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Opioids are effective analgesics in the management of severe pain. However, tolerance, leading to dose escalation and adverse effects are significant limiting factors in their use. The role of peripheral opioid receptors in analgesia has been discussed especially under inflammatory conditions. The results from pharmacological and conditional knockout studies together do not provide a clear picture of the contribution of peripheral opioid receptors on antinociceptive tolerance and this needs to be evaluated. Therefore, we studied whether the peripherally restricted opioid receptor antagonist, methylnaltrexone (MNTX), could prevent morphine tolerance without attenuating the antinociceptive effect of morphine. Male Sprague-Dawley rats were treated for 7 days with increasing subcutaneous doses of morphine (5-30 mg/kg) and were coadministered saline, MNTX (0.5 or 2 mg/kg), or naltrexone (NTX; 2 mg/kg). Nociception was assessed with tail-flick, hotplate, and von Frey tests. Morphine, MNTX, and NTX concentrations in the plasma, brain, and spinal cord were measured by liquid chromatography-tandem mass spectrometry. In acute coadministration, NTX, but not MNTX, abolished the acute antinociceptive effects of morphine in all nociceptive tests. The antinociceptive tolerance after repeated morphine administration was also prevented by NTX but not by MNTX. MNTX penetrated to the spinal cord and the brain to some extent after repeated administration. The results do not support the use of MNTX for preventing opioid tolerance and also suggest that morphine tolerance is mediated by central rather than peripheral opioid receptors in the rat.

Adipogenesis in adulthood replaces fat cells that turn over and can contribute to the development of obesity. However, the proliferative potential of adipocyte progenitors in vivo is unknown (Faust et al., 1976; Faust et al., 1977; Hirsch and Han, 1969; Johnson and Hirsch, 1972). We addressed this by injecting labeled wild-type embryonic stem cells into blastocysts derived from lipodystrophic A-ZIP transgenic mice, which have a genetic block in adipogenesis. In the resulting chimeric animals, wild-type ES cells are the only source of mature adipocytes. We found that when chimeric animals were fed a high-fat-diet, animals with low levels of chimerism showed a significantly lower adipose tissue mass than animals with high levels of chimerism. The difference in adipose tissue mass was attributed to variability in the amount of subcutaneous adipose tissue as the amount of visceral fat was independent of the level of chimerism. Our findings thus suggest that proliferative potential of adipocyte precursors is limited and can restrain the development of obesity.

Infant mortality from dengue disease is a devastating global health burden that could be minimized with the ability to identify susceptibility for severe disease prior to infection. Although most primary infant dengue infections are asymptomatic, maternally derived anti-dengue immunoglobulin G (IgGs) present during infection can trigger progression to severe disease through antibody-dependent enhancement mechanisms. Importantly, specific characteristics of maternal IgGs that herald progression to severe infant dengue are unknown. Here, we define >= 10% afucosylation of maternal anti-dengue IgGs as a risk factor for susceptibility of infants to symptomatic dengue infections. Mechanistic experiments show that afucosylation of anti-dengue IgGs promotes Fc gamma RIIIa signaling during infection, in turn enhancing dengue virus replication in Fc gamma RIIIa(+) monocytes. These studies identify a post-translational modification of anti-dengue IgGs that correlates with risk for symptomatic infant dengue infections and define a mechanism by which afucosylated antibodies and Fc gamma RIIIa enhance dengue infections.

Caspase proteases execute apoptosis but also function in development. In this issue of Developmental Cell, Weaver et al. report that C. elegans CED-3 caspase promotes animal growth through PMK-1/p38 kinase cleavage, and at the expense of pathogen and stress immunity, revealing an unexpected homeostatic relationship between development and disease.

This tutorial and the accompanying poster (10.1038/s41596-020-0307-7) provide a guide for performing quantitative fluorescence imaging using confocal microscopy. It includes advice and troubleshooting information from sample preparation and microscope setup to data analysis and statistics. When used appropriately, a confocal fluorescence microscope is an excellent tool for making quantitative measurements in cells and tissues. The confocal microscope's ability to block out-of-focus light and thereby perform optical sectioning through a specimen allows the researcher to quantify fluorescence with very high spatial precision. However, generating meaningful data using confocal microscopy requires careful planning and a thorough understanding of the technique. In this tutorial, the researcher is guided through all aspects of acquiring quantitative confocal microscopy images, including optimizing sample preparation for fixed and live cells, choosing the most suitable microscope for a given application and configuring the microscope parameters. Suggestions are offered for planning unbiased and rigorous confocal microscope experiments. Common pitfalls such as photobleaching and cross-talk are addressed, as well as several troubling instrumentation problems that may prevent the acquisition of quantitative data. Finally, guidelines for analyzing and presenting confocal images in a way that maintains the quantitative nature of the data are presented, and statistical analysis is discussed. A visual summary of this tutorial is available as a poster (10.1038/s41596-020-0307-7).

The pathogenesis of hidradenitis suppurativa (HS) centers around Th17/Treg dysfunction illustrated by lesional elevation of IL-17A, IL-6, and other inflammatory mediators resulting in a chronic feed-forward inflammatory cascade. Similar inflammatory mechanisms have been identified in psoriasis and rheumatoid arthritis (RA) in which traditional immunosuppressants (including methotrexate) are routinely used with reasonable levels of disease control. Methotrexate's mechanism of action in these instances includes downregulation of the Th17 axis via alterations in dendritic cell and T-cell activity and maturation. Published data suggest methotrexate in an ineffective therapy in HS, which does not pair with our current understanding of the mechanisms of disease. The reasons behind this, including are discussed. Some HS patients may benefit from drugs such as methotrexate, and acknowledgment of the potential of disease heterogeneity will allow exploration of which factors may enable identification of such individuals.

Background. MK-8591 (4'-ethynyl-2-fluoro-2'-deoxyadenosine [EFdA]) is a novel reverse transcriptase-translocation inhibitor. Methods. We assessed MK-8591 as preexposure prophylaxis in the rhesus macaque model of intrarectal challenge with simian/human immunodeficiency virus (SHIV). In study 1, 8 rhesus macaques received 3.9 mg/kg of MK-8591 orally on day 0 and once weekly for the next 14 weeks. Eight controls were treated with vehicle. All rhesus macaques were challenged with SHIV109CP3 on day 6 and weekly for up to 12 challenges or until infection was confirmed. The dose of MK-8591 was reduced to 1.3 and 0.43 mg/kg/week in study 2 and further to 0.1 and 0.025 mg/kg/week in study 3. In studies 2 and 3, each dose was given up to 6 times once weekly, and animals were challenged 4 times once weekly with SHIV109CP3. Results. Control macaques were infected after a median of 1 challenge (range, 1-4 challenges). All treated animals in studies 1 and 2 were protected, consistent with a 41.5-fold lower risk of infection (P < .0001, by the log-rank test). In study 3, at a 0.1-mg/kg dose, 2 rhesus macaques became infected, consistent with a 7.2-fold lower risk of infection (P = .0003, by the log-rank test). The 0.025-mg/kg dose offered no protection. Conclusions. These data support MK-8591's potential as a preexposure prophylaxis agent.

Mu-opioid receptors (MORs) mediate the rewarding properties of oxycodone and other prescription opioid medications, which have played a central role in the current opioid epidemic in the United States. The human mu-opioid receptor gene (OPRM1) contains a functional single nucleotide polymorphism (SNP), A118G, which has been associated with altered opioid addiction risk, however the mechanisms responsible for this are not well understood. To explore this, we examined oxycodone conditioned place preference (CPP) and self-administration behavior (SA) in A112G mice, which possess a functionally analogous SNP in the mouse mu-opioid receptor gene (Oprm1). For CPP, male and female A112G mice homozygous for the A112 (wild-type; AA) or G112 (GG) allele were conditioned with doses of 1 and 3 mg/kg across an 8-day period. For SA, mice were allowed to self administer oxycodone (unit dose 0.25 mg/kg/infusion, FR1) for 4h/day for 10 consecutive days. We observed no effects of genotype or sex on conditioned place preference behavior. Oxycodone 3 mg/kg increased locomotor activity in AA mice but not GG mice, and both male and female GG mice self-administered significantly more oxycodone compared to their wild-type AA littermates. These studies suggest that the G allele promotes greater opioid intake, which may underlie greater opioid addiction morbidity in G-allele carriers.