We describe an automated method to locate the optic nerve in images of the ocular fundus. Our method uses a novel algorithm we call fuzzy convergence to determine the origination of the blood vessel ...network. We evaluate our method using 31 images of healthy retinas and 50 images of diseased retinas, containing such diverse symptoms as tortuous vessels, choroidal neovascularization, and hemorrhages that completely obscure the actual nerve. On this difficult data set, our method achieved 89% correct detection. We also compare our method against three simpler methods, demonstrating the performance improvement. All our images and data are freely available for other researchers to use in evaluating related methods.
Longitudinal studies of chronic wasting disease (CWD) in the native host have provided considerable understanding of how this prion disease continues to efficiently spread among cervid species. These ...studies entail great cost in animal, time and financial support. A variety of methods have emerged including transgenic mouse bioassay, western blot, enzyme-linked immunoassay (ELISA), immunohistochemistry (IHC), serial protein misfolding cyclic amplification (sPMCA) and real time quaking-induced conversion (RT-QuIC), that deepen our understanding of this and other protein misfolding disorders. To further characterize an inoculum source used for ongoing CWD studies and to determine how the readouts from each of these assays compare, we assayed a CWD-positive brain pool homogenate (CBP6) and a mouse dilutional bioassay of this homogenate using the above detection methods. We demonstrate that: (i) amplification assays enhanced detection of amyloid seeding activity in the CWD+ cervid brain pool to levels beyond mouse LD50, (ii) conventional detection methods (IHC and western blot) performed well in identifying the presence of PrPSc in terminal brain tissue yet lack sufficient detection sensitivity to identify all CWD-infected mice, and (iii) the incorporation of amplification assays enhanced detection of CWD-infected mice near the LD50. This cross-platform analysis provides a basis to calibrate the relative sensitivities of CWD detection assays.
The agent responsible for prion diseases is a misfolded form of a normal protein (PrPC). The prion hypothesis stipulates that PrPC must be present for the disease to manifest. Cervid populations ...across the world are infected with chronic wasting disease, a horizontally-transmissible prion disease that is likely spread via oral exposure to infectious prions (PrPCWD). Though PrPCWD has been identified in many tissues, there has been little effort to characterize the overall PrPC expression in cervids and its relationship to PrPCWD accumulation. We used immunohistochemistry (IHC), western blot and enzyme-linked immunosorbent assay to describe PrPC expression in naïve white-tailed deer. We used real-time, quaking-induced conversion (RT-QuIC) to detect prion seeding activity in CWD-infected deer. We assessed tissues comprising the alimentary tract, alimentary-associated lymphoid tissue and systemic lymphoid tissue from 5 naïve deer. PrPC was expressed in all tissues, though expression was often very low compared to the level in the CNS. IHC identified specific cell types wherein PrPC expression is very high. To compare the distribution of PrPC to PrPCWD, we examined 5 deer with advanced CWD infection. Using RT-QuIC, we detected prion seeding activity in all 21 tissues. In 3 subclinical deer sacrificed 4 months post-inoculation, we detected PrPCWD consistently in alimentary-associated lymphoid tissue, irregularly in alimentary tract tissues, and not at all in the brain. Contrary to our hypothesis that PrPC levels dictate prion accumulation, PrPC expression was higher in the lower gastrointestinal tissues than in the alimentary-associated lymphoid system and was higher in salivary glands than in the oropharyngeal lymphoid tissue. These data suggest that PrPC expression is not the sole driver of prion accumulation and that alimentary tract tissues accumulate prions before centrifugal spread from the brain occurs.
The minimum infectious dose required to induce CWD infection in cervids remains unknown, as does whether peripherally shed prions and/or multiple low dose exposures are important factors in CWD ...transmission. With the goal of better understand CWD infection in nature, we studied oral exposures of deer to very low doses of CWD prions and also examined whether the frequency of exposure or prion source may influence infection and pathogenesis. We orally inoculated white-tailed deer with either single or multiple divided doses of prions of brain or saliva origin and monitored infection by serial longitudinal tissue biopsies spanning over two years. We report that oral exposure to as little as 300 nanograms (ng) of CWD-positive brain or to saliva containing seeding activity equivalent to 300 ng of CWD-positive brain, were sufficient to transmit CWD disease. This was true whether the inoculum was administered as a single bolus or divided as three weekly 100 ng exposures. However, when the 300 ng total dose was apportioned as 10, 30 ng doses delivered over 12 weeks, no infection occurred. While low-dose exposures to prions of brain or saliva origin prolonged the time from inoculation to first detection of infection, once infection was established, we observed no differences in disease pathogenesis. These studies suggest that the CWD minimum infectious dose approximates 100 to 300 ng CWD-positive brain (or saliva equivalent), and that CWD infection appears to conform more with a threshold than a cumulative dose dynamic.
Mechanisms of Taxane Resistance Maloney, Sara M; Hoover, Camden A; Morejon-Lasso, Lorena V ...
Cancers,
11/2020, Letnik:
12, Številka:
11
Journal Article
Recenzirano
Odprti dostop
The taxane family of chemotherapy drugs has been used to treat a variety of mostly epithelial-derived tumors and remain the first-line treatment for some cancers. Despite the improved survival time ...and reduction of tumor size observed in some patients, many have no response to the drugs or develop resistance over time. Taxane resistance is multi-faceted and involves multiple pathways in proliferation, apoptosis, metabolism, and the transport of foreign substances. In this review, we dive deeper into hypothesized resistance mechanisms from research during the last decade, with a focus on the cancer types that use taxanes as first-line treatment but frequently develop resistance to them. Furthermore, we will discuss current clinical inhibitors and those yet to be approved that target key pathways or proteins and aim to reverse resistance in combination with taxanes or individually. Lastly, we will highlight taxane response biomarkers, specific genes with monitored expression and correlated with response to taxanes, mentioning those currently being used and those that should be adopted. The future directions of taxanes involve more personalized approaches to treatment by tailoring drug-inhibitor combinations or alternatives depending on levels of resistance biomarkers. We hope that this review will identify gaps in knowledge surrounding taxane resistance that future research or clinical trials can overcome.
The diagnosis of chronic wasting disease (CWD) relies on demonstration of the disease-associated misfolded CWD prion protein (PrPCWD) in brain or retropharyngeal lymph node tissue by immunodetection ...methods, e.g. ELISA and immunohistochemistry (IHC). The success of these methods relies on a quality sample of tissues, which requires both anatomical knowledge and considerable dissection to collect. As the prevalence of CWD continues to increase globally, the development of fast and cost-effective methods to detect the disease is vital to facilitate CWD detection and surveillance. To address these issues, we have evaluated third eyelids from CWD-infected deer and elk using real-time quaking induced conversion (RT-QuIC). We identified prion seeding activity in third eyelids in 24 of 25 (96%) CWD-infected white-tailed deer (Odocoileus virginianus). We detected RT-QuIC positivity in the third eyelid as early as 1 month after experimental CWD exposure. In addition, we identified prion seeding activity in third eyelids of 18 of 25 (72%) naturally exposed asymptomatic CWD-positive rocky mountain elk (Cervus canadensis nelson). We compared CWD detection by RT-QuIC and IHC in third eyelid, retropharyngeal lymph node, and brain in 10 deer in early symptomatic stage of disease. IHC detected PrPCWD deposition in third eyelid lymphoid follicles in 5 of 10 deer (50%) whereas third eyelids of all 10 animals were positive by RT-QuIC. This difference reflected in part a lower requirement for lymphoid follicle presence for seeding activity detection by RT-QuIC. In conclusion, RT-QuIC analysis of the third eyelid, an easily accessed tissue, has potential to advance CWD detection and testing compliance.
CWD is an emergent prion disease that now affects cervid species on three continents. CWD is efficiently spread in wild and captive populations, likely through both direct animal contact and ...environmental contamination. Here, by longitudinally assaying in feces of CWD-exposed white-tailed deer by RT-QuIC, we demonstrate fecal shedding of prion seeding activity months before onset of clinical symptoms and continuing throughout the disease course. We also examine the impact of simulated environmental conditions such as repeated freeze-thaw cycles and desiccation on fecal prion seeding activity. We found that while multiple (n = 7) freeze-thaw cycles substantially decreased fecal seeding activity, desiccation had little to no effect on seeding activity. Finally, we examined whether RT-QuIC testing of landscape fecal deposits could distinguish two premises with substantial known CWD prevalence from one in which no CWD-infected animals had been detected. In the above pilot study, this distinction was possible. We conclude that fecal shedding of CWD prions occurs over much of the disease course, that environmental factors influence prion seeding activity, and that it is feasible to detect fecal prion contamination using RT-QuIC.
Canine parvovirus (CPV) emerged as a new pandemic pathogen of dogs in the 1970s and is closely related to feline panleukopenia virus (FPV), a parvovirus of cats and related carnivores. Although both ...viruses have wide host ranges, analysis of viral sequences recovered from different wild carnivore species, as shown here, demonstrated that>95% were derived from CPV-like viruses, suggesting that CPV is dominant in sylvatic cycles. Many viral sequences showed host-specific mutations in their capsid proteins, which were often close to sites known to control binding to the transferrin receptor (TfR), the host receptor for these carnivore parvoviruses, and which exhibited frequent parallel evolution. To further examine the process of host adaptation, we passaged parvoviruses with alternative backgrounds in cells from different carnivore hosts. Specific mutations were selected in several viruses and these differed depending on both the background of the virus and the host cells in which they were passaged. Strikingly, these in vitro mutations recapitulated many specific changes seen in viruses from natural populations, strongly suggesting they are host adaptive, and which were shown to result in fitness advantages over their parental virus. Comparison of the sequences of the transferrin receptors of the different carnivore species demonstrated that many mutations occurred in and around the apical domain where the virus binds, indicating that viral variants were likely selected through their fit to receptor structures. Some of the viruses accumulated high levels of variation upon passage in alternative hosts, while others could infect multiple different hosts with no or only a few additional mutations. Overall, these studies demonstrate that the evolutionary history of a virus, including how long it has been circulating and in which hosts, as well as its phylogenetic background, has a profound effect on determining viral host range.
Pressure retarded osmosis has the potential to produce renewable energy from natural salinity gradients. This work presents the fabrication of thin-film composite membranes customized for high ...performance in pressure retarded osmosis. We also present the development of a theoretical model to predict the water flux in pressure retarded osmosis, from which we can predict the power density that can be achieved by a membrane. The model is the first to incorporate external concentration polarization, a performance limiting phenomenon that becomes significant for high-performance membranes. The fabricated membranes consist of a selective polyamide layer formed by interfacial polymerization on top of a polysulfone support layer made by phase separation. The highly porous support layer (structural parameter S = 349 μm), which minimizes internal concentration polarization, allows the transport properties of the active layer to be customized to enhance PRO performance. It is shown that a hand-cast membrane that balances permeability and selectivity (A = 5.81 L m(-2) h(-1) bar(-1), B = 0.88 L m(-2) h(-1)) is projected to achieve the highest potential peak power density of 10.0 W/m(2) for a river water feed solution and seawater draw solution. The outstanding performance of this membrane is attributed to the high water permeability of the active layer, coupled with a moderate salt permeability and the ability of the support layer to suppress the undesirable accumulation of leaked salt in the porous support. Membranes with greater selectivity (i.e., lower salt permeability, B = 0.16 L m(-2) h(-1)) suffered from a lower water permeability (A = 1.74 L m(-2) h(-1) bar(-1)) and would yield a lower peak power density of 6.1 W/m(2), while membranes with a higher permeability and lower selectivity (A = 7.55 L m(-2) h(-1) bar(-1), B = 5.45 L m(-2) h(-1)) performed poorly due to severe reverse salt permeation, resulting in a similar projected peak power density of 6.1 W/m(2).
Re-engineering the support layers of membranes for forward and pressure retarded osmosis is critical for making these technologies commercially viable. Real-world applications of forward and pressure ...retarded osmosis, especially those involving natural and waste waters, will require membranes to withstand significant stresses. Therefore, structural changes to the support layer, which are necessary in minimizing internal concentration polarization, must not compromise its critical abilities to resist mechanical stress and provide a suitable surface for the interfacial polymerization of a robust and selective active layer. Electrospinning can provide nanofibers for support layers to potentially overcome the limitations of traditional membrane fabrication techniques in fulfilling these challenging design criteria. In this work, we present the fabrication and evaluation of thin-film composite membranes composed of electrospun polyethylene terephthalate nanofibers, a phase separation formed microporous polysulfone layer, and a polyamide selective layer formed by interfacial polymerization. These membranes have active and support layer transport properties that are suitable for engineered osmosis, with water permeability of 1.13Lm−2h−1bar−1 (3.14×10−7ms−1bar−1), salt permeability of 0.23Lm−2h−1 (6.4×10−8ms−1), and a structural parameter of 651μm. Relevant and easily reproducible tests for membrane resistance to mechanical stress were performed. The use of electrospun fibers in the support layer enhanced membrane resistance to delamination at high cross-flow velocities because the 340nm diameter electrospun fibers enmesh with the microporous polysulfone layer. A broader discussion of the most promising approaches for using electrospun materials to improve membranes for engineered osmosis is provided.
► Thin-film composite membranes for forward and pressure retarded osmosis were fabricated. ► Electrospun mats served as a base onto which a support layer was formed by phase separation. ► Thin-film composite membranes had high salt rejection and high water flux in forward osmosis. ► Membrane resistance to shear stress and hydraulic pressure was evaluated. ► Nanofibers enmeshed with microporous layer to enhance resistance to delamination.
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