Kinetic Evaluation of the Hypoxia Radiotracers [F]FMISO and [F]FAZA in Dogs with Spontaneous Tumors Using Dynamic PET/CT Imaging.

Purpose: We evaluated the kinetics of the hypoxia PET radiotracers, [18F]fluoromisonidazole ([18F]FMISO) and [18F]fluoroazomycin-arabinoside ([18F]FAZA), for tumor hypoxia detection and to assess the correlation of hypoxic kinetic parameters with static imaging measures in canine spontaneous tumors.

Methods: Sixteen dogs with spontaneous tumors underwent a 150-min dynamic PET scan using either [18F]FMISO or [18F]FAZA. The maximum tumor-to-muscle ratio (TMR) > 1.

Double blind trial of a deuterated form of linoleic acid (RT001) in Friedreich ataxia.

Objectives: Friedreich ataxia is (FRDA) an autosomal recessive neurodegenerative disorder associated with intrinsic oxidative damage, suggesting that decreasing lipid peroxidation (LPO) might ameliorate disease progression. The present study tested the ability of RT001, a deuterated form of linoleic acid (D2-LA), to alter disease severity in patients with FRDA in a double-blind placebo-controlled trial.

Methods: Sixty-five subjects were recruited across six sites and received either placebo or active drug for an 11-month study.

Correction: Preservation of myocardial contractility during acute hypoxia with OMX-CV, a novel oxygen delivery biotherapeutic.

[This corrects the article DOI: 10.1371/journal.pbio.

Preservation of myocardial contractility during acute hypoxia with OMX-CV, a novel oxygen delivery biotherapeutic.

The heart exhibits the highest basal oxygen (O2) consumption per tissue mass of any organ in the body and is uniquely dependent on aerobic metabolism to sustain contractile function. During acute hypoxic states, the body responds with a compensatory increase in cardiac output that further increases myocardial O2 demand, predisposing the heart to ischemic stress and myocardial dysfunction. Here, we test the utility of a novel engineered protein derived from the heme-based nitric oxide (NO)/oxygen (H-NOX) family of bacterial proteins as an O2 delivery biotherapeutic (Omniox-cardiovascular [OMX-CV]) for the hypoxic myocardium.

Human stem cells from single blastomeres reveal pathways of embryonic or trophoblast fate specification.

Mechanisms of initial cell fate decisions differ among species. To gain insights into lineage allocation in humans, we derived ten human embryonic stem cell lines (designated UCSFB1-10) from single blastomeres of four 8-cell embryos and one 12-cell embryo from a single couple. Compared with numerous conventional lines from blastocysts, they had unique gene expression and DNA methylation patterns that were, in part, indicative of trophoblast competence.

Human embryonic stem cells derived from embryos at different stages of development share similar transcription profiles.

We have derived hESC from biopsied blastomeres of cleavage stage embryos under virtually the same conditions we used for the derivation of hESC lines from inner cell mass of blastocyst stage embryos. Blastomere-derived hESC lines exhibited all the standard characteristics of hESC including undifferentiated proliferation, genomic stability, expression of pluripotency markers and the ability to differentiate into the cells of all three germ layers both in vitro and in vivo. To examine whether hESC lines derived from two developmental stages of the embryo differ in gene expression, we have subjected three blastomere-derived hESC lines and two ICM-derived hESC lines grown under identical culture conditions to transcriptome analysis using gene expression arrays.

GROα regulates human embryonic stem cell self-renewal or adoption of a neuronal fate.

Previously we reported that feeders formed from human placental fibroblasts (hPFs) support derivation and long-term self-renewal of human embryonic stem cells (hESCs) under serum-free conditions. Here, we show, using antibody array and ELISA platforms, that hPFs secrete ∼6-fold higher amounts of the CXC-type chemokine, GROα, than IMR 90, a human lung fibroblast line, which does not support hESC growth. Furthermore, immunocytochemistry and immunoblot approaches revealed that hESCs express CXCR, a GROα receptor.

Use of human embryonic stem cell-based models for male reproductive toxicity screening.

Male reproductive toxicity examines harmful effects of various agents on all aspects and developmental stages of the male reproductive system, including germ cell development and spermatogenesis. In developing a model for reproductive toxicity screening it is important to define the developmental stage that this model is going to recreate in vitro and to identify critical molecular targets of this stage. In this review we focus our discussion on the potential for using embryonic stem cell (ESC)-derived models for male reproductive toxicity screening.

A human-like senescence-associated secretory phenotype is conserved in mouse cells dependent on physiological oxygen.

Cellular senescence irreversibly arrests cell proliferation in response to oncogenic stimuli. Human cells develop a senescence-associated secretory phenotype (SASP), which increases the secretion of cytokines and other factors that alter the behavior of neighboring cells. We show here that "senescent" mouse fibroblasts, which arrested growth after repeated passage under standard culture conditions (20% oxygen), do not express a human-like SASP, and differ from similarly cultured human cells in other respects.

The senescence-associated secretory phenotype: the dark side of tumor suppression.

Cellular senescence is a tumor-suppressive mechanism that permanently arrests cells at risk for malignant transformation. However, accumulating evidence shows that senescent cells can have deleterious effects on the tissue microenvironment. The most significant of these effects is the acquisition of a senescence-associated secretory phenotype (SASP) that turns senescent fibroblasts into proinflammatory cells that have the ability to promote tumor progression.

Effect of karyotype on successful human embryonic stem cell derivation.

The success rate of human embryonic stem cell (hESC) derivation depends on both culture conditions and embryo quality and is routinely determined by morphological criteria. However, high incidence of chromosomal abnormality even in high-grade cleavage embryos from in vitro fertilization (IVF) patients suggests that the morphological grade of supernumerary embryos obtained from IVF clinics may not be a good prediction factor for successful hESC derivation. We show here that from one donor under identical derivation conditions 12 karyotypically abnormal post-bioptic embryos did not yield hESC lines, whereas two out of four normal embryos did.

Human embryonic stem cells as a model for embryotoxicity screening.

Reproductive toxicity encompasses harmful effects of various agents on all aspects and stages of the reproductive cycle, including infertility and the induction of adverse effects in the embryo/fetus. In developing a model for reproductive toxicity screening, it is important to define the stage of the human reproductive cycle that this specific model is going to recreate in vitro and to identify molecular targets that are critical for this stage of development. In this review, we focus our discussion on modeling pre-implantation embryotoxicity.

Derivation of human embryonic stem cell lines from biopsied blastomeres on human feeders with minimal exposure to xenomaterials.

In a continuous effort to improve the generation of therapeutic grade human embryonic stem cell (hESC) lines, we focused on preserving developmental capacity of the embryos, minimizing the exposure to xenomaterials, increasing derivation efficacy, and reducing the complexity of the derivation procedure. In this study, we describe an improved method for efficient derivation of hESC lines from blastomeres of biopsied embryos. Our protocol substituted feeder cells of mouse origin with human foreskin fibroblasts (HFFs), limited serum exposure of cells to formation of the initial outgrowth, and increased derivation efficacy from 12.

Derivation of hESC from intact blastocysts.

This unit describes protocols for culturing human embryos and deriving human embryonic stem cells from the intact blastocyst. Description of the culturing begins with methods for obtaining human blastocysts using pronuclear or cleavage stage embryos left over after in vitro fertilization. Then there is a description of methods that can be used to derive human embryonic stem cell lines from the blastocyst without trophectoderm removal.

A role for fibroblasts in mediating the effects of tobacco-induced epithelial cell growth and invasion.

Cigarette smoke and smokeless tobacco extracts contain multiple carcinogenic compounds, but little is known about the mechanisms by which tumors develop and progress upon chronic exposure to carcinogens such as those present in tobacco products. Here, we examine the effects of smokeless tobacco extracts on human oral fibroblasts. We show that smokeless tobacco extracts elevated the levels of intracellular reactive oxygen, oxidative DNA damage, and DNA double-strand breaks in a dose-dependent manner.

Efficient method for slow cryopreservation of human embryonic stem cells in xeno-free conditions.

An effective, consistent and xeno-free cryopreservation technique is crucial for any human embryonic stem cell (hESC) laboratory with future perspectives for clinical application. This study presents a new slow freezing-rapid thawing method in serum-free conditions that allows the cryopreservation of a large number of colonies without the use of a programmable freezer. To test its efficacy, this method has been compared with two established vitrification methods and applied to three different hESC lines (H9, VAL-3 and VAL-5).

Disruption of apical-basal polarity of human embryonic stem cells enhances hematoendothelial differentiation.

During murine development, the formation of tight junctions and acquisition of polarity are associated with allocation of the blastomeres on the outer surface of the embryo to the trophoblast lineage, whereas the absence of polarization directs cells to the inner cell mass. Here, we report the results of ultrastructural analyses that suggest a similar link between polarization and cell fate in human embryos. In contrast, the five human embryonic stem cell (hESC) lines displayed apical-basal, epithelial-type polarity with electron-dense tight junctions, apical microvilli, and asymmetric distribution of organelles.

Serum-free derivation of human embryonic stem cell lines on human placental fibroblast feeders.

Objective: To derive new human embryonic stem cell (hESC) lines on pathogen-free human placental fibroblast feeders under serum-free conditions. Because the embryo develops in close contact with extraembryonic membranes, we hypothesized that placental mesenchyme might replicate the stem cell niche in situ.

Design: We isolated and characterized human placental fibroblast lines from individual donors and tested their ability to support growth of federally registered hESC lines.

Stem cell: balancing aging and cancer.

Stem cells are defined by their self-renewing capacity and the ability to differentiate into one or more cell types. Stem cells can be divided, depending on their origin, into embryonic or adult. Embryonic stem cells derive from early stage embryos and can give rise to cells from all three germ layers.

Stromal-epithelial interactions in aging and cancer: senescent fibroblasts alter epithelial cell differentiation.

Cellular senescence suppresses cancer by arresting cells at risk of malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation.

First derivation in Spain of human embryonic stem cell lines: use of long-term cryopreserved embryos and animal-free conditions.

The first two human embryonic stem cell lines (VAL-1 and VAL-2) have been derived in Spain with long-term cryopreserved embryos under animal-free conditions. In the first series, 40 human embryos that had been cryopreserved at day 2 of development were thawed after >5 years. A derivation efficiency of 5% per frozen embryo or 12.

Reversal of human cellular senescence: roles of the p53 and p16 pathways.

Telomere erosion and subsequent dysfunction limits the proliferation of normal human cells by a process termed replicative senescence. Replicative senescence is thought to suppress tumorigenesis by establishing an essentially irreversible growth arrest that requires activities of the p53 and pRB tumor suppressor proteins. We show that, depending on expression of the pRB regulator p16, replicative senescence is not necessarily irreversible.

Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts.

Most mammalian cells do not divide indefinitely, owing to a process termed replicative senescence. In human cells, replicative senescence is caused by telomere shortening, but murine cells senesce despite having long stable telomeres. Here, we show that the phenotypes of senescent human fibroblasts and mouse embryonic fibroblasts (MEFs) differ under standard culture conditions, which include 20% oxygen.