171 research outputs found
Molecular analysis of selected cell cycle regulatory proteins during aerobic and hypoxic maintenance of human ovarian carcinoma cells
Activation of PyMT in Ξ² Cells Induces Irreversible Hyperplasia, but Oncogene-Dependent Acinar Cell Carcinomas When Activated in Pancreatic Progenitors
It is unclear whether the cellular origin of various forms of pancreatic cancer involves transformation or transdifferentiation of different target cells or whether tumors arise from common precursors, with tumor types determined by the specific genetic alterations. Previous studies suggested that pancreatic ductal carcinomas might be induced by polyoma middle T antigen (PyMT) expressed in non-ductal cells. To ask whether PyMT transforms and transdifferentiates endocrine cells toward exocrine tumor phenotypes, we generated transgenic mice that carry tetracycline-inducible PyMT and a linked luciferase reporter. Induction of PyMT in Ξ² cells causes Ξ²-cell hyperplastic lesions that do not progress to malignant neoplasms. When PyMT is de-induced, Ξ² cell proliferation and growth cease; however, regression does not occur, suggesting that continued production of PyMT is not required to maintain the viable expanded Ξ² cell population. In contrast, induction of PyMT in early pancreatic progenitor cells under the control of Pdx1 produces acinar cell carcinomas and Ξ²-cell hyperplasia. The survival of acinar tumor cells is dependent on continued expression of PyMT. Our findings indicate that PyMT can induce exocrine tumors from pancreatic progenitor cells, but cells in the Ξ² cell lineage are not transdifferentiated toward exocrine cell types by PyMT; instead, they undergo oncogene-dependent hyperplastic growth, but do not require PyMT for survival
Cell cycle-dependent phosphorylation of pRb-like protein in root meristem cells of Vicia faba
The retinoblastoma tumor suppressor protein (pRb) regulates cell cycle progression by controlling the G1-to-S phase transition. As evidenced in mammals, pRb has three functionally distinct binding domains and interacts with a number of proteins including the E2F family of transcription factors, proteins with a conserved LxCxE motif (D-type cyclin), and c-Abl tyrosine kinase. CDK-mediated phosphorylation of pRb inhibits its ability to bind target proteins, thus enabling further progression of the cell cycle. As yet, the roles of pRb and pRb-binding factors have not been well characterized in plants. By using antibody which specifically recognizes phosphorylated serines (S807/811) in the c-Abl tyrosine kinase binding C-domain of human pRb, we provide evidence for the cell cycle-dependent changes in pRb-like proteins in root meristems cells of Vicia faba. An increased phosphorylation of this protein has been found correlated with the G1-to-S phase transition
Vesnarinone and glucocorticoids cooperatively induce G1 arrest and have an anti-tumour effect on human non-small cell lung carcinoma cells grown in nude mice
A qualitative study exploring factors associated with mothersβ decisions to formula-feed their infants in Newfoundland and Labrador, Canada
Bicistronic Lentiviruses Containing a Viral 2A Cleavage Sequence Reliably Co-Express Two Proteins and Restore Vision to an Animal Model of LCA1
The disease processes underlying inherited retinal disease are complex and are not completely understood. Many of the corrective gene therapies designed to treat diseases linked to mutations in genes specifically expressed in photoreceptor cells restore function to these cells but fail to stop progression of the disease. There is growing consensus that effective treatments for these diseases will require delivery of multiple therapeutic proteins that will be selected to treat specific aspects of the disease process. The purpose of this study was to design a lentiviral transgene that reliably expresses all of the proteins it encodes and does so in a consistent manner among infected cells. We show, using both in vitro and in vivo analyses, that bicistronic lentiviral transgenes encoding two fluorescent proteins fused to a viral 2A-like cleavage peptide meet these expression criteria. To determine if this transgene design is suitable for therapeutic applications, we replaced one of the fluorescent protein genes with the gene encoding guanylate cyclase -1 (GC1) and delivered lentivirus carrying this transgene to the retinas of the GUCY1*B avian model of Leber congenital amaurosis β 1 (LCA1). GUCY1*B chickens carry a null mutation in the GC1 gene that disrupts photoreceptor function and causes blindness at hatching, a phenotype that closely matches that observed in humans with LCA1. We found that treatment of these animals with the 2A lentivector encoding GC1 restored vision to these animals as evidenced by the presence of optokinetic reflexes. We conclude that 2A-like peptides, with proper optimization, can be successfully incorporated into therapeutic vectors designed to deliver multiple proteins to neural retinal. These results highlight the potential of this vector design to serve as a platform for the development of combination therapies designed to enhance or prolong the benefits of corrective gene therapies
Piezo1 integration of vascular architecture with physiological force
The mechanisms by which physical forces regulate endothelial cells to determine the complexities of vascular structure and function are enigmaticΒΉβ»β΅. Studies of sensory neurons have suggested Piezo proteins as subunits of CaΒ²βΊ-permeable non-selective cationic channels for detection of noxious mechanical impactβΆβ»βΈ. Here we show Piezo1 (Fam38a) channels as sensors of frictional force (shear stress) and determinants of vascular structure in both development and adult physiology. Global or endothelial-specific disruption of mouse Piezo1 profoundly disturbed the developing vasculature and was embryonic lethal within days of the heart beating. Haploinsufficiency was not lethal but endothelial abnormality was detected in mature vessels. The importance of Piezo1 channels as sensors of blood flow was shown by Piezo1 dependence of shear-stress-evoked ionic current and calcium influx in endothelial cells and the ability of exogenous Piezo1 to confer sensitivity to shear stress on otherwise resistant cells. Downstream of this calcium influx there was protease activation and spatial reorganization of endothelial cells to the polarity of the applied force. The data suggest that Piezo1 channels function as pivotal integrators in vascular biology
Co-Regulation of NF-ΞΊB and Inflammasome-Mediated Inflammatory Responses by Myxoma Virus Pyrin Domain-Containing Protein M013
NF-ΞΊB and inflammasomes both play central roles in orchestrating anti-pathogen responses by rapidly inducing a variety of early-response cytokines and chemokines following infection. Myxoma virus (MYXV), a pathogenic poxvirus of rabbits, encodes a member of the cellular pyrin domain (PYD) superfamily, called M013. The viral M013 protein was previously shown to bind host ASC-1 protein and inhibit the cellular inflammasome complex that regulates the activation and secretion of caspase 1-regulated cytokines such as IL-1Ξ² and IL-18. Here, we report that human THP-1 monocytic cells infected with a MYXV construct deleted for the M013L gene (vMyxM013-KO), in stark contrast to the parental MYXV, rapidly induce high levels of secreted pro-inflammatory cytokines like TNF, IL-6, and MCP-1, all of which are regulated by NF-ΞΊB. The induction of these NF-ΞΊB regulated cytokines following infection with vMyxM013-KO was also confirmed in vivo using THP-1 derived xenografts in NOD-SCID mice. vMyxM013-KO virus infection specifically induced the rapid phosphorylation of IKK and degradation of IΞΊBΞ±, which was followed by nuclear translocation of NF-ΞΊB/p65. Even in the absence of virus infection, transiently expressed M013 protein alone inhibited cellular NF-ΞΊB-mediated reporter gene expression and nuclear translocation of NF-ΞΊB/p65. Using protein/protein interaction analysis, we show that M013 protein also binds directly with cellular NF-ΞΊB1, suggesting a direct physical and functional linkage between NF-ΞΊB1 and ASC-1. We further demonstrate that inhibition of the inflammasome with a caspase-1 inhibitor did not prevent the induction of NF-ΞΊB regulated cytokines following infection with vMyxM013-KO virus, but did block the activation of IL-1Ξ². Thus, the poxviral M013 inhibitor exerts a dual immuno-subversive role in the simultaneous co-regulation of both the cellular inflammasome complex and NF-ΞΊB-mediated pro-inflammatory responses
Weak Responses to Auditory Feedback Perturbation during Articulation in Persons Who Stutter: Evidence for Abnormal Auditory-Motor Transformation
Previous empirical observations have led researchers to propose that auditory feedback (the auditory perception of self-produced sounds when speaking) functions abnormally in the speech motor systems of persons who stutter (PWS). Researchers have theorized that an important neural basis of stuttering is the aberrant integration of auditory information into incipient speech motor commands. Because of the circumstantial support for these hypotheses and the differences and contradictions between them, there is a need for carefully designed experiments that directly examine auditory-motor integration during speech production in PWS. In the current study, we used real-time manipulation of auditory feedback to directly investigate whether the speech motor system of PWS utilizes auditory feedback abnormally during articulation and to characterize potential deficits of this auditory-motor integration. Twenty-one PWS and 18 fluent control participants were recruited. Using a short-latency formant-perturbation system, we examined participantsβ compensatory responses to unanticipated perturbation of auditory feedback of the first formant frequency during the production of the monophthong [Ξ΅]. The PWS showed compensatory responses that were qualitatively similar to the controlsβ and had close-to-normal latencies (~150 ms), but the magnitudes of their responses were substantially and significantly smaller than those of the control participants (by 47% on average, p<0.05). Measurements of auditory acuity indicate that the weaker-than-normal compensatory responses in PWS were not attributable to a deficit in low-level auditory processing. These findings are consistent with the hypothesis that stuttering is associated with functional defects in the inverse models responsible for the transformation from the domain of auditory targets and auditory error information into the domain of speech motor commands
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