Journal: International journal of oncology

Article Title: Inhibition of multiple myeloma‑derived exosomes uptake suppresses the functional response in bone marrow stromal cell.

doi: 10.3892/ijo.2019.4685

Figure Lengend Snippet: Figure 5. Inhibitors of macropinocytosis and membrane fusion partially decrease the uptake of multiple myeloma cell‑derived exosomes. Human bone marrow stromal cells were pre‑treated with the indicated concentration of amiloride for 0.5 h and then incubated with 50 µg/ml DIO‑labeled H929 cell‑derived exosomes for another 4 h in the presence of the indicated inhibitors. (A) The uptake was visualized by confocal microscopy and (B) the mean or median fluorescence intensity was determined by flow cytometry. Human bone marrow stromal cells were pre‑treated with the indicated concentration of omeprazole for 0.5 h and then incubated with 50 µg/ml DIO‑labeled H929 cell‑derived exosomes for another 4 h in the presence of the indicated inhibitors. (C) The uptake was visualized by confocal microscopy and (D) the mean or median fluorescence intensity was determined by flow cytometry. **P<0.01 and ***P<0.001, compared with 0 mM amiloride/omeprazole.

Article Snippet: For inhibition experiments, heparin, wortmannin, dynasore, chlorpromazine, amiloride, and omeprazole were purchased from Selleck Chemicals (Houston, TX, USA).

Techniques: Membrane, Concentration Assay, Incubation, Confocal Microscopy, Fluorescence, Flow Cytometry

Journal: International journal of oncology

Article Title: Inhibition of multiple myeloma‑derived exosomes uptake suppresses the functional response in bone marrow stromal cell.

doi: 10.3892/ijo.2019.4685

Figure Lengend Snippet: Figure 6. Endocytosis inhibitors partially suppress the effects of multiple myeloma cell‑derived exosome on BMSCs. (A) Human BMSCs were incubated with heparin, dynasore, amiloride or omeprazole at the indicated final concentration for 4.5 h, and cell viability was examined. **P<0.01 and ***P<0.001, compared with the group without inhibitor treatment. (B) Human BMSCs were pre‑treated with heparin (10 µg/ml) or dynasore (50 µM) for 30 min and incubated with 100 µg/ml RPMI8226 or H929 cell‑derived exosomes for 6 h. Subsequently, total and phosphorylated STAT1, STAT3 and ERK1/2 were quantified via western blot analysis. The GAPDH protein was included as a loading control. The pixel density of phosphorylated and total STAT1, STAT3 and ERK1/2 was quantified. (C) Phosphorylated proteins were normalized to their total protein amounts, and (D) total amounts of proteins were normalized to GAPDH quantities. *P<0.05; **P<0.01; and ***P<0.001. BMSCs, bone marrow stromal cells; Exo, exosomes; p, phosphor; STAT, signal transducer and activator of transcription 1; ERK1/2, extracellular signal‑regulated kinase 1/2.

Article Snippet: For inhibition experiments, heparin, wortmannin, dynasore, chlorpromazine, amiloride, and omeprazole were purchased from Selleck Chemicals (Houston, TX, USA).

Techniques: Incubation, Concentration Assay, Western Blot, Control

Journal: Molecular Therapy. Methods & Clinical Development

Article Title: Intracranial delivery of AAV9 gene therapy partially prevents retinal degeneration and visual deficits in CLN6-Batten disease mice

doi: 10.1016/j.omtm.2020.12.014

Figure Lengend Snippet: i.c.v. delivery of scAAV9.CB.CLN6 expresses throughout the retinal layers, enabling the local translation of hCLN6 in Cln6 nclf mouse retinas (A and B) Endogenous mouse Cln6 ( mCln6 ) expression in a wild-type animal as detected by RNAscope (A) localized primarily to the inner and outer nuclear layers, showing a different expression pattern as compared to the human CLN6 ( hCLN6 ) transgene (B). (C) Retinal whole-mounts immunolabeled with anti-human CLN6 (red) antibodies detected hCLN6 primarily in the central retina. Quantification is also shown in (C). (D–F) AAV9-treated Cln6 nclf mice immunolabeled with anti-human CLN6 (red) and rhodopsin (photoreceptor outer segments, green; D), BRN3a (retinal ganglion cells, green; E), PKCa (rod bipolar cells, green; F), or anti-glutamine synthetase (Müller glia, green; G), show hCLN6 colocalization primarily in RGCs, and also in the INL, ONL, RPE, and choroid. This indicates scAAV9.CB.CLN6 preferentially targeted RGC, INL, ONL, and epithelial layers using this dosing strategy. n = 6/treatment for each time point, represented by equal numbers of males and females. Mean ± SEM, One-way ANOVA, Tukey post-hoc. *p<0.05, ****p<0.0001.

Article Snippet: Briefly, sections were washed with PBS, blocked in 2% horse serum and 0.3% Triton X-100, and incubated with the following primary antibodies overnight at room temperature: rhodopsin (1:1,000, MAB5356, Millipore), glutamine synthetase (1:1,000, cat. no. 302, Millipore), Brn-3a (1:250, cat. no. sc-31984, Santa Cruz), PKCa (1:250, cat. no. sc-208, Santa Cruz), and human CLN6 (1:250, gift from Stella Lee ).

Techniques: Expressing, RNAscope, Immunolabeling

Journal: Frontiers in Endocrinology

Article Title: Quantitative Ubiquitinomics Revealed Abnormal Ubiquitinated ATP7A Involved in Down-Regulation of ACTH in Silent Corticotroph Adenomas

doi: 10.3389/fendo.2022.863017

Figure Lengend Snippet: Function of ATP7A in AtT20 cells. (A) The intensity of ubiquitinated peptides of ATP7A in SCAs and FCAs. (B) Up-regulation of the ACTH level in AtT20 cells with the knockdown of ATP7A by ATP7A siRNA. (C) Up-regulation of the ACTH level in AtT20 cells treated with 20uM omeprazole. All assays were performed in triplicate. (D) The top 20 potential E3 ligases of ATP7A. *Compared to control, p < 0.05.

Article Snippet: AtT20 cells treated with omeprazole (HY-B0113A, MedChemExpress) and the ACTH levels were assessed by an enzyme-linked immunosorbent assay (ELISA) (SBJ-M0483, SBJbio) according to the manufacture’s protocol.

Techniques: Knockdown, Control

Journal: Journal of Immunotoxicology

Article Title: Nebivolol prevents indomethacin-induced gastric ulcer in rats

doi: 10.3109/1547691x.2016.1142488

Figure Lengend Snippet: Figure 5. Sections of gastric mucosa of omeprazole-pre-treated rats. (a) Section indicates intact gastric mucosa (Magnification ¼ 100). (b) Gastric mucosa like normal and gastric glands with slight dilatation (Magnification ¼ 400). H&E stain. Representative images are shown. All of these tissues (and others from the group) were used by the pathologist to generate ulcer scores for each rat and, subsequently, the net protective index value for each group.

Article Snippet: Omeprazole was purchased from Carbosynth LLC (San Diego, CA), while nebivolol was a gift from 4A Pharma for Pharmaceutical Industries (Cairo, Egypt).

Techniques: Staining