Biology subjects

Gu, W.

Publications and source records attributed to Gu, W..

100 records · Page 6Linked to original sources

bioRxiv2020

Core Role of TRPC6 in Regulating Airway Re-modelling in Chronic Obstructive Pulmonary Disease

RationaleThe mechanistic role of canonical transient receptor potential 6 (TRPC6) channel in chronic obstructive pulmonary disease (COPD) is poorly understood. ObjectivesThe purpose of this study is to determine the role of TRPC6 channel in COPD and its underlying signaling mechanisms in human airway smooth muscle cells (HASMCs). Methods and Main ResultsThe present study examined the effects of TRPC6 channel on nicotine and cigarette induced HASMCs proliferation, migration and mouse airway remodeling models. mRNA and protein expression of TRPC6 were increased in cultured HASMCs incubated with nicotine using real-time PCR and western blot analysis. Nicotine treatment significantly increased TRPC6 transcriptional activity through NF-{kappa}B in HASMCs with Co-IP and electrophoretic mobility shift assays (EMSA). Nicotine treatment also increased ROS level in HASMCs, this increase was attenuated by Nox inhibitor apocynin. miR-135a/b-5p down-regulated mRNA and protein level of TRPC6 in HASMCs, while luciferase reporter assay showed that miR-135a/b-5p targeted at the 3-UTR of TRPC6 mRNA. microRNA-135a/b-5p (miR-135a/b-5p), with a negative correlation to TRPC6 expression, was low in airway smooth muscle of COPD patients. Cigarette-induced airway remodeling mice model also exhibited a large increase in smooth muscle cell proliferation and smooth muscle layer mass with immunohistochemistry assay, this well-characterized airway remodeling was eliminated by lentivirus of TRPC6 knockdown or miR-135a/b-5p overexpression. ConclusionsNicotine exposure results in increased HASMCs proliferation and migration through NF-{kappa}B signaling. Inhalation of cigarette causes airway smooth muscle layer re-modeling due to altered TRPC6 elicited Ca2+ influx, miR-135a/b-5p abolishes this change both in vitro and in vivo.

cell biology↗

bioRxiv2019

TGFβ superfamily signaling regulates the state of human stem cell pluripotency and competency to create telencephalic organoids

Telencephalic organoids generated from human pluripotent stem cells (hPSCs) are emerging as an effective system to study the distinct features of the developing human brain and the underlying causes of many neurological disorders. While progress in organoid technology has been steadily advancing, many challenges remain including rampant batch-to-batch and cell line-to-cell line variability and irreproducibility. Here, we demonstrate that a major contributor to successful cortical organoid production is the manner in which hPSCs are maintained prior to differentiation. Optimal results were achieved using fibroblast-feeder-supported hPSCs compared to feeder-independent cells, related to differences in their transcriptomic states. Feeder-supported hPSCs display elevated activation of diverse TGF{beta} superfamily signaling pathways and increased expression of genes associated with naive pluripotency. We further identify combinations of TGF{beta}-related growth factors that are necessary and together sufficient to impart broad telencephalic organoid competency to feeder-free hPSCs and enable reproducible formation of brain structures suitable for disease modeling. HIGHLIGHTSO_LIhPSC maintenance conditions influence outcomes in cortical organoid formation C_LIO_LIIdentification of an intermediate pluripotency state optimal for cortical organoids C_LIO_LIFeeder support involves activation of diverse TGF{beta} signaling pathways C_LIO_LIThe organoid-promoting effects of feeders can be mimicked by a TGF{beta} factor mixture C_LI

developmental biology↗

bioRxiv2019

Quantitative analysis of 1300-nm three-photon calcium imaging in the mouse brain

1300-nm three-photon calcium imaging has emerged as a useful technique to allow calcium imaging in deep brain regions. Application to large-scale neural activity imaging entails a careful balance between recording fidelity and tissue heating. We calculated and experimentally verified the excitation pulse energy to achieve the minimum photon count required for the detection of calcium transients in GCaMP6s-expressing neurons for 920-nm two-photon and 1320-nm three-photon excitation, respectively. Brain tissue heating by continuous three-photon imaging was simulated with Monte Carlo method and experimentally validated with immunohistochemistry. We observed increased immunoreactivity with 150 mW excitation power at 1.0- and 1.2-mm imaging depths. Based on the data, we explained how three-photon excitation achieves better calcium imaging fidelity than two-photon excitation in the deep brain and quantified the imaging depth where three-photon microscopy should be applied. Our analysis presents a translatable model for the optimization of three-photon calcium imaging based on experimentally tractable parameters.

neuroscience↗

bioRxiv2019

Induction of pancreatic tumor-selective ferroptosis through modulation of cystine import

Pancreatic ductal adenocarcinoma (PDA) is the third-leading cause of cancer mortality in the US and is highly resistant to classical, targeted, and immune therapies. We show that human PDA cells are dependent on the provision of exogenous cystine to avert a catastrophic accumulation of lipid reactive oxygen species (ROS) that, left unchecked, leads to ferroptotic cell death, both in vitro and in vivo. Using a dual-recombinase genetically engineered model, we found that acute deletion of Slc7a11 led to tumor-selective ferroptosis, tumor stabilizations/regressions, and extended overall survival. The mechanism of ferroptosis induction in PDA cells required the concerted depletion of both glutathione and coenzyme A, highlighting a novel branch of ferroptosis-relevant metabolism. Finally, we found that cystine depletion in vivo using the pre-IND agent cyst(e)inase phenocopied Slc7a11 deletion, inducing tumor-selective ferroptosis and disease stabilizations/regressions in the well-validated KPC model of PDA.\n\nOne Sentence SummaryGenetic and pharmacological targeting of cystine import induces pancreatic cancer-selective ferroptosis in vivo.

cancer biology↗

bioRxiv2019

In vivo CRISPR screening identifies RNF20/40 as epigenetic regulators of cardiomyocyte maturation

Between birth and adulthood cardiomyocytes (CMs) undergo dramatic changes in size, ultrastructure, metabolism, and gene expression, in a process collectively referred to as CM maturation. The transcriptional network that coordinates CM maturation is poorly understood, creating a bottleneck for cardiac regenerative medicine. Forward genetic screens are a powerful, unbiased method to gain novel insights into transcriptional networks, yet this approach has rarely been used in vivo in mammals because of high resource demands. Here we utilized somatic mutagenesis to perform the first reported in vivo CRISPR genetic screen within a mammalian heart. We discovered and validated several novel transcriptional regulators of CM maturation. Among them were RNF20 and RNF40, which form a complex that monoubiquitinates H2B on lysine 120. Mechanistic studies indicated that this epigenetic mark controls dynamic changes in gene expression required for CM maturation. These insights into CM maturation will inform efforts in cardiac regenerative medicine. More broadly, our approach will enable unbiased forward genetics across mammalian organ systems.

genetics↗

bioRxiv2019

Targeting USP9x/SOX2 axis contributes to the anti-osteosarcoma effect of neogambogic acid

SOX2 has been viewed as a critical oncoprotein in osteosarcoma. Emerging evidence show that inducing the degradation of transcription factors such as SOX2 is a promising strategy to make them druggable. Here, we show that neogambogic acid (NGA), an active ingredients in garcinia, significantly inhibited the proliferation of osteosarcoma cells with ubiquitin proteasome-mediated degradation of SOX2 in vitro and in vivo. We further identified USP9x as a bona fide deubiquitinase for SOX2 and NGA directly interacts with USP9x in cells. Moreover, knockdown of USP9x inhibited the proliferation and colony formation of osteosarcoma cells, which could be rescued by overexpression of SOX2. Consistent with this, knockdown of USP9x inhibited the proliferation of osteosarcoma cells in a xenograft mouse model. Collectively, we identify USP9x as the first deubiquitinating enzyme for controlling the stability of SOX2 and USP9x is a direct target for NGA. We propose that targeting the USP9x/SOX2 axis represents a novel strategy for the therapeutic of osteosarcoma and other SOX2 related cancers.

cancer biology↗

bioRxiv2019

Genetic dissection of femoral and tibial microarchitecture

Our understanding of the genetic control of bone has relied almost exclusively on estimates of bone mineral density. In contrast, here we have used high-resolution x-ray tomography (8 m isotropic voxels) to measure femoral and tibial components across a set of ~600 mice belonging to 60 diverse BXD strains of mice. We computed heritabilities of 25 cortical and trabecular compartments. Males and females have well matched trait heritabilities, ranging from 0.25 to 0.75. We mapped 16 QTLs that collectively cover ~8% of all protein-coding genes in mouse. A majority of loci are detected only in females, and there is also a bias in favor of QTLs for cortical traits. To efficiently evaluate candidate genes we developed a method that couples gene ontologies with expression data to compute bone-enrichment scores for almost all protein-coding genes. We carefully collated and aligned murine candidates with recent human BMD genome-wide association results. We highlight a subset of 50 strong candidates that fall into three categories: 1. those linked to bone function that have already been experimentally validated (Adamts4, Ddr2, Darc, Adam12, Fkbp10, E2f6, Adam17, Grem2, Ifi204); 2. candidates with putative bone function but not yet tested (e.g., Greb1, Ifi202b) but several of which have been linked to phenotypes in humans; and 3. candidates that have high bone-enrichment scores but for which there is not yet any specific link to bone biology or skeletal disease, including Ifi202b, Ly9, Ifi205, Mgmt, F2rl1, Iqgap2. Our results highlight contrasting genetic architecture between the sexes and among major bone compartments. The joint use and alignment of murine and human data should greatly facilitate function analysis and preclinical testing.\n\nDisclosureThe authors declare that no competing interests exist.

genetics↗

bioRxiv2018

Effects of the anesthetic MS-222 on silver pomfret (Pampus argenteus) juveniles under aquaculture treatment stresses

The silver pomfret (Pampus argenteus) is a major economically important marine fish in China. However, P. argenteus is sensitive to many stress factors and susceptible to injury. This problem could be resolved using anesthesia. We determined the lowest effective dose (LED) of tricaine methanesulfonate (MS-222) and assessed the longest safe deep anesthesia time and effect after aquaculture treatment stresses. P. argenteus juveniles were exposed to six concentrations of MS-222 (10, 25, 50, 75, 100, and 125 mg L-1); LED was established at 75 mg L-1. The juveniles were exposed to different deep anesthesia times (4, 7, 10, 12, and 15 min) at 75 mg L-1; the longest safe deep anesthesia time under LED was 10 min. Finally, the juveniles were randomly divided into four groups: control group (CG), draining group (DG, drain), anesthetic group (AG, drain + MS-222 + aquaculture treatment); and non-anesthetic group (NAG, drain + aquaculture treatment). Plasma cortisol levels in the NAG, AG, DG, and CG groups were 38.739 {+/-} 1.065 (highest), 25.083 {+/-} 0.587, 28.644 {+/-} 0.612, and 22.620 {+/-} 0.836 ng mL-1 (lowest). The AG group showed significant differences in superoxide dismutase, catalase, and malondialdehyde activities, except for glutathione. HSP70, HSP90, GR1, and GR2 mRNA levels in the NAG group increased sharply in response to stressors. GR1 and GR2 mRNA levels in the AG group also increased significantly, whereas HSP70 and HSP90 mRNA levels showed no significant differences. Thus, MS-222 can reduce oxidative damage, stress reaction, and resistance to aquaculture treatment stresses in P. argenteus.

animal behavior and cognition↗

bioRxiv2018

Exogenous DCPTA ameliorates the soil drought effect on nitrogen metabolism in maize during the pre-female inflorescence emergence stage

Abstract2-(3,4-Dichlorophenoxy) triethylamine (DCPTA) regulates many aspects of plant development; however, its effects on soil drought tolerance are unknown. We pre-treated maize (Zea mays L.) by foliar application of DCPTA and subsequently exposed the plants to soil drought and rewatering conditions during the pre-female inflorescence emergence stage. Exogenous DCPTA significantly alleviated drought-induced decreases in maize yield, shoot and root relative growth rate (RGR), leaf relative water content (RLWC), net photosynthetic rate (Pn), stomatal conductance (Gs) and transpiration rate (Tr), nitrate (NO3-), nitrite (NO2-), and soluble protein contents, and nitrate reductase (NR), nitrite reductase (NiR), isocitrate dehydrogenase (ICDH), alanine aminotransferase (AlaAT) and aspartate aminotransferase (AspAT) activities; increases in the intercellular CO2 concentration (Ci), the ammonium (NH4+) and free amino acid contents, and the glutamate dehydrogenase (GDH) and protease activities. Simultaneously, exogenous DCPTA improved the spatial and temporal distribution of roots and increased the root hydraulic conductivity (Lp), flow rate of root-bleeding sap and NO3- delivery rates. Moreover, Exogenous DCPTA protected the chloroplast structure from drought injury. Taken together, our results suggest that exogenous DCPTA mitigates the repressive effects of drought on N metabolism and subsequently enhances drought tolerance during the pre-female inflorescence emergence stage of maize.\n\nHighlightsThis is the first article that explores the effects of DCPTA on nitrogen metabolism and the first article that explores the effects of DCPTA on crops under soil drought conditions.

physiology↗

bioRxiv2018

BodyMap transcriptomes reveal unique circular RNA features across tissue types and developmental stages

Circular RNAs (circRNAs) are a novel class of regulatory RNAs. Here, we present a comprehensive investigation of circRNA expression profiles across 11 tissues and 4 developmental stages in rats, along with cross-species analyses in humans and mice. Although positively correlated, circRNAs exhibit higher tissue specificity than cognate mRNAs. Also, genes with higher expression levels exhibit a larger fraction of spliced circular transcripts than their linear counterparts. Intriguingly, while we observed a monotonic increase of circRNA abundance with age in the rat brain, we further discovered a dynamic, age-dependent pattern of circRNA expression in the testes that is characterized by a dramatic increase with advancing stages of sexual maturity and a decrease with aging. The age-sensitive testicular circRNAs are highly associated with spermatogenesis, independent of cognate mRNA expression. The tissue/age implications of circRNAs suggest that they present unique physiological functions rather than simply occurring as occasional by-products of gene transcription.

bioinformatics↗