Noninferiority testing with censoring when the event rate is low.

The PREDICT TB trial tests noninferiority of an abbreviated treatment regimen (arm A) vs a conventional treatment regimen (arm C). Treatment trials of drug-susceptible tuberculosis are expected to have low event rates (ie, relapse probabilities around 3-5%). We examine the question of what is the "best" way to test for noninferiority in a setting with low event rates. In a series of simulations supported by theoretical arguments, we examine operating characteristics of five tests, including normal approximation, exact, and simulation-based tests. Two of these tests are constructed from Kaplan-Meier based-estimators, which account for variable follow-up time (and those lost to follow-up). We evaluate the effect of loss to follow-up via simulations. We also examine the results of the five tests on a data set similar to PREDICT TB, the REMoxTB trial. We find that the normal approximation tests perform well, albeit with small type I error rate inflation. We also find that the Kaplan-Meier methods generally have larger power than the other tests, especially when there is between 10-30% loss to follow-up.

Remdesivir for the Prevention of Invasive Mechanical Ventilation or Death in Coronavirus Disease 2019 (COVID-19): A Post Hoc Analysis of the Adaptive COVID-19 Treatment Trial-1 Cohort Data.

This post hoc analysis of the Adaptive Coronavirus Disease 2019 (COVID-19) Treatment Trial-1 (ACTT-1) shows a treatment effect of remdesivir (RDV) on progression to invasive mechanical ventilation (IMV) or death. Additionally, we create a risk profile that better predicts progression than baseline oxygen requirement alone. The highest risk group derives the greatest treatment effect from RDV.

Bliss' and Loewe's additive and synergistic effects in Plasmodium falciparum growth inhibition by AMA1-RON2L, RH5, RIPR and CyRPA antibody combinations.

Plasmodium invasion of red blood cells involves malaria proteins, such as reticulocyte-binding protein homolog 5 (RH5), RH5 interacting protein (RIPR), cysteine-rich protective antigen (CyRPA), apical membrane antigen 1 (AMA1) and rhoptry neck protein 2 (RON2), all of which are blood-stage malaria vaccine candidates. So far, vaccines containing AMA1 alone have been unsuccessful in clinical trials. However, immunization with AMA1 bound with RON2L (AMA1-RON2L) induces better protection against P. falciparum malaria in Aotus monkeys. We therefore sought to determine whether combinations of RH5, RIPR, CyRPA and AMA1-RON2L antibodies improve their biological activities and sought to develop a robust method for determination of synergy or additivity in antibody combinations. Rabbit antibodies against AMA1-RON2L, RH5, RIPR or CyRPA were tested either alone or in combinations in P. falciparum growth inhibition assay to determine Bliss' and Loewe's additivities. The AMA1-RON2L/RH5 combination consistently demonstrated an additive effect while the CyRPA/RIPR combination showed a modest synergistic effect with Hewlett's [Formula: see text] Additionally, we provide a publicly-available, online tool to aid researchers in analyzing and planning their own synergy experiments. This study supports future blood-stage vaccine development by providing a solid methodology to evaluate additive and/or synergistic (or antagonistic) effect of vaccine-induced antibodies.

µ-Opioid Receptor Activation Directly Modulates Intrinsically Photosensitive Retinal Ganglion Cells.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) encode light intensity and trigger reflexive responses to changes in environmental illumination. In addition to functioning as photoreceptors, ipRGCs are post-synaptic neurons in the inner retina, and there is increasing evidence that their output can be influenced by retinal neuromodulators. Here we show that opioids can modulate light-evoked ipRGC signaling, and we demonstrate that the M1, M2 and M3 types of ipRGCs are immunoreactive for µ-opioid receptors (MORs) in both mouse and rat. In the rat retina, application of the MOR-selective agonist DAMGO attenuated light-evoked firing ipRGCs in a dose-dependent manner (IC50 < 40 nM), and this effect was reversed or prevented by co-application of the MOR-selective antagonists CTOP or CTAP. Recordings from solitary ipRGCs, enzymatically dissociated from retinas obtained from melanopsin-driven fluorescent reporter mice, confirmed that DAMGO exerts its effect directly through MORs expressed by ipRGCs. Reduced ipRGC excitability occurred via modulation of voltage-gated potassium and calcium currents. These findings suggest a potential new role for endogenous opioids in the mammalian retina and identify a novel site of action-MORs on ipRGCs-through which opioids might exert effects on reflexive responses to environmental light.

Retinal ganglion cell distribution and visual acuity in alpacas (Vicugna pacos).

OBJECTIVE: To investigate the distribution of retinal ganglion cells (RGCs) and visual acuity in alpacas (Vicugna pacos) through Brn-3a immunofluorescent labeling. PROCEDURES: Five eyes from four healthy alpacas with normal ophthalmic examination findings were included in the study. The axial length of the globes was measured before fixation. All five retinas were treated with Brn-3a antibodies to label RGCs. Images taken with a fluorescent microscope were used for RGC counting. RGC density maps were reconstructed by computer software. Visual acuity was estimated based on the results of peak RGC density and ocular anatomical parameters. RESULTS: The reconstructed retinal maps from Brn-3a labeling showed a horizontal streak across the retinal meridian superior to the optic nerve head with a temporal, upward extension. The highest RGC densities were in the temporal retinas. The maximal visual acuity was located in the temporal retina and was estimated to range between 12.5 and 13.4 cycles per degree. CONCLUSIONS: Alpacas have a horizontal streak across the retinal meridian superior to the optic disk with a temporal, upward extension based on the Brn-3a labeling of RGCs. The maximal visual acuity was located in the temporal retina. The reconstructed retinal maps indicate the RGC topography of alpacas is similar to that of other herbivores, but is different from that of dromedary camels.

Dopaminergic amacrine cells express opioid receptors in the mouse retina.

The presence of opioid receptors has been confirmed by a variety of techniques in vertebrate retinas including those of mammals; however, in most reports, the location of these receptors has been limited to retinal regions rather than specific cell types. Concurrently, our knowledge of the physiological functions of opioid signaling in the retina is based on only a handful of studies. To date, the best-documented opioid effect is the modulation of retinal dopamine release, which has been shown in a variety of vertebrate species. Nonetheless, it is not known if opioids can affect dopaminergic amacrine cells (DACs) directly, via opioid receptors expressed by DACs. This study, using immunohistochemical methods, sought to determine whether (1) µ- and δ-opioid receptors (MORs and DORs, respectively) are present in the mouse retina, and if present, (2) are they expressed by DACs. We found that MOR and DOR immunolabeling were associated with multiple cell types in the inner retina, suggesting that opioids might influence visual information processing at multiple sites within the mammalian retinal circuitry. Specifically, colabeling studies with the DAC molecular marker anti-tyrosine hydroxylase antibody showed that both MOR and DOR immunolabeling localize to DACs. These findings predict that opioids can affect DACs in the mouse retina directly, via MOR and DOR signaling, and might modulate dopamine release as reported in other mammalian and nonmammalian retinas.

beta-Endorphin expression in the mouse retina.

Evidence showing expression of endogenous opioids in the mammalian retina is sparse. In the present study we examined a transgenic mouse line expressing an obligate dimerized form of Discosoma red fluorescent protein (DsRed) under the control of the pro-opiomelanocortin promoter and distal upstream regulatory elements to assess whether pro-opiomelanocortin peptide (POMC), and its opioid cleavage product, beta-endorphin, are expressed in the mouse retina. Using double label immunohistochemistry we found that DsRed fluorescence was restricted to a subset of GAD-67-positive cholinergic amacrine cells of both orthotopic and displaced subtypes. About 50% of cholinergic amacrine cells colocalized DsRed and a large fraction of DsRed-expressing amacrine cells was positive for beta-endorphin immunostaining, whereas beta-endorphin-immunoreactive neurons were absent in retinas of POMC null mice. Our findings contribute to a growing body of evidence demonstrating that opioid peptides are an integral component of vertebrate retinas, including those of mammals.