RhoB controls endothelial cell morphogenesis in part via negative regulation of RhoA

Recent studies have suggested a role for the small GTPase RhoB in the control of processes required for angiogenesis. However, the mechanisms whereby RhoB exerts control over these processes are not well understood. Given the role of vascular endothelial growth factor (VEGF) in pathological angiogenesis, we were interested in examining whether RhoB contributed to VEGF-induced angiogenic processes. To assess this, RhoB was specifically depleted in human umbilical vein endothelial cells (HUVEC), using siRNA-targeted strategies. The effects of RhoB depletion on VEGF-induced angiogenic activities were assessed using a variety of standard in vitro angiogenesis assays to assess endothelial cell viability, migration and capillary morphogenesis. Effects of RhoB depletion on signaling from other Rho family member proteins was also assessed using specific activity assays for RhoA and RhoC. We observed that although RhoB appeared dispensable for HUVEC viability, RhoB was required for endothelial cell migration, sprouting, and capillary morphogenesis. We also observed that siRNA-mediated depletion of RhoB in HUVEC resulted in increased RhoA activation in response to VEGF stimulation. This increased RhoA activation contributed to the cellular morphogenesis defects observed in RhoB-depleted cells, as inhibition of RhoA activity using C3 transferase, or inhibition of the activity of the downstream RhoA effectors Rho-dependent kinases I and II (ROCK I and II) led to a partial restoration of capillary morphogenesis in the absence of RhoB. Thus our data indicate that RhoB plays a significant role in VEGF-induced endothelial cell morphogenesis in part by negatively regulating the activity of RhoA and the RhoA/ROCK pathway

Reflections on a Mentoring Partnership Journey

This commentary offers reflection on the mentoring partnership journey between a senior Fellow of the American Academy of Health Behavior (AAHB) and three early- or mid-career AAHB members. Their partnership was supported by the AAHB Research Scholars Mentorship Program. The authors discuss the nature of their working relationship, products they generated, and other lessons learned from the experience. The authors also offer their perspectives on effective mentorship characteristics

PSMA Redirects Cell Survival Signaling From The MAPK To The PI3K-AKT Pathways To Promote The Progression Of Prostate Cancer

Increased abundance of the prostate-specific membrane antigen (PSMA) on prostate epithelium is a hallmark of advanced metastatic prostate cancer (PCa) and correlates negatively with prognosis. However, direct evidence that PSMA functionally contributes to PCa progression remains elusive. We generated mice bearing PSMA-positive or PSMA-negative PCa by crossing PSMA-deficient mice with transgenic PCa (TRAMP) models, enabling direct assessment of PCa incidence and progression in the presence or absence of PSMA. Compared with PSMA-positive tumors, PSMA-negative tumors were smaller, lower-grade, and more apoptotic with fewer blood vessels, consistent with the recognized proangiogenic function of PSMA. Relative to PSMA-positive tumors, tumors lacking PSMA had less than half the abundance of type 1 insulin-like growth factor receptor (IGF-1R), less activity in the survival pathway mediated by PI3K-AKT signaling, and more activity in the proliferative pathway mediated by MAPK-ERK1/2 signaling. Biochemically, PSMA interacted with the scaffolding protein RACK1, disrupting signaling between the β1 integrin and IGF-1R complex to the MAPK pathway, enabling activation of the AKT pathway instead. Manipulation of PSMA abundance in PCa cell lines recapitulated this signaling pathway switch. Analysis of published databases indicated that IGF-1R abundance, cell proliferation, and expression of transcripts for antiapoptotic markers positively correlated with PSMA abundance in patients, suggesting that this switch may be relevant to human PCa. Our findings suggest that increase in PSMA in prostate tumors contributes to progression by altering normal signal transduction pathways to drive PCa progression and that enhanced signaling through the IGF-1R/β1 integrin axis may occur in other tumor

Human GLB1 knockout cerebral organoids: A model system for testing AAV9-mediated GLB1 gene therapy for reducing GM1 ganglioside storage in GM1 gangliosidosis

GM1 gangliosidosis is an autosomal recessive neurodegenerative disorder caused by the deficiency of lysosomal gangliosidebeta-galactosidase (beta-gal) and resulting in accumulation of GM1 ganglioside. The disease spectrum ranges from infantile to late onset and is uniformly fatal, with no effective therapy currently available. Although animal models have been useful for understanding disease pathogenesis and exploring therapeutic targets, no relevant human central nervous system (CNS) model system has been available to study its early pathogenic events or test therapies. To develop a model of human GM1 gangliosidosis in the CNS, we employed CRISPR/Cas9 genome editing to target GLB1 exons 2 and 6, common sites for mutations in patients, to create isogenic induced pluripotent stem (iPS) cell lines with lysosomal beta-gal deficiency. We screened for clones with \u3c 5% of parental cell line beta-gal enzyme activity and confirmed GLB1 knockout clones using DNA sequencing. We then generated GLB1 knockout cerebral organoids from one of these GLB1 knockout iPS cell clones. Analysis of GLB1 knockout organoids in culture revealed progressive accumulation of GM1 ganglioside. GLB1 knockout organoids microinjected with AAV9-GLB1 vector showed a significant increase in beta-gal activity and a significant reduction in GM1 ganglioside content compared with AAV9-GFP-injected organoids, demonstrating the efficacy of an AAV9 gene therapy-based approach in GM1 gangliosidosis. This proof-of-concept in a human cerebral organoid model completes the pre-clinical studies to advance to clinical trials using the AAV9-GLB1 vector

Coupling snowpack and groundwater dynamics to interpret historical streamflow trends in the western United States

A key challenge for resource and land managers is predicting the consequences of climate warming on streamflow and water resources. During the last century in the western United States, significant reductions in snowpack and earlier snowmelt have led to an increase in the fraction of annual streamflow during winter and a decline in the summer. Previous work has identified elevation as it relates to snowpack dynamics as the primary control on streamflow sensitivity to warming. But along with changes in the timing of snowpack accumulation and melt, summer streamflows are also sensitive to intrinsic, geologically mediated differences in the efficiency of landscapes in transforming recharge (either as rain or snow) into discharge; we term this latter factor drainage efficiency. Here we explore the conjunction of drainage efficiency and snowpack dynamics in interpreting retrospective trends in summer streamflow during 1950–2010 using daily streamflow from 81 watersheds across the western United States. The recession constant (k) and fraction of precipitation falling as snow (S[subscript f]) were used as metrics of deep groundwater and overall precipitation regime (rain and/or snow), respectively. This conjunctive analysis indicates that summer streamflows in watersheds that drain slowly from deep groundwater and receive precipitation as snow are most sensitive to climate warming. During the spring, however, watersheds that drain rapidly and receive precipitation as snow are most sensitive to climate warming. Our results indicate that not all trends in western United States are associated with changes in snowpack dynamics; we observe declining streamflow in late fall and winter in rain-dominated watersheds as well. These empirical findings support both theory and hydrologic modelling and have implications for how streamflow sensitivity to warming is interpreted across broad regions. Copyright © 2012 John Wiley & Sons, Ltd.Keywords: Groundwater processes, Climate, Warming, Hydrologic processes, Streamflow tren

High-Performance Silicon Photonic Single-Sideband Modulators for Cold Atom Interferometry

The most complicated and challenging system within a light-pulse atom interferometer (LPAI) is the laser system, which controls the frequencies and intensities of multiple laser beams over time to configure quantum gravity and inertial sensors. The main function of an LPAI laser system is to perform cold-atom generation and state-selective detection and to generate coherent two-photon process for the light-pulse sequence. Substantial miniaturization and ruggedization of the laser system can be achieved by bringing together most key functions of the laser and optical system onto a photonic integrated circuit (PIC). Here we demonstrate a high-performance silicon photonic carrier-suppressed single-sideband (CS-SSB) modulator PIC with dual-parallel Mach-Zehnder modulators (DP-MZMs) operating near 1560 nm, which can dynamically shift the frequency of the light for the desired function within the LPAI. Independent RF control of channels in SSB modulator enables the extensive study of imbalances in both the optical and RF phases and amplitudes to simultaneously reach 30 dB carrier suppression and unprecedented 47.8 dB sideband suppression with peak conversion efficiency of -6.846 dB (20.7 %). Using a silicon photonic SSB modulator with time-multiplexed frequency shifting in an LPAI laser system, we demonstrate cold-atom generation, state-selective detection, and the realization of atom interferometer fringes to estimate gravitational acceleration, $g \approx 9.77 \pm 0.01 \,\rm{m/s^2}$, in a Rubidium ($^{87}$Rb) atom system.Comment: 18 pages, 9 figure

HPV Genotypes in High Grade Cervical Lesions and Invasive Cervical Carcinoma as Detected by Two Commercial DNA Assays, North Carolina, 2001–2006

HPV typing using formalin fixed paraffin embedded (FFPE) cervical tissue is used to evaluate HPV vaccine impact, but DNA yield and quality in FFPE specimens can negatively affect test results. This study aimed to evaluate 2 commercial assays for HPV detection and typing using FFPE cervical specimens.Four large North Carolina pathology laboratories provided FFPE specimens from 299 women ages18 and older diagnosed with cervical disease from 2001 to 2006. For each woman, one diagnostic block was selected and unstained serial sections were prepared for DNA typing. Extracts from samples with residual lesion were used to detect and type HPV using parallel and serial testing algorithms with the Linear Array and LiPA HPV genotyping assays.LA and LiPA concordance was 0.61 for detecting any high-risk (HR) and 0.20 for detecting any low-risk (LR) types, with significant differences in marginal proportions for HPV16, 51, 52, and any HR types. Discordant results were most often LiPA-positive, LA-negative. The parallel algorithm yielded the highest prevalence of any HPV type (95.7%). HR type prevalence was similar using parallel (93.1%) and serial (92.1%) approaches. HPV16, 33, and 52 prevalence was slightly lower using the serial algorithm, but the median number of HR types per woman (1) did not differ by algorithm. Using the serial algorithm, HPV DNA was detected in >85% of invasive and >95% of pre-invasive lesions. The most common type was HPV16, followed by 52, 18, 31, 33, and 35; HPV16/18 was detected in 56.5% of specimens. Multiple HPV types were more common in lower grade lesions.We developed an efficient algorithm for testing and reporting results of two commercial assays for HPV detection and typing in FFPE specimens, and describe HPV type distribution in pre-invasive and invasive cervical lesions in a state-based sample prior to HPV vaccine introduction