Journal: Frontiers in Oncology

Article Title: Salubrinal Exposes Anticancer Properties in Inflammatory Breast Cancer Cells by Manipulating the Endoplasmic Reticulum Stress Pathway

doi: 10.3389/fonc.2021.654940

Figure Lengend Snippet: BiP/GRP78 is expressed abundantly in human IBC tissue sections. (A) Hematoxylin and Eosin (H & E) staining of healthy (left) (Aa) and IBC tissue (right) (Ab) is shown at 10X and 40X magnification. Healthy (Ac, Ad) and IBC tissue sections (Ae, Af) were stained using an antibody against BiP/GRP78. Human IBC tissue sections (Ag, Ah) were stained using an anti-IgG antibody. (Aa–Ac, Ae, Ag) are 10X, whereas (Ad, Af, Ah) represent 40X magnification. (B) BiP/GRP78 immunostaining (red) and healthy and IBC tissue are shown at 10X magnification. Tissue sections were developed with Alexa-594 coupled secondary antibody (red). Nuclei were visualized using DAPI as the counterstain (blue).

Article Snippet: Total RNA was isolated using TRIzol Reagent (#15596026, Life Technologies Corporation, Grand Island, NY) from IBC tissue samples (Biochain, breast tumor tissue array # T22350862-2) and treated with DNase I (#18068015, Life Technologies Corporation) at 37°C for 30 min for DNA removal.

Techniques: Staining, Immunostaining

Journal: Frontiers in Oncology

Article Title: Salubrinal Exposes Anticancer Properties in Inflammatory Breast Cancer Cells by Manipulating the Endoplasmic Reticulum Stress Pathway

doi: 10.3389/fonc.2021.654940

Figure Lengend Snippet: ER stress genes are abundantly expressed in IBC tissue sections and IBC cell lines. (A) RNA isolated from healthy and IBC tumor tissue was prepared, converted to cDNA, and gene expression was quantified by real-time RT-PCR using specific primers for ER stress markers as indicated. Each data point represents the average gene expression from six IBC and six healthy control samples. Each point represents the average ± the standard deviation of three experiments. (***) p<0.005, (****) p<0.001 indicates a statistically significant difference compared with healthy tissue. Each reaction was done in triplicate. (B) RNA isolated from HMEC, SUM149PT, and SUM190PT cells was converted to cDNA, and gene expression was quantified by real-time RT-PCR using specific primers for ATF4, CHOP, GADD34, GRP78, IRE1α, and XBP-1. Each point represents the average ± the standard deviation of three experiments. (**) p<0.01, (***) p<0.005, (****) p<0.001 indicate a statistically significant difference compared with HMEC cells. Each reaction was done in triplicate. (C) ER stress genes proteins are abundantly expressed in IBC cell lines. Lysates prepared from HMEC, SUM149PT, and SUM190PT cells, were tested for protein levels of PERK, IRE1α, calnexin, ERO1α, and PDI. Blots were reprobed with anti-β-actin antibody as a loading control for normalization. Fold expression of each protein was calculated by considering the expression of the protein in HMEC as 1. (D) Immunostaining of HMEC and SUM149PT cells seeded in eight-well chamber slides. Cells were fixed, permeabilized, and then stained with primary monoclonal antibodies against ER stress markers, including calnexin, ERO1α, and IRE1α. Cells were developed with Alexa-488 coupled secondary antibody (green). Nuclei were visualized using DAPI as the counterstain (blue).

Article Snippet: Total RNA was isolated using TRIzol Reagent (#15596026, Life Technologies Corporation, Grand Island, NY) from IBC tissue samples (Biochain, breast tumor tissue array # T22350862-2) and treated with DNase I (#18068015, Life Technologies Corporation) at 37°C for 30 min for DNA removal.

Techniques: Isolation, Expressing, Quantitative RT-PCR, Standard Deviation, Immunostaining, Staining

Journal: Biomedicines

Article Title: Potent Small-Molecule Inhibitors Targeting Acetylated Microtubules as Anticancer Agents Against Triple-Negative Breast Cancer

doi: 10.3390/biomedicines8090338

Figure Lengend Snippet: Inhibition of triple-negative breast cancer (TNBC) progression by blocking microtubule acetylation. ( A ) Representative images showing acetyl-α-tubulin expression in various subtypes of human breast cancer. Scale bar, 500 µm. The intensity was analyzed using Aperio Image Scope software. ( B ) Expression of microtubule acetylation in breast cancer cell lines (upper panel) categorized according to the subtype (lower panel). ( C ) Confirmation of acetyl-α-tubulin expression in alpha-tubulin acetyltransferase 1 (αTAT1) Knockout (KO) MDA-MB-231 cell lysates. ( D ) Anchorage-independent growth assays using αTAT1 KO MDA-MB-231 cells for 3 weeks. The number of colonies was quantified using ImageJ software. Scale bar, 1 mm. ***, p ≤ 0.001. ( E ) Mock and αTAT1 KO MDA-MB-231 cells were injected into the mammary fat pad of non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice ( n = 5, each). *, p ≤ 0.05; ***, p ≤ 0.001.

Article Snippet: Paraffin-embedded human breast cancer tissue samples and patient pathological data were obtained from US Biomax, Inc., Derwood, MD, USA (BR10011).

Techniques: Inhibition, Blocking Assay, Expressing, Software, Knock-Out, Injection