PCID2 dysregulates transcription and viral RNA processing to promote HIV-1 latency.

HIV-1 latency results from tightly regulated molecular processes that act at distinct steps of HIV-1 gene expression. Here, we characterize PCI domain-containing 2 (PCID2) protein, a subunit of the transcription and export complex 2 (TREX2) complex, to enforce transcriptional repression and post-transcriptional blocks to HIV-1 gene expression during latency. PCID2 bound the latent HIV-1 LTR (long terminal repeat) and repressed transcription initiation during latency. Depletion of PCID2 remodeled the chromatin landscape at the HIV-1 promoter and resulted in transcriptional activation and latency reversal. Immunoprecipitation coupled to mass spectrometry identified PCID2-interacting proteins to include negative viral RNA (vRNA) splicing regulators, and PCID2 depletion resulted in over-splicing of intron-containing vRNA in cell lines and primary cells obtained from PWH. MCM3AP and DSS1, two other RNA-binding TREX2 complex subunits, also inhibit transcription initiation and vRNA alternative splicing during latency. Thus, PCID2 is a novel HIV-1 latency-promoting factor, which in context of the TREX2 sub-complex PCID2-DSS1-MCM3AP blocks transcription and dysregulates vRNA processing.

Equilibrio ocupacional en recursos de rehabilitación psicosocial durante el confinamiento por covid-19, estudio observacional prospectivo / Occupational balance in psychosocial rehabilitation resources during confinement by covid-19, prospective observational study

Objetivos: estudiar el impacto en el equilibrio ocupacional durante el primer cuatrimestre de la pandemia COVID-19 en España. Métodos: estudio observacional prospectivo en el que han participado 411 personas con diagnóstico de trastorno mental grave atendidas en dispositivos de rehabilitación psicosocial distribuidos por todo el territorio estatal. Se ha empleado el OBQ-E y la escala ACO, creada por las propias personas investigadoras. Resultados: el equilibrio ocupacional disminuye en la mayoría de las personas durante el periodo de confinamiento y vuelve a aumentar a un nivel similar al de la pre- pandemia según comienza la desescalada de medidas de confinamiento. Hay actividades como el uso de las nuevas tecnologías en las que se observa un incremento en la sensación de cambio ocupacional, que se mantiene tras el cierre del dispositivo. Conclusiones: el equilibrio ocupacional se ve alterado al modificar la rutina diaria y las condiciones del ambiente. Se puede observar una clara capacidad de resiliencia cuando las condiciones cambian y/o vuelven a la normalidad.(AU)

Objective: to study the impact on occupational balance during the first quarter of the COVID-19 pandemic in Spain. Methods: prospective observational study involving 411 persons with diagnosis of severe mental disorder treated in psychosocial rehabilitation devices distributed throughout the state territory. OBQ-E and the ACO scale, created by the researchers themselves, have been used. Results: n most of the participants their occupational balance drops during the confinement period and increases again to a level similar to that of the pre-pandemic as the de-escalation of confinement measures begins. There are activities such as the use of new technologies in which there is an increase in the feeling of occupational change, which continues after the closure of the device. Conclusions: the occupational balance is altered by modifying daily routine and environmental conditions. A clear resilience can be observed when conditions change and/or return to normal. (AU)

The BAF complex inhibitor pyrimethamine reverses HIV-1 latency in people with HIV-1 on antiretroviral therapy.

Reactivation of the latent HIV-1 reservoir is a first step toward triggering reservoir decay. Here, we investigated the impact of the BAF complex inhibitor pyrimethamine on the reservoir of people living with HIV-1 (PLWH). Twenty-eight PLWH on suppressive antiretroviral therapy were randomized (1:1:1:1 ratio) to receive pyrimethamine, valproic acid, both, or no intervention for 14 days. The primary end point was change in cell-associated unspliced (CA US) HIV-1 RNA at days 0 and 14. We observed a rapid, modest, and significant increase in (CA US) HIV-1 RNA in response to pyrimethamine exposure, which persisted throughout treatment and follow-up. Valproic acid treatment alone did not increase (CA US) HIV-1 RNA or augment the effect of pyrimethamine. Pyrimethamine treatment did not result in a reduction in the size of the inducible reservoir. These data demonstrate that the licensed drug pyrimethamine can be repurposed as a BAF complex inhibitor to reverse HIV-1 latency in vivo in PLWH, substantiating its potential advancement in clinical studies.

HibeRNAtion: HIV-1 RNA Metabolism and Viral Latency.

HIV-1 infection remains non-curative due to the latent reservoir, primarily a small pool of resting memory CD4+ T cells bearing replication-competent provirus. Pharmacological reversal of HIV-1 latency followed by intrinsic or extrinsic cell killing has been proposed as a promising strategy to target and eliminate HIV-1 viral reservoirs. Latency reversing agents have been extensively studied for their role in reactivating HIV-1 transcription in vivo, although no permanent reduction of the viral reservoir has been observed thus far. This is partly due to the complex nature of latency, which involves strict intrinsic regulation at multiple levels at transcription and RNA processing. Still, the molecular mechanisms that control HIV-1 latency establishment and maintenance have been almost exclusively studied in the context of chromatin remodeling, transcription initiation and elongation and most known LRAs target LTR-driven transcription by manipulating these. RNA metabolism is a largely understudies but critical mechanistic step in HIV-1 gene expression and latency. In this review we provide an update on current knowledge on the role of RNA processing mechanisms in viral gene expression and latency and speculate on the possible manipulation of these pathways as a therapeutic target for future cure studies.

Catchet-MS identifies IKZF1-targeting thalidomide analogues as novel HIV-1 latency reversal agents.

A major pharmacological strategy toward HIV cure aims to reverse latency in infected cells as a first step leading to their elimination. While the unbiased identification of molecular targets physically associated with the latent HIV-1 provirus would be highly valuable to unravel the molecular determinants of HIV-1 transcriptional repression and latency reversal, due to technical limitations, this has been challenging. Here we use a dCas9 targeted chromatin and histone enrichment strategy coupled to mass spectrometry (Catchet-MS) to probe the differential protein composition of the latent and activated HIV-1 5'LTR. Catchet-MS identified known and novel latent 5'LTR-associated host factors. Among these, IKZF1 is a novel HIV-1 transcriptional repressor, required for Polycomb Repressive Complex 2 recruitment to the LTR. We find the clinically advanced thalidomide analogue iberdomide, and the FDA approved analogues lenalidomide and pomalidomide, to be novel LRAs. We demonstrate that, by targeting IKZF1 for degradation, these compounds reverse HIV-1 latency in CD4+ T-cells isolated from virally suppressed people living with HIV-1 and that they are able to synergize with other known LRAs.

A Two-Color Haploid Genetic Screen Identifies Novel Host Factors Involved in HIV-1 Latency.

To identify novel host factors as putative targets to reverse HIV-1 latency, we performed an insertional mutagenesis genetic screen in a latent HIV-1 infected pseudohaploid KBM7 cell line (Hap-Lat). Following mutagenesis, insertions were mapped to the genome, and bioinformatic analysis resulted in the identification of 69 candidate host genes involved in maintaining HIV-1 latency. A select set of candidate genes was functionally validated using short hairpin RNA (shRNA)-mediated depletion in latent HIV-1 infected J-Lat A2 and 11.1 T cell lines. We confirmed ADK, CHD9, CMSS1, EVI2B, EXOSC8, FAM19A, GRIK5, IRF2BP2, NF1, and USP15 as novel host factors involved in the maintenance of HIV-1 latency. Chromatin immunoprecipitation assays indicated that CHD9, a chromodomain helicase DNA-binding protein, maintains HIV-1 latency via direct association with the HIV-1 5' long terminal repeat (LTR), and its depletion results in increased histone acetylation at the HIV-1 promoter, concomitant with HIV-1 latency reversal. FDA-approved inhibitors 5-iodotubercidin, trametinib, and topiramate, targeting ADK, NF1, and GRIK5, respectively, were characterized for their latency reversal potential. While 5-iodotubercidin exhibited significant cytotoxicity in both J-Lat and primary CD4+ T cells, trametinib reversed latency in J-Lat cells but not in latent HIV-1 infected primary CD4+ T cells. Importantly, topiramate reversed latency in cell line models, in latently infected primary CD4+ T cells, and crucially in CD4+ T cells from three people living with HIV-1 (PLWH) under suppressive antiretroviral therapy, without inducing T cell activation or significant toxicity. Thus, using an adaptation of a haploid forward genetic screen, we identified novel and druggable host factors contributing to HIV-1 latency. IMPORTANCE A reservoir of latent HIV-1 infected cells persists in the presence of combination antiretroviral therapy (cART), representing a major obstacle for viral eradication. Reactivation of the latent HIV-1 provirus is part of curative strategies which aim to promote clearance of the infected cells. Using a two-color haploid screen, we identified 69 candidate genes as latency-maintaining host factors and functionally validated a subset of 10 of those in additional T-cell-based cell line models of HIV-1 latency. We further demonstrated that CHD9 is associated with HIV-1's promoter, the 5' LTR, while this association is lost upon reactivation. Additionally, we characterized the latency reversal potential of FDA compounds targeting ADK, NF1, and GRIK5 and identify the GRIK5 inhibitor topiramate as a viable latency reversal agent with clinical potential.

External validation of first trimester combined screening for pre-eclampsia in Brazil: An observational study.

OBJECTIVE: To validate a combined algorithm for early prediction of pre-eclampsia (PE) in the Brazilian population. STUDY DESIGN: This is an unplanned secondary analysis of a cohort study. Consecutive singleton pregnancies undergoing first-trimester screening for PE involving examination of maternal characteristics, medical history, and biophysical markers were considered eligible. Women were classified as low-or high-risk using a cutoff of 1/200, but the individual risk was not used to dictate management, as aspirin prophylaxis was given to women based solely on clinical risk factors. Receiver-operating characteristics (ROC) curves for PE, preterm PE(PE < 37) and early 34(PE < 34) were constructed and detection rates(DR) and false-positive rates(FPR) were calculated, adjusting for the effect of aspirin. Propensity score analysis was utilized to account for possible confounding by indication. MAIN OUTCOME MEASURES: Screening performance and PE rates. RESULTS: Among 1695 women, 323(19.1%) were classified as high-risk for PE and 1372(80.9%) were considered low-risk. Aspirin use was registered in 62(3.7%) in the high-risk group and 33(1.9%) in the low-risk group. There were 164(9.7%) women who developed PE, including 41(2.4%) with PE < 37 and 18(1.1%) PE < 34.Subgroups with aspirin had higher incidence of PE, suggest confounding by indication. The algorithm had an AUC of 0.87, DR of 72% for PE < 34; an AUC of 0.8, DR of 59% for PE < 37, both with FPR of 18%. Accounting for effect of aspirin, we observed an improvement in DR of PE < 37 to 67%. CONCLUSION: Using combined predictive algorithm for preterm PE prediction is feasible in clinical practice in low/middle-income countries. Aspirin use needs to be accounted for when evaluating the performance of screening.

Selective cell death in HIV-1-infected cells by DDX3 inhibitors leads to depletion of the inducible reservoir.

An innovative approach to eliminate HIV-1-infected cells emerging out of latency, the major hurdle to HIV-1 cure, is to pharmacologically reactivate viral expression and concomitantly trigger intracellular pro-apoptotic pathways in order to selectively induce cell death (ICD) of infected cells, without reliance on the extracellular immune system. In this work, we demonstrate the effect of DDX3 inhibitors on selectively inducing cell death in latent HIV-1-infected cell lines, primary CD4+ T cells and in CD4+ T cells from cART-suppressed people living with HIV-1 (PLWHIV). We used single-cell FISH-Flow technology to characterise the contribution of viral RNA to inducing cell death. The pharmacological targeting of DDX3 induced HIV-1 RNA expression, resulting in phosphorylation of IRF3 and upregulation of IFNß. DDX3 inhibition also resulted in the downregulation of BIRC5, critical to cell survival during HIV-1 infection, and selectively induced apoptosis in viral RNA-expressing CD4+ T cells but not bystander cells. DDX3 inhibitor treatment of CD4+ T cells from PLWHIV resulted in an approximately 50% reduction of the inducible latent HIV-1 reservoir by quantitation of HIV-1 RNA, by FISH-Flow, RT-qPCR and TILDA. This study provides proof of concept for pharmacological reversal of latency coupled to induction of apoptosis towards the elimination of the inducible reservoir.

Gliotoxin, identified from a screen of fungal metabolites, disrupts 7SK snRNP, releases P-TEFb, and reverses HIV-1 latency.

A leading pharmacological strategy toward HIV cure requires "shock" or activation of HIV gene expression in latently infected cells with latency reversal agents (LRAs) followed by their subsequent clearance. In a screen for novel LRAs, we used fungal secondary metabolites as a source of bioactive molecules. Using orthogonal mass spectrometry (MS) coupled to latency reversal bioassays, we identified gliotoxin (GTX) as a novel LRA. GTX significantly induced HIV-1 gene expression in latent ex vivo infected primary cells and in CD4+ T cells from all aviremic HIV-1+ participants. RNA sequencing identified 7SK RNA, the scaffold of the positive transcription elongation factor b (P-TEFb) inhibitory 7SK small nuclear ribonucleoprotein (snRNP) complex, to be significantly reduced upon GTX treatment of CD4+ T cells. GTX directly disrupted 7SK snRNP by targeting La-related protein 7 (LARP7), releasing active P-TEFb, which phosphorylated RNA polymerase II (Pol II) C-terminal domain (CTD), inducing HIV transcription.

Management of ectopic pregnancies with poor prognosis through ultrasound guided intrasacular injection of methotrexate, series of 14 cases.

PURPOSE: The purpose of this study is to describe our experience in cases of tubal ectopic pregnancy with heartbeat, abdominal, interstitial (corneal) and cervical ectopic pregnancies treated with intrasacular injection of methotrexate (MTX) administered under ultrasound guidance associated with a single systemic dose of MTX. METHODS: Descriptive retrospective study of 14 cases of extrauterine pregnancies treated with intrasacular injection of MTX under ultrasound control, attended in the Maternal-Fetal Medicine Unit of the Miguel Servet University Hospital, in Zaragoza, Spain, between January of 2009 and June of 2010. RESULTS: Of the 14 ectopic pregnancies, 7 were tubal with heartbeat, 3 cornual, 2 cervical and 2 abdominal. The average gestational age was 7 + 3 weeks and the average ß-hCG value on the date of puncture was 22,885.69 mIU/mL. Surgical treatment was required in two cases, the first due to post-puncture haemoperitoneum and the second as a consequence of the rupture of the corneal ectopic pregnancy. In post-treatment monitoring, an asymptomatic increase of ß-hCG on the seventh day post-puncture was observed in two cases. The success rate of the treatment was 92.96%. CONCLUSIONS: Ultrasound guided intrasacular injection of MTX associated with a systemic dose adjusted to the body surface of the patient is a minimally invasive, safe and effective treatment in the cases of tubal ectopic pregnancy with heartbeat, abdominal, cornual or cervical ectopic pregnancy.

Gersad, boletín informativo para directivos de la Salud de Camagüey / Gersad, news bulletin for executives of Camagüey´s health care institutions

Se presentan los resultados de una investigación de innovación tecnológica desarrollada entre marzo y septiembre de 2009, en la Dirección Provincial de Salud de Camagüey. Se aplicó una encuesta a los directivos del territorio para diseñar un boletín que les informe sobre los procesos gerenciales del Sistema Nacional de Salud cubano. Se estableció una estrategia para la diseminación del mismo a través del correo electrónico y de su inclusión en la Web. Los destinatarios expresaron elevados niveles de satisfacción con el producto. Se recomienda incluir el boletín en el sitio Web de la Dirección Provincial de Salud de Camagüey (AU)

This paper presents the results of a technological innovation research carried out at Camagüey´s Health Care Provincial Department, from March to September 2009. Territorial health care executives answered a survey aimed to designing a news bulletin to inform them about the managing processes of Cuba´s National System of Health. A strategy for its circulation through emails and publication in the Internet was developed. Results showed high levels of satisfaction with the product. As recommendations, the bulletin should be included in the website of Camagüey´s Health Care Provincial Department (AU)

Gersad, boletín informativo para directivos de la Salud de Camagüey / Gersad, news bulletin for executives of Camagüey´s health care institutions

Se presentan los resultados de una investigación de innovación tecnológica desarrollada entre marzo y septiembre de 2009, en la Dirección Provincial de Salud de Camagüey. Se aplicó una encuesta a los directivos del territorio para diseñar un boletín que les informe sobre los procesos gerenciales del Sistema Nacional de Salud cubano. Se estableció una estrategia para la diseminación del mismo a través del correo electrónico y de su inclusión en la Web. Los destinatarios expresaron elevados niveles de satisfacción con el producto. Se recomienda incluir el boletín en el sitio Web de la Dirección Provincial de Salud de Camagüey, así como dar continuidad y enriquecimiento al producto a través del estudio periódico de las necesidades de información de los dirigentes de la Salud en la provincia.

This paper presents the results of a technological innovation research carried out at Camagüey´s Health Care Provincial Department, from March to September 2009. Territorial health care executives answered a survey aimed to designing a news bulletin to inform them about the managing processes of Cuba´s National System of Health. A strategy for its circulation through emails and publication in the Internet was developed. Results showed high levels of satisfaction with the product. As recommendations, the bulletin should be included in the website of Camagüey´s Health Care Provincial Department and given continuation and enrichment by means of periodically studying the executives information needs.