Jkl activation cancer

Source: Kubler K, et al. Disclosures: Susan F. Jeselsohn reports research funding from Eli Lilly and Pfizer, as well as consultant roles with Carrick Therapeutics and Luminex Corporation. Read next. December 08, Receive an email when new articles are posted on. Please provide your email address to receive an email when new articles are posted on.

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Article menu. Recent advances in basic science. Targeting G protein-coupled receptors for the treatment of chronic pain in the digestive system. Abstract Chronic pain is a hallmark of functional disorders, inflammatory diseases and cancer of the digestive system. Statistics from Altmetric. No commercial re-use. See rights and permissions. Published by BMJ. Read the full text or download the PDF:.

Log in. The stress mediator p38 has been previously shown to phosphorylate EZH2 and inhibit its activity In support of the latter hypothesis, the mainly fat-containing mouse mammary fat pad is biologically less stiff microenvironment for epithelial glands than the fibroblast-enriched human breast stroma Supplementary Fig.

A metal grid was placed on top of the cultures and two magnets were placed on the opposite sides of the cultures to generate the compression Fig. Sequential sections were stained. Together, since phosphorylation of EZH2 at T suppresses the protein activity, our results from both mouse and human explant cultures altogether are consistent with a mechanistic model presented in Fig. Accordingly, specifically a stiff matrix induces p38 mediated stress pathway, which keeps EZH2 phosphorylated at T thus suppressing the activity of this key enzyme that catalyzes the addition of methyl groups to histone H3 at lysine The high MBD reflects a greater amount of glandular and connective tissue compared to the fat as well as enhanced tissue stiffness 45 , 46 , Furthermore, women with the highest MBD exhibit a four- to sixfold increase in breast cancer risk compared to women with nondense breasts 48 , 49 , Preoperative mammography is performed prior to noncosmetic reduction mammoplasty in Finland, and, therefore, each reduction mammoplasty RMP sample in our series could be annotated with a pre-existing clinical MBD score for the clinical data, IHC stainings, and scoring, see Fig.

The current results from ex vivo culture studies are summarized in Fig. We show that the epithelial cell identity is not a stable feature in a culture but highly sensitive to changes mediated by the matrix environment. In Matrigel, mammary epithelial tissue explants underwent a rapid phenotypic switch from the luminal to the basal cell identity. However, we observed species-specific differences; MMECs formed normal-like bilayered epithelial structures with the basal cells facing the matrix and the luminal cells forming the inner layer.

In contrast, the non-cancerous human mammary epithelial cells PDEC-N primarily assumed the basal phenotype schematic representation of the matrix effects in Fig. A similar phenotypic switch occurred in most, but not in all breast cancer explants PDEC-BC , perhaps implicating confounding genetic alterations that interfere with matrix-dependent plasticity.

Importantly, the originally luminal breast tissue or breast cancer samples retained the luminal phenotype in three bioinert LMx scaffolds. However, even if the cells retained luminal identity, different matrices had a strikingly different impact on the global regulation of genome expression.

Altogether, the soft gel promoted the intergenic and intron enriched global gene expression pattern that is characteristic for stem cells 28 , it enriched stem cell and pluripotency-related gene expression signatures and induced gene repression related H3K27me3 methylation profiles. Interestingly, a chemically induced stress by anisomycin phenocopied the effect of stiff microenvironment in the explant cultures.

However, p38 also regulates the relaxation of chromatin, the phosphorylation of histone H3, and chromatin demethylation via mechanisms that may not linearly exploit EZH2 53 , 54 , Curiously, the genetic profiles of the stiff LMx-Ag cultured human explants were similar to the mouse explants, which were cultured in the soft LMx-Al.

These findings together with the fact that human mammary epithelial cells naturally encounter higher mechanical stresses than mouse mammary epithelial cells Supplementary Fig. This difference may reflect the fact that during the one-week culture period cells soften the surrounding culture matrix through cell secreted proteases and other biomolecules. Hence, the initial stiffness needs to be high enough to provide mechanical stress to the cells until the end of the experiment.

However, the difference between in vivo and ex vivo requirements may also arise from other factors present in the complex breast tissue in vivo but lacking in the ex vivo cultures. It is well established by number of studies that tumors have a high interstitial pressure and they form a rigid mass representing a significantly stiffer tissue environment than the normal breast epithelium These compressive forces range around kPa.

The surprising cell plasticity and matrix-dependent programmability of differentiated mammary epithelial cells and breast cancer cells in authentic patient-derived tissue cultures open up new opportunities to understand breast cancer biology and to develop new breast cancer treatments.

Breast cancer cell lines were purchased from ATCC. Additionally, 0. Patients participated in the study by signing an informed consent form following the Declaration of Helsinki principles.

Tissues were collected from reduction mammoplasty samples, from tumors, and the adjacent normal-like areas of the tumors. Isolated explants were embedded in different 3D matrices and plated on 8-chamber slides Thermo Scientific. The mice used in the study were sacrificed with CO 2 followed by cervical dislocation. Agarose solutions were prepared by first dispersing 0. Alginate gels were prepared by dispersing 0. GrowDex was a pre-prepared commercial matrix.

Matrigel 8. All the components and equipment were pre-cooled and kept on ice during the preparation. First,10 x PBS and water were mixed. Then, a Collagen I rat tail stock solution was added. Egg whites were separated from the yolks and filtered through a sinter to keep only the clear part. Ovomucin gels were prepared according to an established protocol Following, the liquid was pipetted from the bottom of the tube below beads into a fresh 1.

Afterwards, the supernatant was collected into a fresh 1. After adding Lysis Buffer again, the same extraction was performed. The lysate from the second extraction was collected from the top of the tube above beads and collected into separate 1. The specimen for SEM imaging were prepared using different sample preparation methods to obtain aerogels. GrowDex and agarose samples were plunge-freezed in liquid propane and subsequently lyophilized.

The alginate sample was prepared using a supercritical carbon dioxide drying method. Matrigel, ovomucin, collagen and alginate-RGD samples were prepared using glutaraldehyde 3. Sample preparation methods, coatings, and acceleration voltages for each matrix appear in Supplementary Table 1a. We carried out oscillatory rheology using a TA Instrument AR stress-controlled rheometer with a Peltier heated plate.

The sealing lid and silicon oil prevented evaporation during the measurement. The gap temperature compensation parameter was 0. First, the linear viscoelastic region LVR was confirmed with a preliminary measurement including strain sweep. The parameters for the measurements appear in Supplementary Table 1b for the specific matrices.

Data were acquired in triplicate and reported as average unless otherwise stated. The tissue explants were permeabilized with 0. The list of used antibodies are shown in Supplementary Data 2. Slides containing tissue explants were mounted with the ImmuMount reagent Fisher Scientific. The heat-induced antigen retrieval was performed whether with a microwave oven or a pressure cooker in a citrate buffer solution Dako. Finally, the library was assessed with the Agilent Bioanalyzer.

The Gene Set Enrichment Analysis 3. GSEA results were visualized using Cytoscape v. DNA was extracted from the original tumors and the corresponding 3D cultured samples. The DNA integrity was confirmed using gel electrophoresis.

Cell The samples were incubated in T thermal cycler BioRad. The samples were thermocycled in a T thermocycler BioRad. The PCR products were pooled together in sets of 12 samples containing different Indexing Oligos and purified with 0. The purified cDNA was tagmented using the Nextera kit. Primer sequences are shown in the Supplementary Table 2.

The gene-specific primer sets were used at a final concentration of 0. Primer sequences for the human samples were taken from Kangaspeska et al. The primer sequences for mice samples are shown in Supplementary Table 2.

The explants were embedded within LMx-Ag. Following preparation, two magnets Magnet Expert Ltd; FSC were used to compress the cultures between the well-plate bottom and a metallic grid on the top of the cell culture. All the experiments with representative images Western blot, immunohistology, and immunofluorescence stainings have been repeated at least thrice. Statistical analyses were performed using Graphpad Prism 8 Version 8. Whole-cell extracts were isolated using a RIPA lysis buffer supplemented with protease Roche and phosphatase inhibitors Roche.

Further information on research design is available in the Nature Research Reporting Summary linked to this article. Due to the nature of the consent given by the patients, we are not allowed to share the exome sequencing data in any public data repositories. Source data are provided with this paper. Perou, C. Molecular portraits of human breast tumours. Nature , — Cheang, M. Ki67 index, HER2 status, and prognosis of patients with luminal B breast cancer. Cancer Inst. Dai, X.

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