

Techniques: In Vitro, Chemotaxis Assay, Migration, In Vivo, Injection, Control, Imaging, Comparison

Journal: Oncogene

Article Title: Phosphoinositide 3-kinase signaling is critical for ErbB3-driven breast cancer cell motility and metastasis

doi: 10.1038/onc.2011.275

Figure Lengend Snippet: A. Erk phosphorylation was determined in serum-starved cells after 5 minute treatment with 0 (-) or 0.4 nM HRGβ1 (+). Similar results were observed in three independent experiments. B. MTLn3 pLXSN, ErbB3WT, and ErbB3-Mutant cells were stimulated with 0.4 nM HRGβ1for 0, 5, 10, and 15 minutes (Min HRG). A delay in phosphorylation of Akt on Thr 308 was observed in ErbB3-Mutant cells response to HRGβ1. C. Akt (T308) and Erk phosphorylation activation was determined in serum-starved cells after 5 min treatment with 0 (-) or 5 nM (+) EGF. Similar results were observed in three independent experiments. D . Akt (T308) and Erk phosphorylation activation was determined in serum-starved cells after 10 min treatment with PDGF, IGF-I, and EGF. Similar results were observed in three independent experiments. E. Matrigel-coated transwells were used to test 12.5 nM HRGβ1induced in vitro invasion responses of MTLn3 ErbB3WT in the presence of vehicle (gray bars) or inhibitor (patterned bars). PIK-75 is a p110α selective inhibitor, Triciribine is an Akt inhibitor, and NSC23766 is a Rac1 inhibitor. Data are mean and SEM from 3 independent experiments. *: p< 0.05, **: p< 1.83E-05

Article Snippet: HRGβ1 was from R & D Systems (396-HB).

Techniques: Phospho-proteomics, Mutagenesis, Activation Assay, In Vitro