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Lithium (Li) is one of the most effective drugs for treating bipolar disorder (BD), however, there is presently no way to predict response to guide treatment. The aim of this study is to identify functional genes and pathways that distinguish BD Li responders (LR) from BD Li non-responders (NR). An initial Pharmacogenomics of Bipolar Disorder study (PGBD) GWAS of lithium response did not provide any significant results. As a result, we then employed network-based integrative analysis of transcriptomic and genomic data. In transcriptomic study of iPSC-derived neurons, 41 significantly differentially expressed (DE) genes were identified in LR vs NR regardless of lithium exposure. In the PGBD, post-GWAS gene prioritization using the GWA-boosting (GWAB) approach identified 1119 candidate genes. Following DE-derived network propagation, there was a highly significant overlap of genes between the top 500- and top 2000-proximal gene networks and the GWAB gene list (Phypergeometric = 1.28E-09 and 4.10E-18, respectively). Functional enrichment analyses of the top 500 proximal network genes identified focal adhesion and the extracellular matrix (ECM) as the most significant functions. Our findings suggest that the difference between LR and NR was a much greater effect than that of lithium. The direct impact of dysregulation of focal adhesion on axon guidance and neuronal circuits could underpin mechanisms of response to lithium, as well as underlying BD. It also highlights the power of integrative multi-omics analysis of transcriptomic and genomic profiling to gain molecular insights into lithium response in BD.

Combinatorial antiretroviral therapy (cART) has transformed HIV infection from a death sentence to a controllable chronic disease, but cannot eliminate the virus. Latent HIV-1 reservoirs are the major obstacles to cure HIV-1 infection. Previously, we engineered exosomal Tat (Exo-Tat) to reactivate latent HIV-1 from the reservoir of resting CD4+ T cells. Here, we present an HIV-1 eradication platform, which uses our previously described Exo-Tat to activate latent virus from resting CD4+ T cells guided by the specific binding domain of CD4 in interleukin 16 (IL16), attached to the N-terminus of exosome surface protein lysosome-associated membrane protein 2 variant B (Lamp2B). Cells with HIV-1 surface protein gp120 expressed on the cell membranes are then targeted for immune cytolysis by a BiTE molecule CD4-αCD3, which colocalizes the gp120 surface protein of HIV-1 and the CD3 of cytotoxic T lymphocytes. Using primary blood cells obtained from antiretroviral treated individuals, we find that this combined approach led to a significant reduction in replication-competent HIV-1 in infected CD4+ T cells in a clonal in vitro cell system. Furthermore, adeno-associated virus serotype DJ (AAV-DJ) was used to deliver Exo-Tat, IL16lamp2b and CD4-αCD3 genes by constructing them in one AAV-DJ vector (the plasmid was named pEliminator). The coculture of T cells from HIV-1 patients with Huh-7 cells infected with AAV-Eliminator viruses led to the clearance of HIV-1 reservoir cells in the in vitro experiment, which could have implications for reducing the viral reservoir in vivo, indicating that Eliminator AAV viruses have the potential to be developed into therapeutic biologics to cure HIV-1 infection.

We compared four orthogonal technologies for sizing, counting, and phenotyping of extracellular vesicles (EVs) and synthetic particles. The platforms were: single-particle interferometric reflectance imaging sensing (SP-IRIS) with fluorescence, nanoparticle tracking analysis (NTA) with fluorescence, microfluidic resistive pulse sensing (MRPS), and nanoflow cytometry measurement (NFCM). EVs from the human T lymphocyte line H9 (high CD81, low CD63) and the promonocytic line U937 (low CD81, high CD63) were separated from culture conditioned medium (CCM) by differential ultracentrifugation (dUC) or a combination of ultrafiltration (UF) and size exclusion chromatography (SEC) and characterized by transmission electron microscopy (TEM) and Western blot (WB). Mixtures of synthetic particles (silica and polystyrene spheres) with known sizes and/or concentrations were also tested. MRPS and NFCM returned similar particle counts, while NTA detected counts approximately one order of magnitude lower for EVs, but not for synthetic particles. SP-IRIS events could not be used to estimate particle concentrations. For sizing, SP-IRIS, MRPS, and NFCM returned similar size profiles, with smaller sizes predominating (per power law distribution), but with sensitivity typically dropping off below diameters of 60 nm. NTA detected a population of particles with a mode diameter greater than 100 nm. Additionally, SP-IRIS, MRPS, and NFCM were able to identify at least three of four distinct size populations in a mixture of silica or polystyrene nanoparticles. Finally, for tetraspanin phenotyping, the SP-IRIS platform in fluorescence mode was able to detect at least two markers on the same particle, while NFCM detected either CD81 or CD63. Based on the results of this study, we can draw conclusions about existing single-particle analysis capabilities that may be useful for EV biomarker development and mechanistic studies.

Lithium remains the gold standard for the treatment of bipolar disorder (BD); however, its use has declined over the years mainly due to the side effects and the subjective experience of cognitive numbness reported by patients. In the present study, we aim to methodically test the effects of lithium on neurocognitive functioning in the largest single cohort (n = 262) of BD patients reported to date by harnessing the power of a multi-site, ongoing clinical trial of lithium monotherapy. At the cross-sectional level, multivariate analysis of covariance (MANCOVA) was conducted to examine potential group differences across neurocognitive tests [California Verbal Learning Test (CVLT trials 1-5,CVLT delayed recall), Wechsler Digit Symbol, Trail-making Test parts A and B (TMT-A; TMT-B), and a global cognition index]. At the longitudinal level, on a subset of patients (n = 88) who achieved mood stabilization with lithium monotherapy, we explored the effect of lithium treatment across time on neurocognitive functioning. There were no differences at baseline between BD patients that were taking lithium compared with those that were not. At follow-up a significant neurocognitive improvement in the global cognitive index score [F = 31.69; p

ABSTRACT Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, has evolved into multiple variants. Animal models are important to understand variant pathogenesis, particularly for variants with mutations that have significant phenotypic or epidemiological effects. Here, cohorts of naive or previously infected Syrian hamsters (Mesocricetus auratus) were infected with variants to investigate viral pathogenesis and disease protection. Naive hamsters infected with SARS-CoV-2 variants had consistent clinical outcomes, tissue viral titers, and pathology, while hamsters that recovered from initial infection and were reinfected demonstrated less severe clinical disease and lung pathology than their naive counterparts. Males had more frequent clinical signs than females in most variant groups, but few sex variations in tissue viral titers and lung pathology were observed. These findings support the use of Syrian hamsters as a SARS-CoV-2 model and highlight the importance of considering sex differences when using this species. IMPORTANCE With the continued circulation and emergence of new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, understanding differences in the effects between the initial infection and a subsequent reinfection on disease pathogenesis is critical and highly relevant. This study characterizes Syrian hamsters as an animal model to study reinfection with SARS-CoV-2. Previous infection reduced the disease severity of reinfection with different SARS-CoV-2 variants.

Bipolar disorder (BD) is a serious mood disorder associated with circadian rhythm abnormalities. Risk for BD is genetically encoded and overlaps with systems that maintain circadian rhythms. Lithium is an effective mood stabilizer treatment for BD, but only a minority of patients fully respond to monotherapy. Presently, we hypothesized that lithium-responsive BD patients (Li-R) would show characteristic differences in chronotype and cellular circadian rhythms compared to lithium non-responders (Li-NR). Selecting patients from a prospective, multi-center, clinical trial of lithium monotherapy, we examined morning vs. evening preference (chronotype) as a dimension of circadian rhythm function in 193 Li-R and Li-NR BD patients. From a subset of 59 patient donors, we measured circadian rhythms in skin fibroblasts longitudinally over 5 days using a bioluminescent reporter (Per2-luc). We then estimated circadian rhythm parameters (amplitude, period, phase) and the pharmacological effects of lithium on rhythms in cells from Li-R and Li-NR donors. Compared to Li-NRs, Li-Rs showed a difference in chronotype, with higher levels of morningness. Evening chronotype was associated with increased mood symptoms at baseline, including depression, mania, and insomnia. Cells from Li-Rs were more likely to exhibit a short circadian period, a linear relationship between period and phase, and period shortening effects of lithium. Common genetic variation in the IP3 signaling pathway may account for some of the individual differences in the effects of lithium on cellular rhythms. We conclude that circadian rhythms may influence response to lithium in maintenance treatment of BD.

Biological responses to climate change have been widely documented across taxa and regions, but it remains unclear whether species are maintaining a good match between phenotype and environment, i.e. whether observed trait changes are adaptive. Here we reviewed 10,090 abstracts and extracted data from 71 studies reported in 58 relevant publications, to assess quantitatively whether phenotypic trait changes associated with climate change are adaptive in animals. A meta-analysis focussing on birds, the taxon best represented in our dataset, suggests that global warming has not systematically affected morphological traits, but has advanced phenological traits. We demonstrate that these advances are adaptive for some species, but imperfect as evidenced by the observed consistent selection for earlier timing. Application of a theoretical model indicates that the evolutionary load imposed by incomplete adaptive responses to ongoing climate change may already be threatening the persistence of species.

The extinction of native species and introduction of non-native species may lead to the disruption of biotic interactions. Pollination is a critical ecosystem process that often requires mutualisms between animals and plants. Non-native animals may interact with native flowering plants, with the potential to pollinate or steal nectar (larceny) from flowers without pollination. In the Hawaiian Islands, many native plants have lost their original pollinators. Birds and insects are known to visit native plant flowers, but it is unclear whether they pollinate or steal nectar, whether native and non-native species differ in their interactions with flowers, and what influences visitation to flowers. On Oʻahu, we deployed camera traps and conducted in-person observations on four at-risk species of Hawaiian lobelioids (Campanulaceae). We observed birds, mammals, and insects visiting flowers, with a native bird and native bee visiting most frequently. Regardless of native versus non-native status, bees made contact with reproductive structures during most visits (90.5% of visits), while birds stole nectar during most visits (99.3% of visits). Bee and bird visitation increased with the number of flowers on focal plants. Bird visitation also increased with canopy cover and the number of nearby conspecific flowers and decreased with the number of nearby heterospecific flowers. Our results indicate that bees may pollinate plants that were historically bird-pollinated, while native and non-native birds have neutral or negative impacts on these plants. Broadly, we contribute to an understanding of how native plant pollination can be altered in changing ecosystems.

BackgroundBipolar disorder is a serious and common psychiatric disorder characterized by manic and depressive mood switches and a relapsing and remitting course. The cornerstone of clinical management is stabilization and prophylaxis using mood-stabilizing medications to reduce both manic and depressive symptoms. Lithium remains the gold standard of treatment with the strongest data for both efficacy and suicide prevention. However, many patients do not respond to this medication, and clinically there is a great need for tools to aid the clinician in selecting the correct treatment. Large genome wide association studies (GWAS) investigating retrospectively the effect of lithium response are in the pipeline; however, few large prospective studies on genetic predictors to of lithium response have yet been conducted. The purpose of this project is to identify genes that are associated with lithium response in a large prospective cohort of bipolar patients and to better understand the mechanism of action of lithium and the variation in the genome that influences clinical response.Methods/designThis study is an 11-site prospective non-randomized open trial of lithium designed to ascertain a cohort of 700 subjects with bipolar I disorder who experience protocol-defined relapse prevention as a result of treatment with lithium monotherapy. All patients will be diagnosed using the Diagnostic Interview for Genetic Studies (DIGS) and will then enter a 2-year follow-up period on lithium monotherapy if and when they exhibit a score of 1 (normal, not ill), 2 (minimally ill) or 3 (mildly ill) on the Clinical Global Impressions of Severity Scale for Bipolar Disorder (CGI-S-BP Overall Bipolar Illness) for 4 of the 5 preceding weeks. Lithium will be titrated as clinically appropriate, not to exceed serum levels of 1.2 mEq/L. The sample will be evaluated longitudinally using a wide range of clinical scales, cognitive assessments and laboratory tests. On relapse, patients will be discontinued or crossed-over to treatment with valproic acid (VPA) or treatment as usual (TAU). Relapse is defined as a DSM-IV manic, major depressive or mixed episode or if the treating physician decides a change in medication is clinically necessary. The sample will be genotyped for GWAS. The outcome for lithium response will be analyzed as a time to event, where the event is defined as clinical relapse, using a Cox Proportional Hazards model. Positive single nucleotide polymorphisms (SNPs) from past genetic retrospective studies of lithium response, the Consortium on Lithium Genetics (ConLiGen), will be tested in this prospective study sample; a meta-analysis of these samples will then be performed. Finally, neurons will be derived from pluripotent stem cells from lithium responders and non-responders and tested in vivo for response to lithium by gene expression studies. SNPs in genes identified in these cellular studies will also be tested for association to response.DiscussionLithium is an extraordinarily important therapeutic drug in the clinical management of patients suffering from bipolar disorder. However, a significant proportion of patients, 30-40 %, fail to respond, and there is currently no method to identify the good lithium responders before initiation of treatment. Converging evidence suggests that genetic factors play a strong role in the variation of response to lithium, but only a few genes have been tested and the samples have largely been retrospective or quite small. The current study will collect an entirely unique sample of 700 patients with bipolar disorder to be stabilized on lithium monotherapy and followed for up to 2 years. This study will produce useful information to improve the understanding of the mechanism of action of lithium and will add to the development of a method to predict individual response to lithium, thereby accelerating recovery and reducing suffering and cost.Trial registrationClinicalTrials.gov Identifier: NCT01272531 Registered: January 6, 2011.

This study examined the relationship between maternal food source and preparation during pregnancy and the duration of breastfeeding among 751 mother–child dyads in the United States. The data collected from the Environmental influences on Child Health Outcomes (ECHO) Program included twelve cohorts of mothers (age ≥ 18) who delivered infant(s). Three categories of maternal food source and preparation including, High, Moderate, or Low Food Source Quality were derived from the mother report. The mean duration of breastfeeding differed strongly across the three categories. The High Food Source Quality group breastfed an average of 41 weeks, while shorter durations were observed for the Moderate (26 weeks) and Low (16 weeks) Food Source Quality groups. Cox proportional hazards models were used to estimate the relative hazard of time to breastfeeding cessation for each participant characteristic. The full model adjusted for clustering/cohort effect for all participant characteristics, while the final model adjusted for the subset of characteristics identified from variable reduction modeling. The hazard of breastfeeding cessation for those in the High Food Source Quality group was 24% less than the Moderate group (RH = 0.76; 95% CI, 0.63–0.92). Pregnant women in the High Food Source Quality group breastfed longer than the Moderate and Low groups. We encourage more detailed studies in the future to examine this relationship longitudinally.

Abstract We compared four orthogonal technologies for sizing, counting, and phenotyping of extracellular vesicles (EVs) and synthetic particles. The platforms were: single‐particle interferometric reflectance imaging sensing (SP‐IRIS) with fluorescence, nanoparticle tracking analysis (NTA) with fluorescence, microfluidic resistive pulse sensing (MRPS), and nanoflow cytometry measurement (NFCM). EVs from the human T lymphocyte line H9 (high CD81, low CD63) and the promonocytic line U937 (low CD81, high CD63) were separated from culture conditioned medium (CCM) by differential ultracentrifugation (dUC) or a combination of ultrafiltration (UF) and size exclusion chromatography (SEC) and characterized by transmission electron microscopy (TEM) and Western blot (WB). Mixtures of synthetic particles (silica and polystyrene spheres) with known sizes and/or concentrations were also tested. MRPS and NFCM returned similar particle counts, while NTA detected counts approximately one order of magnitude lower for EVs, but not for synthetic particles. SP‐IRIS events could not be used to estimate particle concentrations. For sizing, SP‐IRIS, MRPS, and NFCM returned similar size profiles, with smaller sizes predominating (per power law distribution), but with sensitivity typically dropping off below diameters of 60 nm. NTA detected a population of particles with a mode diameter greater than 100 nm. Additionally, SP‐IRIS, MRPS, and NFCM were able to identify at least three of four distinct size populations in a mixture of silica or polystyrene nanoparticles. Finally, for tetraspanin phenotyping, the SP‐IRIS platform in fluorescence mode was able to detect at least two markers on the same particle, while NFCM detected either CD81 or CD63. Based on the results of this study, we can draw conclusions about existing single‐particle analysis capabilities that may be useful for EV biomarker development and mechanistic studies.

It is unclear whether species’ responses to climate change tend to be adaptive or sufficient to keep up with climate change. Here, Radchuk et al. perform a meta-analysis showing that in birds phenology has advanced adaptively in some species, though not all the way to the new optima.

Erythema nodosum leprosum (ENL), or type 2 lepra reaction, is a multi-system immune-mediated complication in patients with multibacillary leprosy, frequently associated with chronicity and recurrences. Management of ENL requires high doses of oral corticosteroids, which may not be universally effective and pose serious adverse effects. Thalidomide has proven to be a steroid-sparing agent and is useful in controlling the reactions. However, many centres do not employ it in outpatient settings due to adverse effects and teratogenicity risk. Hence, we studied the feasibility of treating ENLs and reported the therapeutic outcome.This is a five-year record-based analysis of ENL leprosy patients treated with thalidomide, includingdescriptive statistics of demographic variables. Clinical characteristics were stratified by treatment compliance status (yes/no). Incidence rates and rate ratios for recovery stratified by bacillary index, type of ENL presentation and MDT treatment status were calculated.Out of 102 ENL patients treated with thalidomide, 68 (66.7%) were compliant and improved. Among them, ENL recurrence was noted in 11(16.2%) patients. The commonest thalidomide side effect was pedal oedema (73.5%). Patients with bacillary index (BI) less than or equal to 4.0 had a 37% increase in the incidence of recovery. Patients with acute ENL were almost twice as likely to recover as those with chronic ENL. Also, the improvement was two and a half times greater among those who completed MDT as compared to those on MDT. The study showed that thalidomide treatment for patients with ENL is possible in outpatientclinics. We also successfully prevented pregnancies to a larger extent through counselling for contraception.We observed that early institution of thalidomide induces faster remission and prevents ENL recurrence.

Studies of evolution in wild populations often find that the heritable phenotypic traits of individuals producing the most offspring do not increase proportionally in the population. This paradox may arise when phenotypic traits influence both fecundity and viability and when there is a tradeoff between these fitness components, leading to opposing selection. Such tradeoffs are the foundation of life history theory, but they are rarely investigated in selection studies. Timing of breeding is a classic example of a heritable trait under directional selection that does not result in an evolutionary response. Using a 22-y study of a tropical parrot, we show that opposing viability and fecundity selection on the timing of breeding is common and affects optimal breeding date, defined by maximization of fitness. After accounting for sampling error, the directions of viability (positive) and fecundity (negative) selection were consistent, but the magnitude of selection fluctuated among years. Environmental conditions (rainfall and breeding density) primarily and breeding experience secondarily modified selection, shifting optimal timing among individuals and years. In contrast to other studies, viability selection was as strong as fecundity selection, late-born juveniles had greater survival than early-born juveniles, and breeding later in the year increased fitness under opposing selection. Our findings provide support for life history tradeoffs influencing selection on phenotypic traits, highlight the need to unify selection and life history theory, and illustrate the importance of monitoring survival as well as reproduction for understanding phenological responses to climate change.

BackgroundRC-101 is a congener of the antiretroviral peptide retrocyclin, which we and others have reported is active against clinical HIV-1 isolates from all major clades, does not hemagglutinate, and is non-toxic and non-inflammatory in cervicovaginal cell culture. Herein, film-formulated RC-101 was assessed for its antiviral activity in vitro, safety in vivo, retention in the cervix and vagina, and ability to remain active against HIV-1 and SHIV after intravaginal application in macaques.Methodology/principal findingsRC-101 was formulated as a quick-dissolving film (2000 µg/film), retained complete activity in vitro as compared to unformulated peptide, and was applied intravaginally in six pigtailed macaques daily for four days. At one and four days following the final application, the presence of RC-101 was assessed in peripheral blood, cervicovaginal lavage, cytobrushed cervicovaginal cells, and biopsied cervical and vaginal tissues by quantitative western blots. One day following the last film application, cervical biopsies from RC-101-exposed and placebo-controlled macaques were collected and were subjected to challenge with RT-SHIV in an ex vivo organ culture model. RC-101 peptide was detected primarily in the cytobrush and biopsied cervical and vaginal tissues, with little to no peptide detected in lavage samples, suggesting that the peptide was associated with the cervicovaginal epithelia. RC-101 remained in the tissues and cytobrush samples up to four days post-application, yet was not detected in any sera or plasma samples. RC-101, extracted from cytobrushes obtained one day post-application, remained active against HIV-1 BaL. Importantly, cervical biopsies from RC-101-treated animals reduced RT-SHIV replication in ex vivo organ culture as compared to placebo-treated animals.Conclusions/significanceFormulated RC-101 was stable in vivo and was retained in the mucosa. The presence of antivirally active RC-101 after five days in vivo suggests that RC-101 would be an important molecule to develop further as a topical microbicide to prevent HIV-1 transmission.

Ovarian cancer, the most deadly gynecological malignancy in U.S. women, metastasizes uniquely, spreading through the peritoneal cavity and often generating widespread metastatic sites before diagnosis. The vast majority of ovarian cancer cases occur in women over 40 and the median age at diagnosis is 63. Additionally, elderly women receive poorer prognoses when diagnosed with ovarian cancer. Despite age being a significant risk factor for the development of this cancer, there are little published data which address the impact of aging on ovarian cancer metastasis. Here we report that the aged host is more susceptible to metastatic success using two murine syngeneic allograft models of ovarian cancer metastasis. This age-related increase in metastatic tumor burden corresponds with an increase in tumor infiltrating lymphocytes (TILs) in tumor-bearing mice and alteration of B cell-related pathways in gonadal adipose tissue. Based on this work, further studies elucidating the status of B cell TILs in mouse models of metastasis and human tumors in the context of aging are warranted.

Shock and kill strategies involving the use of small molecules to induce viral transcription in resting CD4+ T cells (shock) followed by immune mediated clearance of the reactivated cells (kill), have been proposed as a method of eliminating latently infected CD4+ T cells. The combination of the histone deacetylase (HDAC) inhibitor romidepsin and protein kinase C (PKC) agonist bryostatin-1 is very effective at reversing latency in vitro. However, we found that primary HIV-1 specific CD8+ T cells were not able to eliminate autologous resting CD4+ T cells that had been reactivated with these drugs. We tested the hypothesis that the drugs affected primary CD8+ T cell function and found that both agents had inhibitory effects on the suppressive capacity of HIV-specific CD8+ T cells from patients who control viral replication without antiretroviral therapy (elite suppressors/controllers). The inhibitory effect was additive and multi-factorial in nature. These inhibitory effects were not seen with prostratin, another PKC agonist, either alone or in combination with JQ1, a bromodomain-containing protein 4 inhibitor. Our results suggest that because of their adverse effects on primary CD8+ T cells, some LRAs may cause immune-suppression and therefore should be used with caution in shock and kill strategies.

Gonadectomy is widely used to treat and prevent behavior problems including the aggressive behavior of dogs. The aim of this study was to determine whether aggressive behavior toward familiar people, strangers, or other dogs was significantly different in dogs gonadectomized at various ages vs. intact dogs using the Canine Behavioral Assessment Research Questionnaire (C-BARQ) with multivariate analysis. Of 15,370 initial surveys, those for dogs reported to have been gonadectomized at less than 6 weeks of age or to correct a behavior problem, and those with incomplete answers to questions regarding independent or dependent variables were excluded, leaving 13,795 for the analysis of aggressive behavior toward familiar people: 13,498 for aggressive behavior toward strangers and 13,237 for aggressive behavior toward dogs. Aggressive behavior was defined (a) using mean scores for all questions on the C-BARQ for aggressive behavior (range 0–4) and (b) comparing dogs with no aggressive behavior (all questions answered 0) to dogs with moderate or severe aggression (at least one score of 2, 3, or 4). Data for intact dogs were compared with those for dogs gonadectomized at 6 months or less, 7–12 months, 11–18 months, and >18 months. Neither gonadectomy nor age at gonadectomy showed an association with aggression toward familiar people or dogs. However, there was a low but significant increase in the odds of moderate or severe aggression toward strangers for all gonadectomized dogs compared with intact dogs, but this effect was driven entirely by data for dogs gonadectomized at 7–12 months of age, which were 26% more likely to demonstrate aggression toward strangers. This large, comprehensive study of the relationships between gonadectomy and aggressive behavior in dogs demonstrates that when the many factors affecting aggressive behavior are considered, there is no evidence that gonadectomy at any age alters aggressive behavior toward familiar people or dogs, and there is only a minimal increase in aggression toward strangers. Given the increasing evidence of significant negative health effects of gonadectomy, there is an urgent need to systematically examine other means of preventing unwanted procreation, such as vasectomy and hysterectomy.

Fusion inhibitors are a class of antiretroviral drugs used to prevent entry of HIV into host cells. Many of the fusion inhibitors being developed, including the drug enfuvirtide, are peptides designed to competitively inhibit the viral fusion protein gp41. With the emergence of drug resistance, there is an increased need for effective and unique alternatives within this class of antivirals. One such alternative is a class of cyclic, cationic, antimicrobial peptides known as θ-defensins, which are produced by many non-human primates and exhibit broad-spectrum antiviral and antibacterial activity. Currently, the θ-defensin analog RC-101 is being developed as a microbicide due to its specific antiviral activity, lack of toxicity to cells and tissues, and safety in animals. Understanding potential RC-101 resistance, and how resistance to other fusion inhibitors affects RC-101 susceptibility, is critical for future development. In previous studies, we identified a mutant, R5-tropic virus that had evolved partial resistance to RC-101 during in vitro selection. Here, we report that a secondary mutation in gp41 was found to restore replicative fitness, membrane fusion, and the rate of viral entry, which were compromised by an initial mutation providing partial RC-101 resistance. Interestingly, we show that RC-101 is effective against two enfuvirtide-resistant mutants, demonstrating the clinical importance of RC-101 as a unique fusion inhibitor. These findings both expand our understanding of HIV drug-resistance to diverse peptide fusion inhibitors and emphasize the significance of compensatory gp41 mutations.

Pulmonary-renal syndrome is a medical emergency; etiology of which has broad differential diagnosis. Delay in both diagnosis and initiation of management may result in end-organ damage. Management decisions may have to be empiric till a rapid, definitive tissue diagnosis is established. We present such a case where prompt recognition and immediate treatment was initiated, although the patient sustained irreversible end-organ damage. The case also highlights the need to interpret the kidney biopsy data (namely, immunofluroscence findings) in the context of clinical presentation.

RC-101 is a congener of the antiretroviral peptide retrocyclin, which we and others have reported is active against clinical HIV-1 isolates from all major clades, does not hemagglutinate, and is non-toxic and non-inflammatory in cervicovaginal cell culture. Herein, film-formulated RC-101 was assessed for its antiviral activity in vitro, safety in vivo, retention in the cervix and vagina, and ability to remain active against HIV-1 and SHIV after intravaginal application in macaques.RC-101 was formulated as a quick-dissolving film (2000 µg/film), retained complete activity in vitro as compared to unformulated peptide, and was applied intravaginally in six pigtailed macaques daily for four days. At one and four days following the final application, the presence of RC-101 was assessed in peripheral blood, cervicovaginal lavage, cytobrushed cervicovaginal cells, and biopsied cervical and vaginal tissues by quantitative western blots. One day following the last film application, cervical biopsies from RC-101-exposed and placebo-controlled macaques were collected and were subjected to challenge with RT-SHIV in an ex vivo organ culture model. RC-101 peptide was detected primarily in the cytobrush and biopsied cervical and vaginal tissues, with little to no peptide detected in lavage samples, suggesting that the peptide was associated with the cervicovaginal epithelia. RC-101 remained in the tissues and cytobrush samples up to four days post-application, yet was not detected in any sera or plasma samples. RC-101, extracted from cytobrushes obtained one day post-application, remained active against HIV-1 BaL. Importantly, cervical biopsies from RC-101-treated animals reduced RT-SHIV replication in ex vivo organ culture as compared to placebo-treated animals.Formulated RC-101 was stable in vivo and was retained in the mucosa. The presence of antivirally active RC-101 after five days in vivo suggests that RC-101 would be an important molecule to develop further as a topical microbicide to prevent HIV-1 transmission.

Central nervous system (CNS) invasion during acute-stage HIV-infection has been demonstrated in a small number of individuals, but there is no evidence of neurological impairment at this stage and virus infection in brain appears to be controlled until late-stage disease. Using our reproducible SIV macaque model to examine the earliest stages of infection in the CNS, we identified immune responses that differentially regulate inflammation and virus replication in the brain compared to the peripheral blood and lymphoid tissues. SIV replication in brain macrophages and in brain of SIV-infected macaques was detected at 4 days post-inoculation (p.i.). This was accompanied by upregulation of innate immune responses, including IFNbeta, IFNbeta-induced gene MxA mRNA, and TNFalpha. Additionally, IL-10, the chemokine CCL2, and activation markers in macrophages, endothelial cells, and astrocytes were all increased in the brain at four days p.i. We observed synchronous control of virus replication, cytokine mRNA levels and inflammatory markers (MHC Class II, CD68 and GFAP) by 14 days p.i.; however, control failure was followed by development of CNS lesions in the brain. SIV infection was accompanied by induction of the dominant-negative isoform of C/EBPbeta, which regulates SIV, CCL2, and IL6 transcription, as well as inflammatory responses in macrophages and astrocytes. This synchronous response in the CNS is in part due to the effect of the C/EBPbeta on virus replication and cytokine expression in macrophage-lineage cells in contrast to CD4+ lymphocytes in peripheral blood and lymphoid tissues. Thus, we have identified a crucial period in the brain when virus replication and inflammation are controlled. As in HIV-infected individuals, though, this control is not sustained in the brain. Our results suggest that intervention with antiretroviral drugs or anti-inflammatory therapeutics with CNS penetration would sustain early control. These studies further suggest that interventions should target HIV-infected individuals with increased CCL2 levels or HIV RNA in the CNS.

Human immunodeficiency virus (HIV) infection frequently causes neurologic disease even with anti-retroviral treatment. Although associations between MHC class I alleles and acquired immunodeficiency syndrome (AIDS) have been reported, the role MHC class I alleles play in restricting development of HIV-induced organ-specific diseases, including neurologic disease, has not been characterized. This study examined the relationship between expression of the MHC class I allele Mane-A*10 and development of lentiviral-induced central nervous system (CNS) disease using a well-characterized simian immunodeficiency (SIV)/pigtailed macaque model. The risk of developing CNS disease (SIV encephalitis) was 2.5 times higher for animals that did not express the MHC class I allele Mane-A*10 (P = 0.002; RR = 2.5). Animals expressing the Mane-A*10 allele had significantly lower amounts of activated macrophages, SIV RNA, and neuronal dysfunction in the CNS than Mane-A*10 negative animals (P