Restriction and characterization of human breast cancer using a three-dimensional embryonic stem cell model

Human breast cancer is currently the highest diagnosed form of cancer and the second leading cause of cancer-related deaths in American women. Triple negative breast cancer is of the basal subtype and displays the worst prognosis owing to its highly metastatic properties. Current treatments focused on eradicating breast tumors in lieu of or following local therapy include chemotherapy, hormonal therapy, and targeted therapy. Hormonal therapy is not an option for triple negative breast cancer as it does not contain hormone receptors and there are currently no approved biological targeted therapies. Chemotherapy has proven unsuccessful because triple negative breast cancer is highly drug resistant. Here we report that metastatic human breast cancer cells (BCCs) were converted to a less aggressive phenotype and overcame chemotherapeutic drug resistance following exposure to embryonic stem cells (ESCs) encapsulated in alginate microstrands. We also demonstrate that the 3D ESC model restores proper EGFR and canonical Wnt/β-catenin signaling pathway regulation in metastatic BCCs and can be applied to identify a biological treatment for triple negative breast cancer. This study establishes the feasibility of inhibiting highly aggressive human BCCs with 3D cultured ESCs as demonstrated through decreases in metastatic BCC proliferation, abnormal metabolism, migration, invasiveness, chemotherapeutic drug resistance, and survival in vitro. ESCs and BCCs display signaling pathway convergence, which is highly and precisely regulated in ESCs and dysregulated in BCCs. Gene expression at the mRNA and protein level within restricted human BCCs indicates inhibition of the oncogenic EGFR and canonical Wnt/β-catenin signaling pathways. Naked cuticle 2 (NKD2) is a potential point of cross-talk between these two pathways and its increased expression suggests a role in restored regulation of these pathways. Application of this model for mechanistic studies will enable development of a targeted treatment for triple negative human breast cancer

A Case Study of the Eviction of a Female Bobcat and Her Four Kittens from a Suburban Backyard in Fremont, California

Since 2000, the Alameda County Vector Control Services District has received only 10 requests for service regarding bobcats. Seven of those cases have occurred in the past three years. Of those, only one case has involved the bobcats creating a den on a residential property. This is the case study of a mother bobcat and four kittens denning in the backyard of a suburban home in Fremont, California. The eviction of the bobcats from the deck of the vacant home was completed in May and June of 2019. During a four-week period, a wide range of eviction techniques were used, including noise, light, and water harassment, along with eviction fluid and human presence. This experience gave us the opportunity to observe and record the behaviors of bobcats living in a suburban environment and to test various eviction methods that had previously been untested in Alameda County. After four weeks of continuous humane harassment, the eviction was successful

A Case Study of the Eviction of a Female Bobcat and Her Four Kittens from a Suburban Backyard in Fremont, California

Since 2000, the Alameda County Vector Control Services District has received only 10 requests for service regarding bobcats. Seven of those cases have occurred in the past three years. Of those, only one case has involved the bobcats creating a den on a residential property. This is the case study of a mother bobcat and four kittens denning in the backyard of a suburban home in Fremont, California. The eviction of the bobcats from the deck of the vacant home was completed in May and June of 2019. During a four-week period, a wide range of eviction techniques were used, including noise, light, and water harassment, along with eviction fluid and human presence. This experience gave us the opportunity to observe and record the behaviors of bobcats living in a suburban environment and to test various eviction methods that had previously been untested in Alameda County. After four weeks of continuous humane harassment, the eviction was successful

Bioengineering Embryonic Stem Cell Microenvironments for the Study of Breast Cancer

Breast cancer is the most prevalent disease amongst women worldwide and metastasis is the main cause of death due to breast cancer. Metastatic breast cancer cells and embryonic stem (ES) cells display similar characteristics. However, unlike metastatic breast cancer cells, ES cells are nonmalignant. Furthermore, embryonic microenvironments have the potential to convert metastatic breast cancer cells into a less invasive phenotype. The creation of in vitro embryonic microenvironments will enable better understanding of ES cell-breast cancer cell interactions, help elucidate tumorigenesis, and lead to the restriction of breast cancer metastasis. In this article, we will present the characteristics of breast cancer cells and ES cells as well as their microenvironments, importance of embryonic microenvironments in inhibiting tumorigenesis, convergence of tumorigenic and embryonic signaling pathways, and state of the art in bioengineering embryonic microenvironments for breast cancer research. Additionally, the potential application of bioengineered embryonic microenvironments for the prevention and treatment of invasive breast cancer will be discussed

Bioengineering Embryonic Stem Cell Microenvironments for the Study of Breast Cancer

Breast cancer is the most prevalent disease amongst women worldwide and metastasis is the main cause of death due to breast cancer. Metastatic breast cancer cells and embryonic stem (ES) cells display similar characteristics. However, unlike metastatic breast cancer cells, ES cells are nonmalignant. Furthermore, embryonic microenvironments have the potential to convert metastatic breast cancer cells into a less invasive phenotype. The creation of in vitro embryonic microenvironments will enable better understanding of ES cell-breast cancer cell interactions, help elucidate tumorigenesis, and lead to the restriction of breast cancer metastasis. In this article, we will present the characteristics of breast cancer cells and ES cells as well as their microenvironments, importance of embryonic microenvironments in inhibiting tumorigenesis, convergence of tumorigenic and embryonic signaling pathways, and state of the art in bioengineering embryonic microenvironments for breast cancer research. Additionally, the potential application of bioengineered embryonic microenvironments for the prevention and treatment of invasive breast cancer will be discussed

Understanding convergent signaling regulation in metastatic breast cancer cells using a bioengineered stem cell microenvironment

Aim: The convergence of tumorigenic and embryonic signaling pathways drives us to exploit the embryonic stem cell (ESC) microenvironment to restrict metastatic potential of cancer cells. We have previously demonstrated that bioengineered ESC microenvironments could restrict growth and metastatic potential of highly aggressive breast cancer cell (BCC). This study aims to further understand the regulation of convergent EGFR and canonical Wnt/β-catenin signaling pathway function in triple negative metastatic BCCs using the 3D in vitro ESC microenvironment created by encapsulating ESCs in alginate hydrogel microstrands.Methods: Co-culture with ESC-microstrands increased sensitivity to two chemotherapeutic drugs in metastatic BCCs. To test whether these changes were due to restored signaling pathway regulation in BCCs, we probed for changes in gene expression of key molecules related to the EGFR and canonical Wnt/β-catenin signaling pathways using quantitative reverse transcription polymerase chain reaction and Western blot analysis.Results: ESC-microstrands are able to alter the gene expression of highly aggressive BCCs at both mRNA and protein levels. These changes are indicative of a reversal of EGFR and canonical Wnt/β-catenin signaling pathway hyperactivation following co-culture. Increased NKD2 mRNA and protein expression coinciding with dual signaling pathway inhibition within co-cultured BCCs suggests that this reversal may be attributable to restored regulation of NKD2 within these pathways.Conclusion: ESC-microstrands are able to reverse oncogenic signaling pathway hyperactivation and restore signaling pathway regulation in metastatic BCCs. Further studies could provide insight into what role NKD2 up-regulation plays in BCC inhibition, leading to the development of a new targeted therapy for metastatic breast cancer

Restriction of Cancer Metastatic Potential Using Embryonic Stem Cells Encapsulated in Alginate Hydrogel Microstrands

Current treatments focused on eradicating metastatic tumors have proven unsuccessful due to cancer’s ability to quickly undergo epithelial-to-mesenchymal transition (EMT) and metastasize to secondary sites. Using human triple negative breast cancer cells (BCCs) as a model system, this work establishes a platform for the study of aggressive cancer phenotypes by demonstrating the inhibition of human metastatic cancer cells with 3D cultured embryonic stem cells (ESCs) encapsulated in alginate microstrands (ESC-microstrands), which mimic the embryonic microenvironment and recapitulate pluripotent signaling. Coculture with ESC-microstrands significantly decreases triple negative BCC proliferation and survival and reverses abnormal cancer metabolism. In particular, coculture with ESC-microstrands markedly restricts the metastatic potential of highly aggressive cancer cells, demonstrated as decreased migration and invasion, and reversed EMT marker expression. This indicates that pluripotent signaling from 3D ESC-microstrands could restrict cancer metastasis through restriction and reversion of EMT. Furthermore, two soluble factors associated with dysregulated oncogenic signaling were identified which display altered relative mRNA expression following coculture with ESC-microstrands. Future application of this model to mechanistic studies will enable a better understanding of cancer metastasis and the discovery of therapeutic targets for metastatic diseases

Quality standards for the care of people with giant cell arteritis in secondary care

Giant cell arteritis (GCA) is the commonest primary systemic vasculitis in adults. It has significant health economic costs and societal burden (1, 2), which is likely to get worse with an aging population. British and European recommendations endorse early specialist review (3, 4). In England, 49% of centres provide a diagnostic ultrasonography service but there is wide variation in access and speed of delivery (5). 34% of hospitals in England did not have any formal clinical pathway for assessing GCA (5). Primary care physicians require pathways (6), and the experience of secondary care physicians suggests that establishing a robust one is difficult (5). Treatment recommendations provide an impetus for improvement in standards of care. Those with auditable metrics provide an even greater driver for change. For example, adoption of national standards for the treatment of early inflammatory arthritis in the United Kingdom has proven to be a significant catalyst for improvement in care (7). We have formed a multidisciplinary group aiming to create standards to bring about similar nationwide improvement in the care of GCA

A Genome-Scale RNA–Interference Screen Identifies RRAS Signaling as a Pathologic Feature of Huntington's Disease

<div><p>A genome-scale RNAi screen was performed in a mammalian cell-based assay to identify modifiers of mutant huntingtin toxicity. Ontology analysis of suppressor data identified processes previously implicated in Huntington's disease, including proteolysis, glutamate excitotoxicity, and mitochondrial dysfunction. In addition to established mechanisms, the screen identified multiple components of the RRAS signaling pathway as loss-of-function suppressors of mutant huntingtin toxicity in human and mouse cell models. Loss-of-function in orthologous RRAS pathway members also suppressed motor dysfunction in a Drosophila model of Huntington's disease. Abnormal activation of RRAS and a down-stream effector, RAF1, was observed in cellular models and a mouse model of Huntington's disease. We also observe co-localization of RRAS and mutant huntingtin in cells and in mouse striatum, suggesting that activation of R-Ras may occur through protein interaction. These data indicate that mutant huntingtin exerts a pathogenic effect on this pathway that can be corrected at multiple intervention points including RRAS, FNTA/B, PIN1, and PLK1. Consistent with these results, chemical inhibition of farnesyltransferase can also suppress mutant huntingtin toxicity. These data suggest that pharmacological inhibition of RRAS signaling may confer therapeutic benefit in Huntington's disease.</p> </div

Ingenuity Pathway Analysis (IPA) of HD Suppressor Genes.

<p>(A) IPA network of the HD suppressor genes that could be directly connected to each other without intervening nodes. This network was constructed using data from all Ingenuity model organisms. Huntingtin (HTT) was manually added to this network, and its connections colored red. Functions of nodes are indicated with icons. “Direct Relationship” (solid lines) indicates direct physical contact between two molecules, <i>e.g.</i> binding or phosphorylation. “Indirect Relationship” (dotted lines) indicates a functional interaction that does not require physical contact between the two molecules, <i>e.g.</i> signaling events. See also <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003042#pgen.1003042.s003" target="_blank">Figure S3</a>.</p