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Gould, E. A.

Publications and source records attributed to Gould, E. A..

2 recordsLinked to original sources

Differential Expression of Mucins in Murine Olfactory Versus Respiratory Epithelium

Mucins are a key component of the airway surface liquid and serve many functions. Given the numerous differences in olfactory versus respiratory nasal epithelia, we hypothesized that mucins would be differentially expressed between these two areas. Secondarily, we evaluated for changes in mucin expression with radiation exposure, given the clinical observations of nasal dryness, altered mucus rheology, and smell loss in radiated patients. Immunofluorescence staining was performed in a mouse model to determine the expression of mucins 1, 2, 5AC and 5B in nasal respiratory and olfactory epithelia of control mice and one week after exposure to 8 gy of radiation. Mucins 1, 5AC and 5B exhibited differential expression between olfactory and respiratory epithelium while mucin 2 showed no difference. Within the olfactory epithelium, mucin 1 was located in a lattice-like pattern around gaps corresponding to dendritic knobs of olfactory sensory neurons, whereas in respiratory epithelium it was only intermittently expressed. Mucin 5AC was expressed by subepithelial glands in both epithelial types but to a higher degree in the olfactory epithelium. Mucin 5B was expressed by submucosal glands in the olfactory epithelium but by surface epithelial cells in respiratory epithelium. At one-week after exposure to single-dose 8 gy of radiation, no qualitative effects were seen on mucin expression. Our findings demonstrate that murine olfactory and respiratory epithelia express mucins differently, and characteristic patterns of mucins 1, 5AC, and 5B can be used to define the underlying epithelium. Radiation (8 gy) does not appear to affect mucin expression at one week.\n\nAuthor RolesChristopher Kennel conceived, organized and executed the study, performed the analysis, and contributed to the manuscript.\n\nElizabeth Gould conceived and executed the study, and contributed to the manuscript.\n\nDiego Restrepo conceived and executed the study, supervised the experiments, reviewed the analysis, and contributed to the manuscript.\n\nErnesto Salcedo performed experiments and reviewed the manuscript.\n\nThad Vickery performed experiments and reviewed the manuscript.\n\nEric Larson performed experiments and reviewed the manuscript.\n\nVijay Ramakrishnan conceived and executed the study, reviewed the analysis, and contributed to the manuscript.\n\nAll authors discussed the results and implications and contributed to the final manuscript.

neuroscience

An E460D substitution in the NS5 protein of tick-borne encephalitis virus confers resistance to the inhibitor Galidesivir (BCX4430) and also attenuates the virus for mice

The adenosine analogue Galidesivir (BCX4430), a broad-spectrum RNA virus inhibitor, has entered a Phase 1 clinical safety and pharmacokinetics study in healthy subjects and is under clinical development for treatment of Ebola virus infection. Moreover, Galidesivir also inhibits the reproduction of tick-borne encephalitis virus (TBEV) and numerous other medically important flaviviruses. Until now, studies of this antiviral agent have not yielded resistant viruses. Here, we demonstrate that an E460D substitution, in the active site of TBEV RNA-dependent-RNA-polymerase (RdRp), confers resistance to Galidesivir in cell culture. Stochastic molecular simulations indicate that the steric freedom caused by the E460D substitution increases close electrostatic interactions between the inhibitor and the interrogation residue of the TBEV RdRp motif F, resulting in rejection of the analogue as an incorrect/modified nucleotide. Galidesivir-resistant TBEV exhibited no cross-resistance to structurally different antiviral nucleoside analogues, such as 7-deaza-2-C-methyladenosine, 2-C-methyladenosine and 4-azido-aracytidine. Although, the E460D substitution led only to a subtle decrease in viral fitness in cell culture, Galidesivir-resistant TBEV was highly attenuated in vivo, with 100% survival rate and no clinical signs observed in infected mice. Our results contribute to understanding the molecular basis of Galidesivir antiviral activity, flavivirus resistance to nucleoside inhibitors and the potential contribution of viral RdRp to flavivirus neurovirulence. ImportanceTick-borne encephalitis virus (TBEV) is a pathogen that causes severe human neuroinfections in large areas of Europe and Asia and for which there is currently no specific therapy. We have previously found that Galidesivir (BCX4430), a broad-spectrum RNA virus inhibitor, which is under clinical development for treatment of Ebola virus infection, has a strong antiviral effect against TBEV. For any antiviral drug, it is important to generate drug-resistant mutants to understand how the drug works. Here, we produced TBEV mutants resistant to Galidesivir and found that the resistance is caused by a single amino acid substitution in an active site of the viral RNA-dependent RNA polymerase, an enzyme which is crucial for replication of viral RNA genome. Although, this substitution led only to a subtle decrease in viral fitness in cell culture, Galidesivir-resistant TBEV was highly attenuated in a mouse model. Our results contribute to understanding the molecular basis of Galidesivir antiviral activity.

microbiology