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Demodex musculi is a prostigmatid follicular mite that has rarely been reported in laboratory mice. Although prevalence of this species has not been assessed formally, we have found that many imported mouse strains from noncommercial sources harbor Demodex mites. To assess whether an acaricide can be used to eradicate this mite, infested immunocompromised mice were provided ivermectin-compounded (12 ppm) feed without restriction for 8 wk (n = 10), were treated topically with moxidectin and imidacloprid (MI; 3 and 13 mg/kg, respectively) weekly for 8 wk (n = 10), or remained untreated (n = 10). Mice were confirmed to be mite-infested before treatment and were tested after treatment by using fur plucks (FP), deep skin scrapes (DSS), and PCR analysis of fur swabs. In addition, the presence of mites was confirmed through skin biopsies at 2 study endpoints (1 wk [n = 5] and 12 wk [n = 5] after treatment). Samples collected before treatment and from untreated mice were positive for D. musculi at all time points and by all test modalities. After treatment, all ivermectin-treated animals remained infested, whereas mice treated with MI were repeatedly negative by all test modalities. An additional shortened treatment trial revealed that 4 wk of MI (n = 7) was insufficient to eradicate mites. Neither treatment produced any evidence of adverse effects according to hematology, serum chemistry parameters, behavior, body weight, and histopathology. Of the 70 PCR assays performed in treated mice, 14 were positive when FP+DSS was negative. In 6 cases where PCR results were negative, 5 were positive by FP+DSS and a single sample was positive on skin biopsy. Although PCR analysis has a high detection rate for D. musculi, FP+DSS can further enhance the detection rate. In conclusion, topical MI administered for 8 consecutive weeks can safely eradicate D. musculi from an immunocompromised mouse strain.

Tight binding of Gdown1 represses RNA polymerase II (Pol II) function in a manner that is reversed by Mediator, but the structural basis of these processes is unclear. Although Gdownl is intrinsically disordered, its Pol II interacting domains were localized and shown to occlude transcription factor IIF (TFIIF) and transcription factor liB (TFIIB) binding by perfect positioning on their Pol II interaction sites. Robust binding of Gdownl to Pol II is established by cooperative interactions of a strong Pol II binding region and two weaker binding modulatory regions, thus providing a mechanism both for tight Pol II binding and transcription inhibition and for its reversal. In support of a physiological function for Gdownl in transcription repression, Gdownl co-localizes with Pol II in transcriptionally silent nuclei of early Drosophila embryos but re-localizes to the cytoplasm during zygotic genome activation. Our study reveals a self-inactivation through Gdownl binding as a unique mode of repression in Pol II function.

Cocaine addiction treatment is difficult due to the current lack of approved pharmacotherapuetics. Several preclinical and clinical studies have demonstrated that the mu opioid receptor (MOPr) antagonist/kappa opioid receptor (KOPr) partial agonist naltrexone (NTX) reduces the subjective effects and self-administration of cocaine. However, very limited research has examined the ability of the structurally similar MOPr antagonist/KOPr partial agonist nalmefene (NMF) to reduce cocaine reward. Here we examine the effect of low (1 mg/kg) and high (10 mg/kg) doses of NTX or NMF on cocaine place preference. In vivo characterization of these NTX and NMF doses were performed to examine their effectiveness at MOPr and KOPr. Results: Both NTX doses and high dose NMF significantly reduced cocaine place preference. Conversely, a significant place avoidance was observed for high dose NTX and both NMF doses. Interestingly, neither NTX nor NMF blocked cocaine-induced hyperlocomotion. High dose NTX and both NMF doses fully blocked MOPr agonist morphine-induced thermal analgesia as well as KOPr agonist U50,488H-induced locomotor discoordination. However, low dose NTX fully blocked morphine analgesia but not U50,488H locomotor discoordination suggesting that low dose NIX is effective at MOPr but not KOPr. Conclusion: Both NTX and NMF block the place preference, but not locomotor activating, effects of cocaine. These results suggest that both NTX and NMF may be viable pharmacotheraputics for some aspects of cocaine addiction. This is an important step to understanding the potential mechanism(s) of action of NTX and NMF for the development of more efficacious pharmacological treatments for substance use disorders. (C) 2018 Elsevier Ltd. All rights reserved.

Anti-Mullerian hormone (AMH) produced by ovarian granulosa cells (GCs) plays a crucial role in ovarian function. It is used as a diagnostic and/or prognostic marker of fertility as well as for pathophysiological conditions in women. In this study, we investigated the underlying mechanism for regulation of AMH expression in GCs using primary mouse GCs and a human GC tumor-derived KGN cell line. We find that growth differentiation factor 9 (GDF9) and bone morphogenetic factor 15 (BMP15) together (GDF9 + BMP15), but not when tested separately, significantly induce AMH expression in vitro and in vivo (serum AMH). Our results show that GDF9 + BMP15 through the PI3K/Akt and Smad2/3 pathways synergistically recruit the coactivator p300 on the AMH promoter region that promotes acetylation of histone 3 lysine 27 (H3K27ac), facilitating AMH/Amh expression. Intriguingly, we also find that FSH inhibits GDF9 + BMP15-induced increase of AMH/Amh expression. This inhibition occurs through FSH-induced protein kinase A/SF1-mediated expression of gonadotropin inducible ovarian transcription factor 1, a transcriptional repressor, that recruits histone deacetylase 2 to deacetylate H3K27ac, resulting in the suppression of AMH/Amh expression. Furthermore, we report that ovarian Amh mRNA levels are significantly higher in Fshb-null mice (Fsh beta(-/-)) compared with those in wild-type (WT) mice. In addition, ovarian Amh mRNA levels are restored in Fshb-null mice expressing a human WT FSHb transgene (FSH beta(-/-)hFSH beta(WT)). Our study provides a mechanistic insight into the regulation of AMH expression that has many implications in female reproduction/fertility.

Comprised of telomeric TTAGGG repeats and shelterin, telomeres ensure that the natural ends of chromosomes remain impervious to the DNA damage response. Telomeres carry a long constitutive 3 overhang that can bind replication protein A (RPA) and activate the ATR Ser/Thr kinase (ATR), which induces cell cycle arrest. A single-stranded (ss) TTAGGG repeat-binding protein in mouse shelterin, POT1a, has been proposed to repress ATR signaling by preventing RPA binding. Repression of ATR at telomeres requires tethering of POT1a to the other shelterin subunits situated on the double-stranded (ds) telomeric DNA. The simplest model of ATR repression, the tethered exclusion model, suggests that the only critical features of POT1a are its connection to shelterin and its binding to ss telomeric DNA. In agreement with the model, we show here that a shelterin-tethered variant of RPA70 (lacking the ATR recruitment domain) can repress ATR signaling at telomeres that lack POT1a. However, arguing against the tethered exclusion model, the nearly identical POT1b subunit of shelterin has been shown to be much less proficient than POT1a in repression of ATR. We now show that POT1b has the intrinsic ability to fully repress ATR but is prevented from doing so when bound to Ctc1, Stn1, Ten1 (CST), the complex needed for telomere end processing. These results establish that shelterin represses ATR with a tethered ssDNA-binding domain that excludes RPA from the 3 overhang and also reveal an unexpected effect of CST on the ability of POT1b to repress ATR.

Background: The Kreek-McHugh-Schluger-Kellogg (KMSK) scales provide a rapid assessment of maximal self-exposure to specific drugs and can be used as a dimensional instrument. This study provides a re-evaluation of the KMSK scales for cannabis, alcohol, cocaine, and heroin in a relatively large multi-ethnic cohort, and also the first systematic comparison of gender-specific profiles of drug exposure with this scale. Methods: This was an observational study of n = 1,133 consecutively ascertained adult volunteers. The main instruments used were the SCID-I interview (DSM-IV criteria) and KMSK scales for cannabis, alcohol, cocaine, and heroin. Results: Participants were 852 volunteers (297 female) with specific DSM-IV abuse or dependence diagnoses, and 281 volunteers without any drug diagnoses (154 female). Receiver operating characteristic (ROC) curves were calculated for concurrent validity of KMSK scores with the respective DSM-IV dependence diagnoses. The areas under the ROC curves for men and women combined were 99.5% for heroin, 97% for cocaine, 93% for alcohol, and 85% for cannabis. Newly determined optimal KMSK "cutpoint" scores were identical for men and women for cocaine and heroin dependence diagnoses, but were higher in men than in women, for cannabis and alcohol dependence diagnoses. Conclusions: This study confirms the scales' effectiveness in performing rapid dimensional analyses for cannabis, alcohol, cocaine, and heroin exposure, in a cohort larger than previously reported, with "cutpoints" changed from initial determinations, based on this larger sample. The KMSK scales also detected gender differences in self-exposure to alcohol and cannabis that are associated with the respective dependence diagnoses.

Patients with epidermodysplasia verruciformis (EV) and biallelic null mutations of TMC6 (encoding EVER1) or TMC8 (EVER2) are selectively prone to disseminated skin lesions due to keratinocyte-tropic human beta-papillomaviruses (beta-HPVs), which lack E5 and E8. We describe EV patients homozygous for null mutations of the CIB1 gene encoding calcium-and integrin-binding protein-1 (CIB1). CIB1 is strongly expressed in the skin and cultured keratinocytes of controls but not in those of patients. CIB1 forms a complex with EVER1 and EVER2, and CIB1 proteins are not expressed in EVER1- or EVER2-deficient cells. The known functions of EVER1 and EVER2 in human keratinocytes are not dependent on CIB1, and CIB1 deficiency does not impair keratinocyte adhesion or migration. In keratinocytes, the CIB1 protein interacts with the HPV E5 and E8 proteins encoded by alpha-HPV16 and gamma-HPV4, respectively, suggesting that this protein acts as a restriction factor against HPVs. Collectively, these findings suggest that the disruption of CIB1-EVER1-EVER2-dependent keratinocyte-intrinsic immunity underlies the selective susceptibility to beta-HPVs of EV patients.

Background: Immunity decreases with age, which leads to reactivation of varicella zoster virus (VZV). In human subjects age-associated immune changes are usually measured in blood leukocytes; however, this might not reflect alterations in tissue specific immunity. Objectives: We used a VZV antigen challenge system in the skin to investigate changes in tissue-specific mechanisms involved in the decreased response to this virus during aging. Methods: We assessed cutaneous immunity based on the extent of erythema and induration after intradermal VZV antigen injection. We also performed immune histology and transcriptomic analyses on skin biopsy specimens taken from the challenge site in young (<40 years) and old (>65 years) subjects. Results: Old human subjects exhibited decreased erythema and induration, CD4+ and CD8+ T-cell infiltration, and attenuated global gene activation at the site of cutaneous VZV antigen challenge compared with young subjects. This was associated with increased sterile inflammation in the skin in the same subjects related to p38 mitogen-activated protein kinase-related proinflammatory cytokine production (P <.0007). We inhibited systemic inflammation in old subjects by means of pretreatment with an oral small-molecule p38 mitogen-activated protein kinase inhibitor (Losmapimod; GlaxoSmithKline, Brentford, United Kingdom), which reduced both serum C-reactive protein levels and peripheral blood monocyte secretion of IL-6 and TNF-alpha. In contrast, cutaneous responses to VZV antigen challenge were increased significantly in the same subjects (P <.0003). Conclusion: Excessive inflammation in the skin early after antigen challenge retards antigen-specific immunity. However, this can be reversed by inhibition of inflammatory cytokine production that can be used to promote vaccine efficacy and the treatment of infections and malignancy during aging.