Differential contributions of nuclear lamina association and genome compartmentalization to gene regulation.

Chromatin regions that interact with the nuclear lamina are often heterochromatic, repressed in gene expression, and in the spatial B compartment. However, exceptions to this trend allow us to examine the relative impact of lamin association and spatial compartment on gene regulation. Here, we compared lamin association, gene expression, Hi-C, and histone mark datasets from cell lines representing different states of differentiation across different cell-type lineages. With these data, we compare, for example, gene expression differences when a B compartment region is associated with the nuclear lamina in one cell type but not in another. In general, we observed an additive rather than redundant effect of lamin association and compartment status. But, whether compartment status or lamin association had a dominant influence on gene expression varied by cell type. Finally, we identified how compartment and lamin association influence the likelihood of gene induction or repression in response to physicochemical treatment.

Recruiting women faculty through inclusive, family-friendly practices.

Recruitment of STEM faculty is biased against parents and caregivers. Specifically, women experience discrimination associated with childrearing and marriage. Underestimating the value of these candidates leads to a tremendous loss of talent. Here, we present a toolkit to facilitate the recruitment of talented women caregivers by providing guidelines for hiring.

Transcriptional profiling of Hutchinson-Gilford Progeria syndrome fibroblasts reveals deficits in mesenchymal stem cell commitment to differentiation related to early events in endochondral ossification.

The expression of a mutant Lamin A, progerin, in Hutchinson-Gilford Progeria Syndrome leads to alterations in genome architecture, nuclear morphology, epigenetic states, and altered phenotypes in all cells of the mesenchymal lineage. Here, we report a comprehensive analysis of the transcriptional status of patient derived HGPS fibroblasts, including nine cell lines not previously reported, in comparison with age-matched controls, adults, and old adults. We find that Progeria fibroblasts carry abnormal transcriptional signatures, centering around several functional hubs: DNA maintenance and epigenetics, bone development and homeostasis, blood vessel maturation and development, fat deposition and lipid management, and processes related to muscle growth. Stratification of patients by age revealed misregulated expression of genes related to endochondral ossification and chondrogenic commitment in children aged 4-7 years old, where this differentiation program starts in earnest. Hi-C measurements on patient fibroblasts show weakening of genome compartmentalization strength but increases in TAD strength. While the majority of gene misregulation occurs in regions which do not change spatial chromosome organization, some expression changes in key mesenchymal lineage genes coincide with lamin associated domain misregulation and shifts in genome compartmentalization.

SnapShot: Chromosome organization.

Chromosomes in higher eukaryotes are folded at different length scales into loop extrusion domains, spatial compartments, and chromosome territories and exhibit interactions with nuclear structures such as the lamina. Microscopic methods can probe this structure by measuring positions of chromosomes in the nuclear space in individual cells, while sequencing-based contact capture approaches can report the frequency of contacts of different regions within these structural layers. To view this SnapShot, open or download the PDF.

Chromosome compartmentalization alterations in prostate cancer cell lines model disease progression.

Prostate cancer aggressiveness and metastatic potential are influenced by gene expression and genomic aberrations, features that can be influenced by the 3D structure of chromosomes inside the nucleus. Using chromosome conformation capture (Hi-C), we conducted a systematic genome architecture comparison on a cohort of cell lines that model prostate cancer progression, from normal epithelium to bone metastasis. We describe spatial compartment identity (A-open versus B-closed) changes with progression in these cell lines and their relation to gene expression changes in both cell lines and patient samples. In particular, 48 gene clusters switch from the B to the A compartment, including androgen receptor, WNT5A, and CDK14. These switches are accompanied by changes in the structure, size, and boundaries of topologically associating domains (TADs). Further, compartment changes in chromosome 21 are exacerbated with progression and may explain, in part, the genesis of the TMPRSS2-ERG translocation. These results suggest that discrete 3D genome structure changes play a deleterious role in prostate cancer progression. .

Overstretched and overlooked: solving challenges faced by early-career investigators after the pandemic.

The coronavirus disease 2019 (COVID-19) pandemic has had a detrimental effect on research. However, little has been done to identify and solve the unique challenges faced by early career investigators (ECIs). As a group of American Cancer Society-funded ECIs, we provide recommendations for solving these challenges in the aftermath of the pandemic.

Radiation-induced DNA damage and repair effects on 3D genome organization.

The three-dimensional structure of chromosomes plays an important role in gene expression regulation and also influences the repair of radiation-induced DNA damage. Genomic aberrations that disrupt chromosome spatial domains can lead to diseases including cancer, but how the 3D genome structure responds to DNA damage is poorly understood. Here, we investigate the impact of DNA damage response and repair on 3D genome folding using Hi-C experiments on wild type cells and ataxia telangiectasia mutated (ATM) patient cells. We irradiate fibroblasts, lymphoblasts, and ATM-deficient fibroblasts with 5 Gy X-rays and perform Hi-C at 30 minutes, 24 hours, or 5 days after irradiation. We observe that 3D genome changes after irradiation are cell type-specific, with lymphoblastoid cells generally showing more contact changes than irradiated fibroblasts. However, all tested repair-proficient cell types exhibit an increased segregation of topologically associating domains (TADs). This TAD boundary strengthening after irradiation is not observed in ATM deficient fibroblasts and may indicate the presence of a mechanism to protect 3D genome structure integrity during DNA damage repair.

UDP-glucose 6-dehydrogenase regulates hyaluronic acid production and promotes breast cancer progression.

An improved understanding of the biochemical alterations that accompany tumor progression and metastasis is necessary to inform the next generation of diagnostic tools and targeted therapies. Metabolic reprogramming is known to occur during the epithelial-mesenchymal transition (EMT), a process that promotes metastasis. Here, we identify metabolic enzymes involved in extracellular matrix remodeling that are upregulated during EMT and are highly expressed in patients with aggressive mesenchymal-like breast cancer. Activation of EMT significantly increases production of hyaluronic acid, which is enabled by the reprogramming of glucose metabolism. Using genetic and pharmacological approaches, we show that depletion of the hyaluronic acid precursor UDP-glucuronic acid is sufficient to inhibit several mesenchymal-like properties including cellular invasion and colony formation in vitro, as well as tumor growth and metastasis in vivo. We found that depletion of UDP-glucuronic acid altered the expression of PPAR-gamma target genes and increased PPAR-gamma DNA-binding activity. Taken together, our findings indicate that the disruption of EMT-induced metabolic reprogramming affects hyaluronic acid production, as well as associated extracellular matrix remodeling and represents pharmacologically actionable target for the inhibition of aggressive mesenchymal-like breast cancer progression.

Tenascin-C and Integrin α9 Mediate Interactions of Prostate Cancer with the Bone Microenvironment.

Deposition of the extracellular matrix protein tenascin-C is part of the reactive stroma response, which has a critical role in prostate cancer progression. Here, we report that tenascin C is expressed in the bone endosteum and is associated with formation of prostate bone metastases. Metastatic cells cultured on osteo-mimetic surfaces coated with tenascin C exhibited enhanced adhesion and colony formation as mediated by integrin α9ß1. In addition, metastatic cells preferentially migrated and colonized tenascin-C-coated trabecular bone xenografts in a novel system that employed chorioallantoic membranes of fertilized chicken eggs as host. Overall, our studies deepen knowledge about reactive stroma responses in the bone endosteum that accompany prostate cancer metastasis to trabecular bone, with potential implications to therapeutically target this process in patients. Cancer Res; 77(21); 5977-88. ©2017 AACR.

Reactive stroma in the prostate during late life: The role of microvasculature and antiangiogenic therapy influences.

BACKGROUND: Prostate cancer is associated to a reactive stroma microenvironment characterized by angiogenic processes that are favorable for tumor progression. Senescence has been identified as a predisposing factor for prostate malignancies. In turn, the relationships between aging, reactive stroma, and the mechanisms that induce this phenotype are largely unknown. Thus, we investigated the occurrence of reactive stroma in the mouse prostate during advanced age as well as the effects of antiangiogenic and androgen ablation therapies on reactive stroma recruitment. METHODS: Male mice (52-week-old FVB) were treated with two classes of angiogenesis inhibitors: direct (TNP-470; 15 mg/kg; s.c.) and/or indirect (SU5416; 6 mg/kg; i.p.). Androgen ablation was carried out by finasteride administration (20 mg/kg; s.c.), alone or in association to both inhibitors. The Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) model was used as a paradigm of cancer-associated reactive stroma. The dorsolateral prostate was collected for α-actin (αSMA), vimentin (VIM), and transforming growth factor-beta (TGF-ß) immunohistochemical and Western blotting analyses as well as for CD34/αSMA and CD34/VIM colocalization. RESULTS: Senescence was associated with increased αSMA, VIM, and TGF-ß expression as well as with the recruitment of CD34/αSMA and CD34/VIM dual-positive fibroblasts. These observations were similar to those verified in TRAMP mice. Antiangiogenic treatment promoted the recovery of senescence-associated stromal changes. Hormonal ablation, despite having led to impaired CD34/αSMA and CD34/VIM dual-positive cell recruitment, did not result in decreased stimulus to reactive stroma development, due to enhanced TGF-ß expression in relation to the aged controls. CONCLUSIONS: Reactive stroma develops in the prostate of non-transgenic mice as a result of aging. The periacinar microvasculature is a candidate source for the recruitment of reactive stroma-associated cells, which may be derived either from perivascular-resident mesenchymal stem cells (MSCs) or from an endothelial-to-mesenchymal transition (EndMT) process. Thus, antiangiogenic therapy is a promising approach for preventing age-associated prostate malignancies by means of its negative interference in the development of reactive stroma phenotype from the vascular wall.

RUNX1 is essential for mesenchymal stem cell proliferation and myofibroblast differentiation.

Myofibroblasts are a key cell type in wound repair, cardiovascular disease, and fibrosis and in the tumor-promoting microenvironment. The high accumulation of myofibroblasts in reactive stroma is predictive of the rate of cancer progression in many different tumors, yet the cell types of origin and the mechanisms that regulate proliferation and differentiation are unknown. We report here, for the first time to our knowledge, the characterization of normal human prostate-derived mesenchymal stem cells (MSCs) and the TGF-ß1-regulated pathways that modulate MSC proliferation and myofibroblast differentiation. Human prostate MSCs combined with prostate cancer cells expressing TGF-ß1 resulted in commitment to myofibroblasts. TGF-ß1-regulated runt-related transcription factor 1 (RUNX1) was required for cell cycle progression and proliferation of progenitors. RUNX1 also inhibited, yet did not block, differentiation. Knockdown of RUNX1 in prostate or bone marrow-derived MSCs resulted in cell cycle arrest, attenuated proliferation, and constitutive differentiation to myofibroblasts. These data show that RUNX1 is a key transcription factor for MSC proliferation and cell fate commitment in myofibroblast differentiation. This work also shows that the normal human prostate gland contains tissue-derived MSCs that exhibit multilineage differentiation similar to bone marrow-derived MSCs. Targeting RUNX1 pathways may represent a therapeutic approach to affect myofibroblast proliferation and biology in multiple disease states.

Recruitment of CD34(+) fibroblasts in tumor-associated reactive stroma: the reactive microvasculature hypothesis.

Reactive stroma co-evolves with cancer, exhibiting tumor-promoting properties. It is also evident at sites of wound repair and fibrosis, playing a key role in tissue homeostasis. The specific cell types of origin and the spatial/temporal patterns of reactive stroma initiation are poorly understood. In this study, we evaluated human tumor tissue arrays by using multiple labeled, quantitative, spectral deconvolution microscopy. We report here a novel CD34/vimentin dual-positive reactive fibroblast that is observed in the cancer microenvironment of human breast, colon, lung, pancreas, thyroid, prostate, and astrocytoma. Recruitment of these cells occurred in xenograft tumors and Matrigel plugs in vivo and was also observed in stromal nodules associated with human benign prostatic hyperplasia. Because spatial and temporal data suggested the microvasculature as a common site of origin for these cells, we analyzed microvasculature fragments in organ culture. Interestingly, fibroblasts with identical phenotypic properties and markers expanded radially from microvasculature explants. We propose the concept of reactive microvasculature for the evolution of reactive stroma at sites of epithelial disruption common in both benign and malignant disorders. Data suggest that the reactive stroma response is conserved among tissues, in normal repair, and in different human cancers. A more clear understanding of the nature and origin of reactive stroma is needed to identify novel therapeutic targets in cancer and fibrosis.

Detección y tipificación del virus papiloma humano en lesiones laríngeas con técnica de hibridación molecular in situ / Detection and typification of human papilloma virus in laryngeal lesions with the technique of molecular hybridization in situ

Se ensayó la técnica de hibridación molecular in situ en lesiones papilomatosas de la laringe, con el objetivo de detectar material genético de virus papiloma humano (VPH), identificar los tipos virales involucrados en la infección, establecer relación de diagnóstico histológico con el virológico y verificar si, a determinados cambios histopatológicos, se asocian genotipos específicos. En ocho pacientes adultos portadores de papilomatosis laríngea se revisaron, retrospectivamente, aspectos clínicos y del tejido de biopsia. Se analizaron histopatológicamente las muestras, verificando la existencia de displasia y actividad viral, y se sometieron los tejidos al estudio de hibridación molecular de ácidos nucleicos in situ. Estos aspectos se realizaron independientemente siendo por tanto un estudio doble ciego. Detectamos genotipos virales en 7 de los 8 pacientes. Tres de ellos fueron positivos para los tipos 6-11 y se asociaron a grados de displasia leves y signos de actividad viral evidente. Cuatro pacientes fueron positivos para los tipos 16-18 y presentaron grados de displasia moderada y severa con pocos signos de actividad viral. Los resultados inducen a pensar que los genotipos 6-11 se asocian a lesiones de pronóstico benigno y los 16-18 a lesiones que podrían resultar malignas debiendo considerarse la infecciones por VPH en la laringe, especialmente las producidas por subtipos 16-18, como de riesgo, dado que estas serian cofactores carcinogenéticos

Resultados de la detección de anticuerpos anticocleares en diversas patologías del oído / Results of anticochlear antibodies detection in some ear diseases

Se presentan los resultados de aplicar un método de detección de anticuerpos anticocleares a 212 persona, 186 de ellas portadoras de hipoacusias atribuída a diversos cuadros clínicos y 26 normales. Se empleó la técnica de inmunofluorescencia indirecta, utilizando como antígeno material membranosoobtenido del conducto coclear humano de cadáveres frescos con menos de 6 horas de deceso. En 31 hipoacusias sensorioneurales, de las cuales 24 tenían un inicio y curso clínico que indicaban una posible etiología autoinmune y 7 que se presentaban asociadas a un cuadro sistémico de reconocida etiología autoinmune, la positividad en la detección de anticuerpos anticocleares se encontró en 17 de ellas (54.8 por ciento ). En 25 hipoacusias catalogadas como hidropesías endolinfáticas retardadas sean ipsi como contralaterales, la prueba fue positiva en 13 (52 por ciento ). En estos dos grupos los porcentajes son significativamente semejantes lo que sugiere una etipatogenia similar (p<0.01). En 47 hidropesias endolinfáticas idiopáticas o enfermedad de Meniere la prueba fue positiva en 12 (25.5 por ciento ). En 26 pacientes que presentaron una sordera súbita sensorioneural la prueba fue positiva en 8 (32 por ciento ). Otros 42 pacientes portadores de una hipoacusia sensorioneural no clasificable tuvieron una prueba positiva en 8 de ellos (19 por ciento ). La prueba fue también positiva en 2 de 8 hipoacusias consideradas congénitas y en 2 de 4 otoesclerosis. Cuatro hipoacusias consideradas como presbiacusia tuvieron la prueba negativa. Ninguno de los 26 sujetos sanos presentó la prueba positiva. Esto significa que la prueba de detección de anticuerpos anticocleares tiene una sensibilidad de 54.8 por ciento , una especificidad de 100 por ciento , un valor predictivo positivo de 100 por ciento y un valor predictivo negativo de 65 por ciento . Se concluye que no sólo la denominada enfermedad autoinmune del oído interno sería atribuíble a un fenómeno de autoinmunidad, sino que otros cuadros clínicos podrían tener una etiopatogenia similar

Técnica inmunohistoquímica en hueso temporal en enfermedad autoinmune de oido interno / Immunohistochemical technic in temporal bone in autoimmune disease of the inner ear

Diecisiete sueros de pacientes portadores de enfermedades del oído interno que ya habían sido sometidos a la búsqueda de anticuerpos anticocleares mediante la técnica de inmunofluorescencia indirecta, fueron estudiados mediante una técnica inmunohistoquímica usando peroxidasa de rábano picante. El objetivo fue el de precisar el lugar en el que el fenómeno antígeno anticuerpo tuvo lugar. Así mismo, se quiso determinar la especificidad y sensibilidad del nuevo método. Los resultados positivos demostraron que los complejos marcados con inmunoperoxidasa tuvieron afinidad por aquellas estructuras que tienen relación con el territorio arteriolar de la coclea, en especial con la estría vascular y la membrana bacilar. Comparado con la técnica de inmunofluorescencia indirecta se demostró que la especificidad se mantuvo, pero que la sensibilidad disminuyó. Esto último se atribuyó a que el procedimiento de fijación y descalcificación del hueso temporal obtenido, y que se usó como antígeno, pudo alterar la antigenicidad del mismo