Spermatogonial Stem Cell Numbers Are Reduced by Transient Inhibition of GDNF Signaling but Restored by Self-Renewing Replication when Signaling Resumes.

One cause of human male infertility is a scarcity of spermatogonial stem cells (SSCs) in testes with Sertoli cells that neither produce adequate amounts of GDNF nor form the Sertoli-Sertoli junctions that form the blood-testis barrier (BTB). These patients raise the issue of whether a pool of SSCs, depleted due to inadequate GDNF stimulation, will expand if normal signaling is restored. Here, we reduce adult mouse SSC numbers by 90% using a chemical-genetic approach that reversibly inhibits GDNF signaling. Signal resumption causes all remaining SSCs to replicate immediately, but they primarily form differentiating progenitor spermatogonia. Subsequently, self-renewing replication restores SSC numbers. Testicular GDNF levels are not increased during restoration. However, SSC replication decreases as numbers of SSCs and progenitors increase, suggesting important regulatory interactions among these cells. Finally, sequential loss of SSCs and then pachytene spermatocytes causes dissolution of the BTB, thereby recapitulating another important characteristic of some infertile men.

A pharmacological characterization of electrocardiogram PR and QRS intervals in conscious telemetered rats.

INTRODUCTION: The conscious telemetered rat is widely used as an early in vivo screening model for assessing the cardiovascular safety of novel pharmacological agents. The current study aimed to identify its utility in assessing electrocardiogram (ECG) PR and QRS interval changes. METHOD: Male Han-Wistar rats (~250 g) were implanted with radio-telemetry devices for the recording of ECG and haemodynamic parameters. Animals (n = 4-8) were treated with single doses of calcium (nifedipine, diltiazem or verapamil; CCBs) or sodium channel blockers (quinidine or flecainide; SCBs) or their corresponding vehicles in an ascending dose design. Data was recorded continuously up to 24 h post-dose. Pharmacokinetic analysis of blood samples was performed to allow comparison of effects to published data in other species. RESULTS: Of the CCBs, only diltiazem (300 mg/kg) prolonged the PR interval (49 ± 2 versus vehicle: 43 ± 1 ms), although this was not statistically significant (p = .11). QA interval decreased with nifedipine (30 ± 1 versus 24 ± 0 ms) and diltiazem (34 ± 1 versus 27 ± 1 ms) but increased with verapamil (30 ± 0 versus 37 ± 1 ms) demonstrating pharmacological activity of each agent. Both SCBs, caused statistically significant (p < .05) increases in both intervals - quinidine (100 mg/kg; PR: 50 ± 2 versus 43 ± 1 ms; QRS: 22 ± 2 versus 18 ± 1 ms) and flecainide (9 mg/kg; PR: 56 ± 1 versus 46 ± 1 ms; QRS: 27 ± 1 versus 21 ± 1 ms). Drug plasma exposure was confirmed in all animals. DISCUSSION: At similar plasma concentrations to other species, the conscious telemetered rat demonstrates limited utility in assessing PR interval prolongation by CCBs, despite significant contractility effects being observed. However, results with SCBs demonstrate a potential application for evaluating drug-induced QRS prolongation.

Glioblastoma Recurrence Correlates With Increased APE1 and Polarization Toward an Immuno-Suppressive Microenvironment.

While treatment with surgery, radiotherapy and/or chemotherapy may prolong life for patients with glioblastoma, recurrence is inevitable. What is still being discovered is how much these treatments and recurrence of disease affect the molecular profiles of these tumors and how these tumors adapt to withstand these treatment pressures. Understanding such changes will uncover pathways used by the tumor to evade destruction and will elucidate new targets for treatment development. Nineteen matched pre-treatment and post-treatment glioblastoma tumors were subjected to gene expression profiling (Fluidigm, TaqMan assays), MGMT promoter methylation analysis (pyrosequencing) and protein expression analysis of the DNA repair pathways, known to be involved in temozolomide resistance (immunohistochemistry). Gene expression profiling to molecularly subtype tumors revealed that 26% of recurrent post-treatment specimens did not match their primary diagnostic specimen subtype. Post-treatment specimens had molecular changes which correlated with known resistance mechanisms including increased expression of APEX1 (p < 0.05) and altered MGMT methylation status. In addition, genes associated with immune suppression, invasion and aggression (GPNMB, CCL5, and KLRC1) and polarization toward an M2 phenotype (CD163 and MSR1) were up-regulated in post-treatment tumors, demonstrating an overall change in the tumor microenvironment favoring aggressive tumor growth and disease recurrence. This was confirmed by in vitro studies that determined that glioma cell migration was enhanced in the presence of M2 polarized macrophage conditioned media. Further, M2 macrophage-modulated migration was markedly enhanced in post-treatment (temozolomide resistant) glioma cells. These findings highlight the ability of glioblastomas to evade not only the toxic onslaught of therapy but also to evade the immune system suggesting that immune-altering therapies may be of value in treating this terrible disease.

Variable flip angle 3D ultrashort echo time (UTE) T1 mapping of mouse lung: A repeatability assessment.

BACKGROUND: Lung T1 is a potential translational biomarker of lung disease. The precision and repeatability of variable flip angle (VFA) T1 mapping using modern 3D ultrashort echo time (UTE) imaging of the whole lung needs to be established before it can be used to assess response to disease and therapy. PURPOSE: To evaluate the feasibility of regional lung T1 quantification with VFA 3D-UTE and to investigate long- and short-term T1 repeatability in the lungs of naive mice. STUDY TYPE: Prospective preclinical animal study. POPULATION: Eight naive mice and phantoms. FIELD STRENGTH/SEQUENCE: 3D free-breathing radial UTE (8 µs) at 4.7T. ASSESSMENT: VFA 3D-UTE T1 calculations were validated against T1 values measured with inversion recovery (IR) in phantoms. Lung T1 and proton density (S0 ) measurements of whole lung and muscle were repeated five times over 1 month in free-breathing naive mice. Two consecutive T1 measurements were performed during one of the imaging sessions. STATISTICAL TESTS: Agreement in T1 between VFA 3D-UTE and IR in phantoms was assessed using Bland-Altman and Pearson 's correlation analysis. The T1 repeatability in mice was evaluated using coefficient of variation (CV), repeated-measures analysis of variance (ANOVA), and paired t-test. RESULTS: Good T1 agreement between the VFA 3D-UTE and IR methods was found in phantoms. T1 in lung and muscle showed a 5% and 3% CV (1255 ± 63 msec and 1432 ± 42 msec, respectively, mean ± SD) with no changes in T1 or S0 over a month. Consecutive measurements resulted in an increase of 2% in both lung T1 and S0 . DATA CONCLUSION: VFA 3D-UTE shows promise as a reliable T1 mapping method that enables full lung coverage, high signal-to-noise ratio (∼25), and spatial resolution (300 µm) in freely breathing animals. The precision of the VFA 3D-UTE method will enable better design and powering of studies. LEVEL OF EVIDENCE: 1 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2018.

Larval western bean cutworm feeding damage encourages the development of Gibberella ear rot on field corn.

BACKGROUND: A 2 year study was conducted to determine whether western bean cutworm (Striacosta albicosta Smith) (WBC) larval feeding damage increases severity of the fungal disease Gibberella ear rot [Fusarium graminearum (Schwein.) Petch] in field corn (Zea mays L.). The effect of a quinone-outside inhibiting fungicide, pyraclostrobin, on Gibberella ear rot severity and mycotoxin production, both with and without WBC pressure, was also evaluated. The impact of each variable was assessed individually and in combination to determine the effect of each upon ear disease severity. RESULTS: There was a positive correlation between the presence of WBC larvae in field corn and Gibberella ear rot severity under inoculated conditions in the 2 years of the experiment. An application of pyraclostrobin did not impact Gibberella ear rot development when applied at corn growth stage R1 (silks first emerging). CONCLUSION: Feeding damage from WBC larvae significantly increases the development of F. graminearum in field corn. We conclude that an effective integrated management strategy for Gibberella ear rot should target the insect pest first, in an effort to limit disease severity and subsequent mycotoxin production by F. graminearum in kernels. © 2016 Society of Chemical Industry.

Analysis of Arabidopsis Accessions Hypersensitive to a Loss of Chloroplast Translation.

Natural accessions of Arabidopsis (Arabidopsis thaliana) differ in their ability to tolerate a loss of chloroplast translation. These differences can be attributed in part to variation in a duplicated nuclear gene (ACC2) that targets homomeric acetyl-coenzyme A carboxylase (ACCase) to plastids. This functional redundancy allows limited fatty acid biosynthesis to occur in the absence of heteromeric ACCase, which is encoded in part by the plastid genome. In the presence of functional ACC2, tolerant alleles of several nuclear genes, not yet identified, enhance the growth of seedlings and embryos disrupted in chloroplast translation. ACC2 knockout mutants, by contrast, are hypersensitive. Here we describe an expanded search for hypersensitive accessions of Arabidopsis, evaluate whether all of these accessions are defective in ACC2, and characterize genotype-to-phenotype relationships for homomeric ACCase variants identified among 855 accessions with sequenced genomes. Null alleles with ACC2 nonsense mutations, frameshift mutations, small deletions, genomic rearrangements, and defects in RNA splicing are included among the most sensitive accessions examined. By contrast, most missense mutations affecting highly conserved residues failed to eliminate ACC2 function. Several accessions were identified where sensitivity could not be attributed to a defect in either ACC2 or Tic20-IV, the chloroplast membrane channel required for ACC2 uptake. Overall, these results underscore the central role of ACC2 in mediating Arabidopsis response to a loss of chloroplast translation, highlight future applications of this system to analyzing chloroplast protein import, and provide valuable insights into the mutational landscape of an important metabolic enzyme that is highly conserved throughout eukaryotes.

Notes from the Field: Increase in Neisseria meningitidis-Associated Urethritis Among Men at Two Sentinel Clinics - Columbus, Ohio, and Oakland County, Michigan, 2015.

Neisseria meningitidis (Nm) urogenital infections, although less common than infections caused by Neisseria gonorrhoeae (Ng), have been associated with urethritis, cervicitis, proctitis, and pelvic inflammatory disease. Nm can appear similar to Ng on Gram stain analysis (gram-negative intracellular diplococci) (1-5). Because Nm colonizes the nasopharynx, men who receive oral sex (fellatio) can acquire urethral Nm infections (1,3,5). This report describes an increase in Nm-associated urethritis in men attending sexual health clinics in Columbus, Ohio, and Oakland County, Michigan.

Intratumoral heterogeneity identified at the epigenetic, genetic and transcriptional level in glioblastoma.

Heterogeneity is a hallmark of glioblastoma with intratumoral heterogeneity contributing to variability in responses and resistance to standard treatments. Promoter methylation status of the DNA repair enzyme O(6)-methylguanine DNA methyltransferase (MGMT) is the most important clinical biomarker in glioblastoma, predicting for therapeutic response. However, it does not always correlate with response. This may be due to intratumoral heterogeneity, with a single biopsy unlikely to represent the entire lesion. Aberrations in other DNA repair mechanisms may also contribute. This study investigated intratumoral heterogeneity in multiple glioblastoma tumors with a particular focus on the DNA repair pathways. Transcriptional intratumoral heterogeneity was identified in 40% of cases with variability in MGMT methylation status found in 14% of cases. As well as identifying intratumoral heterogeneity at the transcriptional and epigenetic levels, targeted next generation sequencing identified between 1 and 37 unique sequence variants per specimen. In-silico tools were then able to identify deleterious variants in both the base excision repair and the mismatch repair pathways that may contribute to therapeutic response. As these pathways have roles in temozolomide response, these findings may confound patient management and highlight the importance of assessing multiple tumor biopsies.

Molecular heterogeneity in glioblastoma: potential clinical implications.

Glioblastomas, (grade 4 astrocytomas), are aggressive primary brain tumors characterized by histopathological heterogeneity. High-resolution sequencing technologies have shown that these tumors also feature significant inter-tumoral molecular heterogeneity. Molecular subtyping of these tumors has revealed several predictive and prognostic biomarkers. However, intra-tumoral heterogeneity may undermine the use of single biopsy analysis for determining tumor genotype and has implications for potential targeted therapies. The clinical relevance and theories of tumoral molecular heterogeneity in glioblastoma are discussed.

Hepatitis A outbreak among adults with developmental disabilities in group homes--Michigan, 2013.

Hepatitis A virus (HAV) infections among persons with developmental disabilities living in institutions were common in the past, but with improvements in care and fewer persons institutionalized, the number of HAV infections has declined in these institutions. However, residents in institutions are still vulnerable if they have not been vaccinated. On April 24, 2013, a resident of a group home (GH) for adults with disabilities in southeast Michigan (GH-A) was diagnosed with hepatitis A and died 2 days later of fulminant liver failure. Four weeks later, a second GH-A resident was diagnosed with hepatitis A. None of the GH-A residents or staff had been vaccinated against hepatitis A. Over the next 3 months, six more cases of hepatitis A were diagnosed in residents in four other Michigan GHs. Three local health departments were involved in case investigation and management, including administration of postexposure prophylaxis (PEP). Serum specimens from seven cases were found to have an identical strain of HAV genotype 1A. This report describes the outbreak investigation, the challenges of timely delivery of PEP for hepatitis A, and the need for preexposure vaccination against hepatitis A for adults living or working in GHs for the disabled.

Factors that may influence the willingness of cancer patients to consent for biobanking.

BACKGROUND: Broad or general consent given by cancer patients for their tissue, blood, and clinical information to be stored in institutional biorepositories is fundamental to enable future ethical translational cancer research. The decision to consent for biobanking will contribute to the development of advanced diagnostic and prognostic tests, as well as new therapies to improve patient outcomes. While the rate of patient participation in biobanking programs is generally reported as high worldwide, few studies have investigated factors that may influence this decision. Biobanking at our medical research institute, an associated public (government-run) university hospital, and private hospital has been established for over 20 years, with collection of certain tumor types embedded in the research culture of these institutions. In this study, we investigated factors that may influence a cancer patient's decision to give broad consent for biobanking of their specimens. METHODS: Data on patient consent were collected over a 6-month period from both government and private hospitals associated with our medical research institute. Factors considered included gender, patient age at surgery, type of malignancy (breast, duodenal, cervical, endometrial, gastric, liver, esophageal, ovarian, pancreatic, pelvic, uterine, or vulval), type of institution where surgery was performed, and timing of consent. RESULTS: Of 171 cancer patients, 159 (93%) gave broad consent for biobanking of their tissue and blood specimens for future cancer research projects receiving ethical and scientific approval. None of the factors analyzed was shown to influence a patient's decision to contribute biological specimens and clinical data to a biorepository for future medical research. CONCLUSION: Biobanking for future ethically and scientifically approved research projects in an established institution is an initiative that receives strong support from patients undergoing cancer surgery, independent of factors including gender, age, type of tumor, type of institution where surgery was performed, or timing of consent.

Natural variation in sensitivity to a loss of chloroplast translation in Arabidopsis.

Mutations that eliminate chloroplast translation in Arabidopsis (Arabidopsis thaliana) result in embryo lethality. The stage of embryo arrest, however, can be influenced by genetic background. To identify genes responsible for improved growth in the absence of chloroplast translation, we examined seedling responses of different Arabidopsis accessions on spectinomycin, an inhibitor of chloroplast translation, and crossed the most tolerant accessions with embryo-defective mutants disrupted in chloroplast ribosomal proteins generated in a sensitive background. The results indicate that tolerance is mediated by ACC2, a duplicated nuclear gene that targets homomeric acetyl-coenzyme A carboxylase to plastids, where the multidomain protein can participate in fatty acid biosynthesis. In the presence of functional ACC2, tolerance is enhanced by a second locus that maps to chromosome 5 and heightened by additional genetic modifiers present in the most tolerant accessions. Notably, some of the most sensitive accessions contain nonsense mutations in ACC2, including the "Nossen" line used to generate several of the mutants studied here. Functional ACC2 protein is therefore not required for survival in natural environments, where heteromeric acetyl-coenzyme A carboxylase encoded in part by the chloroplast genome can function instead. This work highlights an interesting example of a tandem gene duplication in Arabidopsis, helps to explain the range of embryo phenotypes found in Arabidopsis mutants disrupted in essential chloroplast functions, addresses the nature of essential proteins encoded by the chloroplast genome, and underscores the value of using natural variation to study the relationship between chloroplast translation, plant metabolism, protein import, and plant development.

Responses to glial cell line-derived neurotrophic factor change in mice as spermatogonial stem cells form progenitor spermatogonia which replicate and give rise to more differentiated progeny.

Spermatogonial stem cells (SSCs) are the foundation of spermatogenesis. These cells are classically defined as a subset of morphologically defined A single (As) spermatogonia, which can produce more SSCs or they can give rise to nonstem As cells that, upon replication, generate A paired (Apr) and then A aligned (Aal) spermatogonia. These latter two cell types, along with the nonstem As cells, function as transit-amplifying progenitor cells. It is known that glial cell line-derived neurotrophic factor (GDNF) is essential for maintaining all of these cells, but it is unknown if or how the responses of these cells change as they progress down the pathway to differentiated type A1 spermatogonia. We address this issue by using a chemical-genetic approach to inhibit GDNF signaling in vivo and an in vitro approach to increase GDNF stimulation. We show that inhibition for 2 days suppresses replication of As, Apr, and Aal spermatogonia to an equal extent, whereas stimulation by GDNF preferentially increases replication of As and Apr spermatogonia. We also test if inhibiting GDNF signaling causes As, Apr, and Aal spermatogonia to express Kit, an essential step in their differentiation into type A1 spermatogonia. Inhibition for 3 or 7 days produces a progressive increase in the percentages of As, Apr, and Aal undergoing differentiation, with the largest increase observed in Aal spermatogonia. Finally, we demonstrate that numbers of SSCs decrease more slowly than numbers of progenitor spermatogonia when GDNF signaling is inhibited. Taken together, these data suggest that there are significant changes in the responses to GDNF as SSCs give rise to progenitor spermatogonia, which replicate and gradually differentiate into type A1 spermatogonia.

Differences in respiratory syncytial virus and influenza infection in a house-dust-mite-induced asthma mouse model: consequences for steroid sensitivity.

A significant number of clinical asthma exacerbations are triggered by viral infection. We aimed to characterize the effect of virus infection in an HDM (house dust mite) mouse model of asthma and assess the effect of oral corticosteroids. HDM alone significantly increased eosinophils, lymphocytes, neutrophils, macrophages and a number of cytokines in BAL (bronchoalveolar lavage), all of which were sensitive to treatment with prednisolone (with the exception of neutrophils). Virus infection also induced cell infiltration and cytokines. RSV (respiratory syncytial virus) infection in HDM-treated animals further increased all cell types in BAL (except eosinophils, which declined), but induced no further increase in HDM-elicited cytokines. However, while HDM-elicited TNF-α (tumour necrosis factor-α), IFN-γ (interferon-γ), IL (interleukin)-2, IL-5 and IL-10 were sensitive to prednisolone treatment, concomitant infection with RSV blocked the sensitivity towards steroid. In contrast, influenza infection in HDM- challenged animals resulted in increased BAL lymphocytes, neutrophils, IFN-γ, IL-1ß, IL-4, IL-5, IL-10 and IL-12, but all were attenuated by prednisolone treatment. HDM also increased eNO (exhaled NO), which was further increased by concomitant virus infection. This increase was only partially attenuated by prednisolone. RSV infection alone increased BAL mucin. However, BAL mucin was increased in HDM animals with virus infection. Chronic HDM challenge in mice elicits a broad inflammatory response that shares many characteristics with clinical asthma. Concomitant influenza or RSV infection elicits differing inflammatory profiles that differ in their sensitivity towards steroids. This model may be suitable for the assessment of novel pharmacological interventions for asthmatic exacerbation.

A resurrected mammalian hAT transposable element and a closely related insect element are highly active in human cell culture.

Chromosome structure and function are influenced by transposable elements, which are mobile DNA segments that can move from place to place. hAT elements are a superfamily of DNA cut and paste elements that move by excision and integration. We have characterized two hAT elements, TcBuster and Space Invaders (SPIN), that are members of a recently described subfamily of hAT elements called Buster elements. We show that TcBuster, from the red flour beetle Tribolium castaneum, is highly active in human cells. SPIN elements are currently inactive elements that were recently highly active in multiple vertebrate genomes, and the high level of sequence similarity across widely diverged species and patchy phylogenetic distribution suggest that they may have moved between genomes by horizontal transfer. We have generated an intact version of this element, SPIN(ON), which is highly active in human cells. In vitro analysis of TcBuster and SPIN(ON) shows that no proteins other than transposase are essential for recombination, a property that may contribute to the ability of SPIN to successfully invade multiple organisms. We also analyze the target site preferences of de novo insertions in the human genome of TcBuster and SPIN(ON) and compare them with the preferences of Sleeping Beauty and piggyBac, showing that each superfamily has a distinctive pattern of insertion. The high-frequency transposition of both TcBuster and SPIN(ON) suggests that these transposon systems offer powerful tools for genome engineering. Finally, we describe a Saccharomyces cerevisiae assay for TcBuster that will provide a means for isolation of hyperactive and other interesting classes of transposase mutants.

Correlation of MicroRNA 132 Up-regulation with an Unfavorable Clinical Outcome in Patients with Primary Glioblastoma Multiforme Treated with Radiotherapy Plus Concomitant and Adjuvant Temozolomide Chemotherapy.

BACKGROUND: MicroRNA 132 (miR-132) is dysregulated in a range of human malignancies; however, its role in glioma has not been reported. The aim of this study was to profile miR-132 expression in a cohort of patients with primary glioblastoma multiforme (GBM) treated with the Stupp regimen and to correlate microRNA levels with patient outcome. METHODS: miR-132 levels relative to RNU44 were assessed by quantitative reverse transcription-polymerase chain reaction in 43 GBMs and normal brain tissue. The cohort comprised patients less than 72 years of age with Eastern Cooperative Oncology Group (ECOG) scores between 0 and 2 who had undergone 6-week concomitant radiation and temozolomide followed by adjuvant temozolomide. Survival data were available for all cases. Tumors were characterized for O6-methylguanine-DNA methyltransferase (MGMT) methylation and isocitrate dehydrogenase (IDH) 1/2 mutation status. Associations between miR-132 expression and clinical indicators were analyzed. RESULTS: Tumor miR-132 levels ranged from 0.07- to 40.4-fold increase (mean = 5.5-fold increase) relative to normal brain. High-level miR-132 (above the mean) independently predicted for a significantly shorter overall survival (P = .008). miR-132 was a stronger prognostic indicator than ECOG score (P = .012) and age at diagnosis (P = .026) but did not correlate with MGMT methylation status or extent of tumor resection. Cox regression analysis confirmed high miR-132 as the strongest predictor of outcome (P = .010) with a hazard ratio of 2.8. CONCLUSIONS: This study identified high miR-132 expression as a biomarker of poor prognosis in patients with primary GBM treated with the Stupp regimen.

An in vivo model of epithelial to mesenchymal transition reveals a mitogenic switch.

The epithelial to mesenchymal transition (EMT) is a process by which differentiated epithelial cells transition to a mesenchymal phenotype. EMT enables the escape of epithelial cells from the rigid structural constraints of the tissue architecture to a phenotype more amenable to cell migration and, therefore, invasion and metastasis. We characterized an in vivo model of EMT and discovered that marked changes in mitogenic signaling occurred during this process. DNA microarray analysis revealed that the expression of a number of genes varied significantly between post-EMT and pre-EMT breast cancer cells. Post-EMT cancer cells upregulated mRNA encoding c-Met and the PDGF and LPA receptors, and acquired increased responsiveness to HGF, PDGF, and LPA. This rendered the post-EMT cells responsive to the growth inhibitory effects of HGF, PDGF, and LPA receptor inhibitors/antagonists. Furthermore, post-EMT cells exhibited decreased basal Raf and Erk phosphorylation, and in comparison to pre-EMT cells, their proliferation was poorly inhibited by a MEK inhibitor. These studies suggest that therapies need to be designed to target both pre-EMT and post-EMT cancer cells and that signaling changes in post-EMT cells may allow them to take advantage of paracrine signaling from the stroma in vivo.

E-cadherin plasticity in prostate cancer stem cell invasion.

Prostate cancer that has progressed to metastatic disease remains largely untreatable. Nearly 90% of patients with advanced prostate cancer develop skeletal metastases, resulting in a substantial reduction in the quality of life and a drastic worsening of patient prognosis. The mechanisms involved in prostate cancer cell dissemination, however, remain poorly understood. We previously reported the identification of a highly tumorigenic E-cadherin positive prostate tumor stem cell subpopulation that expressed the embryonic stem cell markers SOX2 and OCT3/4. We herein demonstrate that this subpopulation is also highly invasive and, importantly, is capable of altering its E-cadherin expression during the process of invasion. The non-tumorigenic E-cadherin negative subpopulation which minimally expresses SOX2 or OCT3/4 was found to be poorly invasive. In addition, targeted knockdown of SOX2 or OCT3/4 markedly suppressed the invasion of prostate cancer cells. Taken together, these findings indicate that the expression of SOX2 or OCT3/4 is required for invasive cell capacity, but the ability to modulate E-cadherin is the key permissive factor enabling cancer stem cell invasion in vitro. We therefore propose a model in which the post-epithelial to mesenchymal transition phenotype progresses to a frank, aggressive, and invasive phenotype by a process requiring the acquisition of E-cadherin plasticity. Considering the clinical significance of the metastatic complications of prostate adenocarcinoma, the identification of factors that promote the dissemination of the malignant prostate phenotype is essential to establish effective therapies to combat this disease in future.

A versatile and tunable coating strategy allows control of nanocrystal delivery to cell types in the liver.

There are many liver diseases that could be treated with delivery of therapeutics such as DNA, proteins, or small molecules. Nanoparticles are often proposed as delivery vectors for such therapeutics; however, achieving nanoparticle accumulations in the therapeutically relevant hepatocytes is challenging. In order to address this issue, we have synthesized polymer coated, fluorescent iron oxide nanoparticles that bind and deliver DNA, as well as produce contrast for magnetic resonance imaging (MRI), fluorescence imaging, and transmission electron microscopy (TEM). The composition of the coating can be varied in a facile manner to increase the quantity of poly(ethylene glycol) (PEG) from 0% to 5%, 10%, or 25%, with the aim of reducing opsonization but maintaining DNA binding. We investigated the effect of the nanoparticle coating on DNA binding, cell uptake, cell transfection, and opsonization in vitro. Furthermore, we exploited MRI, fluorescence imaging, and TEM to investigate the distribution of the different formulations in the liver of mice. While MRI and fluorescence imaging showed that each formulation was heavily taken up in the liver at 24 h, the 10% PEG formulation was taken up by the therapeutically relevant hepatocytes more extensively than either the 0% PEG or the 5% PEG, indicating its potential for delivery of therapeutics to the liver.

Immunohistochemistry for SDHB divides gastrointestinal stromal tumors (GISTs) into 2 distinct types.

The Carney triad (CT) is gastrointestinal stromal tumor (GIST), paraganglioma, and pulmonary chondroma. The GISTs of CT show different clinical, molecular, and morphologic features to usual adult GISTs but are similar to the majority of pediatric GISTs. We postulated that these GISTs would show negative staining for succinate dehydrogenase B (SDHB). We performed SDHB immunohistochemistry on GISTs arising in 5 individuals with CT, 1 child, 7 individuals with GIST in young adulthood including 2 with germline KIT mutations, 3 individuals with neurofibromatosis 1, one 63-year-old female with multifocal gastric epithelioid GIST with lymph node metastases, and 104 consecutive unselected individuals with apparently sporadic GIST. The GISTs and paragangliomas arising in CT, the pediatric GIST, and the multifocal gastric GIST from the 63-year-old showed negative SDHB staining. GISTs from the 7 young adults and 3 with neurofibromatosis were SDHB positive. Of the unselected GISTs, 101 (97%) were positive. One of the negative GISTs arose in a 48-year-old female with previous recurrent multifocal gastric GISTs and the other 2 arose in females also in their 40s with gastric GISTs with epithelioid morphology. We conclude that negative staining for SDHB is characteristic of the GISTs of CT and the subgroup of pediatric GISTs which it resembles. Furthermore, when negative staining occurs in apparently sporadic GISTs in adults, the GISTs show morphologic and clinical features similar to pediatric and CT type GISTs. GISTs may therefore be divided into type 1 (SDHB positive) and type 2 (SDHB negative) subtypes.