Ab-Externo AAV-Mediated Gene Delivery to the Suprachoroidal Space Using a 250 Micron Flexible Microcatheter

The current method of delivering gene replacement to the posterior segment of the eye involves a three-port pars plana vitrectomy followed by injection of the agent through a 37-gauge cannula, which is potentially wrought with retinal complications. In this paper we investigate the safety and efficacy of delivering adeno-associated viral (AAV) vector to the suprachoroidal space using an ab externo approach that utilizes an illuminated microcatheter.6 New Zealand White rabbits and 2 Dutch Belted rabbits were used to evaluate the ab externo delivery method. sc-AAV5-smCBA-hGFP vector was delivered into the suprachoroidal space using an illuminated iTrackTM 250A microcatheter. Six weeks after surgery, the rabbits were sacrificed and their eyes evaluated for AAV transfection using immunofluorescent antibody staining of GFP.Immunostaining of sectioned and whole-mounted eyes demonstrated robust transfection in all treated eyes, with no fluorescence in untreated control eyes. Transfection occurred diffusely and involved both the choroid and the retina. No apparent adverse effects caused by either the viral vector or the procedure itself could be seen either clinically or histologically.The ab externo method of delivery using a microcatheter was successful in safely and effectively delivering a gene therapy agent to the suprachoroidal space. This method presents a less invasive alternative to the current method of virally vectored gene delivery

AAV-Mediated Cone Rescue in a Naturally Occurring Mouse Model of CNGA3-Achromatopsia

Achromatopsia is a rare autosomal recessive disorder which shows color blindness, severely impaired visual acuity, and extreme sensitivity to bright light. Mutations in the alpha subunits of the cone cyclic nucleotide-gated channels (CNGA3) are responsible for about 1/4 of achromatopsia in the U.S. and Europe. Here, we test whether gene replacement therapy using an AAV5 vector could restore cone-mediated function and arrest cone degeneration in the cpfl5 mouse, a naturally occurring mouse model of achromatopsia with a CNGA3 mutation. We show that gene therapy leads to significant rescue of cone-mediated ERGs, normal visual acuities and contrast sensitivities. Normal expression and outer segment localization of both M- and S-opsins were maintained in treated retinas. The therapeutic effect of treatment lasted for at least 5 months post-injection. This study is the first demonstration of substantial, relatively long-term restoration of cone-mediated light responsiveness and visual behavior in a naturally occurring mouse model of CNGA3 achromatopsia. The results provide the foundation for development of an AAV5-based gene therapy trial for human CNGA3 achromatopsia

A neuron-in-capillary platform for facile collection and mass spectrometric characterization of a secreted neuropeptide

The integration of microfluidic devices-which efficiently handle small liquid volumes-with separations/mass spectrometry (MS) is an effective approach for profiling the neurochemistry occurring in selected neurons. Interfacing the microfluidic cell culture to the mass spectrometer is challenging because of geometric and scaling issues. Here we demonstrate the hyphenation of a neuron-in-capillary platform to a solid phase extraction device and off-line MS. A primary neuronal culture of Aplysia californica neurons was established directly inside a cylindrical polyimide capillary. The approach also uses a particle-embedded monolith to condition neuropeptide releasates collected from several Aplysia neurons cultured in the capillary, with the subsequent characterization of released peptides via MS. This system presents a number of advances compared to more traditional microfluidic devices fabricated with polydimethylsiloxane. These include low cost, easy access to cell culture, rigidity, ease of transport, and minimal fluid handling. The cylindrical geometry of the platform allows convenient interface with a wide range of analytical tools that utilize capillary columnsclos

A prospective study of changes in anxiety, depression, and problems in living during chemotherapy treatments: effects of age and gender

PurposeMonitoring distress assessment in cancer patients during the treatment phase is a component of good quality care practice. Yet, there is a dearth of prospective studies examining distress. In an attempt to begin filling this gap and inform clinical practice, we conducted a prospective, longitudinal study examining changes in distress (anxiety, depression, and problems in living) by age and gender and the roles of age and gender in predicting distress.MethodsNewly diagnosed Brazilian cancer patients (N = 548) were assessed at three time points during chemotherapy. Age and gender were identified on the first day of chemotherapy (T1); anxiety, depression, and problems in living were self-reported at T1, the planned midway point (T2), and the last day of chemotherapy (T3).ResultsAt T1, 37 and 17% of patients reported clinically significant levels of anxiety and depression, respectively. At T3, the prevalence was reduced to 4.6% for anxiety and 5.1% for depression (p < .001). Patients 40-55 years, across all time points, reported greater anxiety and practical problems than patients >70 years (p < .03). Female patients reported greater emotional, physical, and family problems than their male counterparts (p < .04).ConclusionsFor most patients, elevated levels of distress noted in the beginning of treatment subsided by the time of treatment completion. However, middle-aged and female patients continued to report heightened distress. Evidence-based psychosocial intervention offered to at risk patients during early phases of the treatment may provide distress relief and improve outcomes over the illness trajectory while preventing psychosocial and physical morbidity due to untreated chronic distress

Composition, diversity and structure of vascular epiphytes in two contrasting Central Amazonian floodplain ecosystems

Research focusing on assemblages of vascular epiphytes in the Amazon are scarce. This is especially true for Amazonian floodplain forests, for which only two previous studies have been published. We compared composition, richness and structure of epiphyte assemblages in white-water and black-water floodplains (várzea and igapó) in Central Amazonia in order to close knowledge gaps concerning the distribution and richness of epiphytes. We established sixteen 25x25 m plots in each forest type, and counted and identified all species of vascular epiphytes occurring on trees with a diameter at breast height (DBH) ≥10 cm. We observed a clear distinction in epiphytic species composition (r2=0.83, p=0.001) and diversity (t=3.24, P=0.003) between the two environments, with 61.5 % of species being restricted to várzea, 22.9 % restricted to igapó and only 15.6 % common to both ecosystems. The floodplains were also structurally different for the most abundant species and those with the highest Epiphytic Importance Value (IVe). The diversity of trees did not influence the epiphyte diversity in either ecosystem. The forests were found to differ in the composition, diversity and structure of their epiphytic assemblages, which must be taken into account when designing conservation action plans for these ecosystems and for their vascular epiphytes

Non-standard errors

In statistics, samples are drawn from a population in a data-generating process (DGP). Standard errors measure the uncertainty in estimates of population parameters. In science, evidence is generated to test hypotheses in an evidence generating process (EGP). We claim that EGP variation across researchers adds uncertainty: Non-standard errors (NSEs). We study NSEs by letting 164 teams test the same hypotheses on the same data. NSEs turn out to be sizable, but smaller for better reproducible or higher rated research. Adding peer-review stages reduces NSEs. We further find that this type of uncertainty is underestimated by participants

Evaluation of distributed recharge in an upland semi-arid karst system : the West Bank Mountain Aquifer, Middle East

Assessment of recharge in a structurally complex upland karst limestone aquifer situated in a semi-arid environment is difficult. Resort to surrogate indicators such as measurement of spring outflow and borehole discharge, is a common alternative, and attempts to apply conventional soil moisture deficit analysis may not adequately account for the intermittent spate conditions that arise in such environments. A modelling approach has been made using the West Bank Mountain Aquifer system in the Middle East as a trial. The model uses object oriented software which allows various objects to be switched on and off. Each of the main recharge processes identified in the West Bank is incorporated. The model allows either conventional soil moisture deficit analysis calculations or wetting threshold calculations to be made as appropriate, and accommodates both direct recharge and secondary recharge. Daily time steps enable recharge and runoff routing to be calculated for each node. Model runs have enabled a series of simulations for each of the three aquifer basins in the West Bank and for the whole of the West Bank. These provide recharge estimates comparable to those prepared by earlier workers by conventional means. The model is adaptable and has been successfully used in other environments

Consistency in performance rankings: the Peru water sector

Regulators can utilize a number of alternative methodologies for comparing firm efficiency, but these approaches need to be robust to be accepted by stakeholders. This study evaluates the consistency of water-utility performance rankings for Peruvian water utilities. The results indicate that data envelopment analysis (DEA) and stochastic frontier analysis (SFA) yield similar rankings in this case. In addition, the techniques have comparable success for identifying the best and worst performing utilities. However, these rankings based on sophisticated statistical techniques are not highly correlated with those developed by the Peruvian water regulator (SUNASS). This result does not invalidate the performance rankings obtained by the regulator, since those rankings are based on more dimensions of utility performance. However, they illustrate the importance of developing sound techniques for identifying weak utilities. Improvements in sector performance require that benchmarking be given greater attention than in the past.