The East African Community mobile laboratory network prepares for monkeypox outbreaks.

In response to the largest recorded monkeypox virus outbreak outside of endemic Central and Western Africa, the East African Community (EAC), in cooperation with the Bernhard-Nocht- Institute for Tropical Medicine, coordinated an emergency monkeypox diagnostic training for the East African Region. As of June 2022, the Democratic Republic of Congo reported a steady increase of suspected monkeypox cases, increasing the risk of spill-over into the remaining six EAC Partner States. Within the existing EAC Mobile Laboratories project, laboratory experts of the National Public Health Laboratories of the remaining six EAC Partner States (Burundi, Rwanda, Tanzania, Kenya, Uganda, and South Sudan) participated in the workshop and were trained in the reception of suspect samples, DNA extraction and diagnosis using real-time polymerase chain reaction (RT-PCR). The EAC region is now equipped with the tools to prepare and rapidly respond to any emerging monkeypox outbreak.

Cardiopulmonary interactions in left heart failure.

The primary impact of ventilation and ventilatory efforts on left ventricular (LV) function in left ventricular dysfunction relate to how changes in intrathoracic pressure (ITP) alter the pressure gradients for venous return into the chest and LV ejection out of the chest. Spontaneous inspiratory efforts by decreasing ITP increase both of these pressure gradients increasing venous blood flow and impeding LV ejection resulting in increased intrathoracic blood volume. In severe heart failure states when lung compliance is reduced, or airway resistance is increased these negative swings in ITP can be exacerbated leading to LV failure and acute cardiogenic pulmonary edema. By merely reversing these negative swings in ITP by the use of non-invasive continuous positive airway pressure (CPAP), these profoundly detrimental forces can be immediately reversed, and cardiovascular stability can be restored in moments. This forms the clinical rationale for the immediate use of CPAP for the treatment of acute cardiogenic pulmonary edema. Increasing ITP during positive pressure ventilation decreases the pressure gradients for venous return and LV ejection decreasing intrathoracic blood volume. In a hypovolemic patient even with LV dysfunction this can result in hypotension due to inadequate LV preload. Minor increases in ITP as occur using pressure-limited positive-pressure ventilation primarily reverse the increased LV afterload of negative swings in ITP and if fluid overload was already present, minimally alter cardiac output. The effect of changes in lung volume on LV function are related primarily to its effects on right ventricular (RV) function through changes in pulmonary vascular resistance and overdistention (hyperinflation). In acute lung injury with alveolar collapse, positive pressure ventilation may reduce pulmonary vascular resistance if alveolar recruitment predominates. Hyperinflation, however, impedes diastolic filling while simultaneously increasing pulmonary vascular resistance. Thus, increasing lung volume can reduce RV afterload by reversing hypoxic pulmonary vasoconstriction or increase afterload by overdistention. Hyperinflation can also impede RV filling. All of these processes can be readily identified at the bedside using echocardiography.

A high-fat diet increases hepatic mitochondrial turnover through restricted acetylation in a NAFLD mouse model.

Lysine acetylation of proteins has emerged as a key posttranslational modification (PTM) that regulates mitochondrial metabolism. Acetylation may regulate energy metabolism by inhibiting and affecting the stability of metabolic enzymes and oxidative phosphorylation (OxPhos) subunits. Although protein turnover can be easily measured, due to the low abundance of modified proteins, it has been difficult to evaluate the effect of acetylation on the stability of proteins in vivo. We applied 2H2O-metabolic labeling coupled with immunoaffinity and high-resolution mass spectrometry method to measure the stability of acetylated proteins in mouse liver based on their turnover rates. As a proof-of-concept, we assessed the consequence of high-fat diet (HFD)-induced altered acetylation in protein turnover in LDL receptor-deficient (LDLR-/-) mice susceptible to diet-induced nonalcoholic fatty liver disease (NAFLD). HFD feeding for 12 wk led to steatosis, the early stage of NAFLD. A significant reduction in acetylation of hepatic proteins was observed in NAFLD mice, based on immunoblot analysis and label-free quantification with mass spectrometry. Compared with control mice on a normal diet, NAFLD mice had overall increased turnover rates of hepatic proteins, including mitochondrial metabolic enzymes (0.159 ± 0.079 vs. 0.132 ± 0.068 day-1), suggesting their reduced stability. Also, acetylated proteins had slower turnover rates (increased stability) than native proteins in both groups (0.096 ± 0.056 vs. 0.170 ± 0.059 day-1 in control, and 0.111 ± 0.050 vs. 0.208 ± 0.074 day-1 in NAFLD). Furthermore, association analysis revealed a relationship between the HFD-induced decrease in acetylation and increased turnover rates for hepatic proteins in NAFLD mice. These changes were associated with increased expressions of the hepatic mitochondrial transcriptional factor (TFAM) and complex II subunit without any changes to other OxPhos proteins, suggesting that enhanced mitochondrial biogenesis prevented restricted acetylation-mediated depletion of mitochondrial proteins. We conclude that decreased acetylation of mitochondrial proteins may contribute to adaptive improved hepatic mitochondrial function in the early stages of NAFLD.NEW & NOTEWORTHY This is the first method to quantify acetylome dynamics in vivo. This method revealed acetylation-mediated altered hepatic mitochondrial protein turnover in response to a high-fat diet in a mouse model of NAFLD.

Precision targeting of autoantigen-specific B cells in muscle-specific tyrosine kinase myasthenia gravis with chimeric autoantibody receptor T cells.

Muscle-specific tyrosine kinase myasthenia gravis (MuSK MG) is an autoimmune disease that causes life-threatening muscle weakness due to anti-MuSK autoantibodies that disrupt neuromuscular junction signaling. To avoid chronic immunosuppression from current therapies, we engineered T cells to express a MuSK chimeric autoantibody receptor with CD137-CD3ζ signaling domains (MuSK-CAART) for precision targeting of B cells expressing anti-MuSK autoantibodies. MuSK-CAART demonstrated similar efficacy as anti-CD19 chimeric antigen receptor T cells for depletion of anti-MuSK B cells and retained cytolytic activity in the presence of soluble anti-MuSK antibodies. In an experimental autoimmune MG mouse model, MuSK-CAART reduced anti-MuSK IgG without decreasing B cells or total IgG levels, reflecting MuSK-specific B cell depletion. Specific off-target interactions of MuSK-CAART were not identified in vivo, in primary human cell screens or by high-throughput human membrane proteome array. These data contributed to an investigational new drug application and phase 1 clinical study design for MuSK-CAART for the treatment of MuSK autoantibody-positive MG.

Metabolic labeling unveils alterations in the turnover of HDL-associated proteins during diabetes progression in mice.

Several aspects of diabetes pathophysiology and complications result from hyperglycemia-induced alterations in the structure and function of plasma proteins. Furthermore, insulin has a significant influence on protein metabolism by affecting both the synthesis and degradation of proteins in various tissues. To understand the role of progressive hyperglycemia on plasma proteins, in this study, we measured the turnover rates of high-density lipoprotein (HDL)-associated proteins in control (chow diet), prediabetic [a high-fat diet (HFD) for 8 wk] or diabetic [HFD for 8 wk with low-dose streptozotocin (HFD + STZ) in weeks 5-8 of HFD] C57BL/6J mice using heavy water (2H2O)-based metabolic labeling approach. Compared with control mice, HFD and HFD + STZ mice showed elevations of fasting plasma glucose levels in the prediabetic and diabetic range, respectively. Furthermore, the HFD and HFD + STZ mice showed increased hepatic triglyceride (TG) levels, total plasma cholesterol, and plasma TGs. The kinetics of 40 proteins were quantified using the proteome dynamics method, which revealed an increase in the fractional synthesis rate (FSR) of HDL-associated proteins in the prediabetic mice compared with control mice, and a decrease in FSR in the diabetic mice. The pathway analysis revealed that proteins with altered turnover rates were involved in acute-phase response, lipid metabolism, and coagulation. In conclusion, prediabetes and diabetes have distinct effects on the turnover rates of HDL proteins. These findings suggest that an early dysregulation of the HDL proteome dynamics can provide mechanistic insights into the changes in protein levels in these conditions.NEW & NOTEWORTHY This study is the first to examine the role of gradual hyperglycemia during diabetes disease progression on HDL-associated protein dynamics in the prediabetes and diabetic mice. Our results show that the fractional synthesis rate of HDL-associated proteins increased in the prediabetic mice whereas it decreased in the diabetic mice compared with control mice. These kinetic changes can help to elucidate the mechanism of altered protein levels and HDL dysfunction during diabetes disease progression.

Integrated In Silico Analyses Identify PUF60 and SF3A3 as New Spliceosome-Related Breast Cancer RNA-Binding Proteins.

More women are diagnosed with breast cancer (BC) than any other type of cancer. Although large-scale efforts have completely redefined cancer, a cure remains unattainable. In that respect, new molecular functions of the cell should be investigated, such as post-transcriptional regulation. RNA-binding proteins (RBPs) are emerging as critical post-transcriptional modulators of tumorigenesis, but only a few have clear roles in BC. To recognize new putative breast cancer RNA-binding proteins, we performed integrated in silico analyses of all human RBPs (n = 1392) in three major cancer databases and identified five putative BC RBPs (PUF60, TFRC, KPNB1, NSF, and SF3A3), which showed robust oncogenic features related to their genomic alterations, immunohistochemical changes, high interconnectivity with cancer driver genes (CDGs), and tumor vulnerabilities. Interestingly, some of these RBPs have never been studied in BC, but their oncogenic functions have been described in other cancer types. Subsequent analyses revealed PUF60 and SF3A3 as central elements of a spliceosome-related cluster involving RBPs and CDGs. Further research should focus on the mechanisms by which these proteins could promote breast tumorigenesis, with the potential to reveal new therapeutic pathways along with novel drug-development strategies.

Morphological, molecular and ecological characterization of a native isolate of Steinernema feltiae (Rhabditida: Steinernematidae) from southern Chile.

BACKGROUND: Steinernema feltiae is an entomopathogenic nematode used in biological control programs with a global distribution. Populations of this species show phenotypic plasticity derived from local adaptation and vary in different traits, such as location and host penetration. The aim of this work was to describe a Chilean isolate of this nematode species, using integrative approaches. METHODS: Nematode morphological and morphometric studies were conducted along with molecular analysis of nuclear genes. The symbiotic bacterium was also identified by sequencing the 16S rRNA gene. Some ecological characteristics were described, including the temperature requirements for the nematode life cycle and the effect of soil water content for optimal reproduction. RESULTS: Morphometric characterization revealed a large intra-specific variability. The isolate identity was also corroborated with the analysis of nuclear genes. Based on the 16S gene, its symbiont bacteria, Xenorhabdus bovienii, was identified. The lowest, optimal and highest temperatures found to limit the infestation and reproduction on Galleria mellonella were 10, 20 and 30 °C, respectively; the emergence from the host larvae occurred approximately 10 days after inoculation. Differences were observed in offspring, and 120 infective juveniles (IJ)/larva was the most prolific dose at 20 °C. The soil water content did not affect the number of IJ invaders, penetration efficacy and IJ emergence time or offspring per larva, but it caused a delay in achieving full mortality at the permanent wilting point with respect to saturation and field capacity. CONCLUSIONS: For the first time, a Chilean isolate of S. feltiae is described in detail considering morphological, molecular and ecological aspects. The isolate was shown to be efficient in soil containing water, with optimal temperatures ranging from 15 to 25 °C for host infestation and production of an abundant offspring; these characteristics would allow its potential use as control agents in a wide geographical area of the country.

Measuring acetyl-CoA and acetylated histone turnover in vivo: Effect of a high fat diet.

Cellular availability of acetyl-CoA, a central intermediate of metabolism, regulates histone acetylation. The impact of a high-fat diet (HFD) on the turnover rates of acetyl-CoA and acetylated histones is unknown. We developed a method for simultaneous measurement of acetyl-CoA and acetylated histones kinetics using a single 2H2O tracer, and used it to examine effect of HFD-induced perturbations on hepatic histone acetylation in LDLR-/- mice, a mouse model of non-alcoholic fatty liver disease (NAFLD). Mice were given 2H2O in the drinking water and the kinetics of hepatic acetyl-CoA, histones, and acetylated histones were quantified based on their 2H-labeling. Consumption of a high fat Western-diet (WD) for twelve weeks led to decreased acetylation of hepatic histones (p< 0.05), as compared to a control diet. These changes were associated with 1.5-3-fold increased turnover rates of histones without any change in acetyl-CoA flux. Acetylation significantly reduced the stability of histones and the turnover rates of acetylated peptides were correlated with the number of acetyl groups in neighboring lysine sites. We conclude that 2H2O-method can be used to study metabolically controlled histone acetylation and acetylated histone turnover in vivo.

Distinct effects of complement and of NLRP3- and non-NLRP3 inflammasomes for choroidal neovascularization.

NLRP3 inflammasome activation and complement-mediated inflammation have been implicated in promoting choroidal neovascularization (CNV) in age-related macular degeneration (AMD), but central questions regarding their contributions to AMD pathogenesis remain unanswered. Key open questions are (1) whether NLRP3 inflammasome activation mainly in retinal pigment epithelium (RPE) or rather in non-RPE cells promotes CNV, (2) whether inflammasome activation in CNV occurs via NLRP3 or also through NLRP3-independent mechanisms, and (3) whether complement activation induces inflammasome activation in CNV. Here we show in a neovascular AMD mouse model that NLRP3 inflammasome activation in non-RPE cells but not in RPE cells promotes CNV. We demonstrate that both NLRP3-dependent and NLRP3-independent inflammasome activation mechanisms induce CNV. Finally, we find that complement and inflammasomes promote CNV through independent mechanisms. Our findings uncover an unexpected role of non-NLRP3 inflammasomes for CNV and suggest that combination therapies targeting inflammasomes and complement may offer synergistic benefits to inhibit CNV.

Incidence and Outcomes of Dermatofibrosarcoma Protuberans in the US Pediatric Population.

BACKGROUND: Dermatofibrosarcoma protuberans (DFSP) is a low-grade soft tissue sarcoma. In the pediatric population, DFSP is exceedingly rare. Aim of this study was to describe the epidemiology and clinical outcomes in a large pediatric cohort. METHODS: Surveillance, Epidemiology, and End Results (SEER) database (1973-2010) was analyzed for all patients with dermatofibrosarcoma occurring in patients <20 years of age. Data were extracted based on age, gender, race, anatomic site, histology, stage, treatment modalities, and survival. Incidence rates were standardized to the 2000 US population. RESULTS: A total of 451 patients were identified. Overall annual incidence was 0.10 per 100,000. Incidence was highest among black children and adolescents (ages 15 to 19 years). Trunk was most common site, followed by extremities. Head and neck region was least common site (P < 0.05). Majority (54%) of patients presented with localized disease. Overall, 95% underwent surgery. Only 2.2% were treated with perioperative radiation therapy. Overall prognosis was favorable with 5-year overall survival (OS) of 100%, 15-year OS of 98%, and 30-year OS of 97%. Median survival was 117 months. Male patients had lower 15- and 30-year OS compared with females (P < 0.05). CONCLUSION: Pediatric DFSP has lower incidence but similar clinical characteristics to adults. Incidence is higher in black children and in the trunk region. While prognosis is favorable, male sex is associated with decreased OS.

Jubilación: cambios percibidos en relación a rutinas y roles en adultos mayores no institucionalizados de la cuidad de Punta Arenas / Retirement: perceived changes in relation to routine and roles in non-institutionalized elder people from Punta Arenas city

Esta investigación tiene como principal propósito identificar las percepciones de adultos mayores no institucionalizados de la ciudad de Punta Arenas, respecto a los cambios producidos en sus rutinas y roles luego del proceso de jubilación, desde el enfoque de la ciencia de la ocupación y la terapia ocupacional en la región de Magallanes y Antártica Chilena. La metodología empleada tiene un enfoque cualitativo. Para la recolección de datos, se utilizó una entrevista semiestructurada y el análisis de información se basa en la teoría fundamentada de Glaser & Strauss (1967). Como principales resultados, se obtiene que la existencia de cambios en las rutinas y roles, incluyen percepciones positivas y negativas asociadas. Además, se identifica que en el desempeño de rutinas y roles de las personas, existe una estrecha relación. Finalmente, se puede concluir que para los adultos mayores, la adaptación a dichos cambios producidos después de vivir el proceso de jubilación, depende de las oportunidades de involucramiento en actividades significativas, junto con la posibilidad de contar con redes de apoyo.

This research has as main purpose to identify the perception of non institutionalized elderly people from Punta Arenas city, on the matter of all changes produce in their routines and roles after the retirement process; seeing from the approach of occupation’s science and occupational therapy in the Magellan and Antarctic Chilean region. The methodology used has a qualitative approach. It was used a semi-structured interview for the data collection, and the analysis information is based upon the grounded theory of Glaser & Strauss (1967). As main results, it was obtained that the existence of changes in the routines and roles, include positive and negative perception associated. Furthermore, it was identified that a narrow relationship exists in the performance of the routines and roles of the people. Lastly, it can be concluded that for elderly people, the adaptation to those changes produced after living the retirement process depends on the opportunities of getting involved in meaningful activities, together with the possibility of counting with support networks.

Role of zinc in catalytic activity of carbonic anhydrase IX.

The carbonic anhydrases (CAs) in the α class are zinc-dependent metalloenzymes. Previous studies have reported that recombinant forms of carbonic anhydrase IX (CAIX), a membrane-bound form of CA expressed in solid tumors, appear to be activated by low levels of zinc independent of its well-studied role at the catalytic site. In this study, we sought to determine if CAIX is stimulated by zinc in its native environment. MDA-MB-231 breast cancer cells express CAIX in response to hypoxia. We compared CAIX activity associated with membrane ghosts isolated from hypoxic cells with that in intact hypoxic cells. We measured CA activity directly using (18)O exchange from (13)CO(2) into water determined by membrane inlet mass spectrometry. In membrane ghosts, there was little effect of zinc at low concentrations on CAIX activity, although at high concentration zinc was inhibitory. In intact cells, zinc had no significant effect on CAIX activity. This suggests that there is an appreciable decrease in sensitivity to zinc when CAIX is in its natural membrane milieu compared to the purified forms.

Anticariogenic activity of the active fraction from isertia laevis against S. mutans and S. sobrinus: comparison of two extraction methods

Dental caries is considered a multi-factorial, infectious, chronic, localized, post-eruptive, transmissible disease that leads to the destruction of dental hard tissue. The recognition of Streptococcus mutans as the major bacterial species involved in dental caries has led to the implementation of prevention and control measures for eliminating or reducing it in oral cavity. The main goal of research on medicinal plants is the search for substances or compounds with antimicrobial activity. The aim of this study was to evaluate the antimicrobial activity of fractions obtained by two methods from Isertia laevis against S. mutans and S. sobrinus. The plant material was collected in Medina (Colombia), at an elevation of 550 meters above sea level. From the ethanol extract of leaves of I. laevis, fractions were obtained by two methods: extraction by column vacuum chromatography (CVC) and extraction by continuous liquid / liquid partitioning (CLLP). The evaluation of the antimicrobial activity of fractions against S. mutans and S. sobrinus was performed by well diffusion and bioautography assays. From the CVC technique, only the methanol and methanol-dichloromethane fractions showed activity against S. mutans and S. sobrinus, with a minimum inhibitory concentration of 2 mg/well. From the CLLP technique, only the dichloromethane fraction showed activity against both microorganisms, with a minimum inhibitory concentration of 1 mg/well. Compounds C1 and C2 were isolated from the three active fractions, and showed a minimum inhibitory concentration of 0.4 mg/well for S. mutans and S. sobrinus, with zones of inhibition measuring 6.5 and 6.2 mm, respectively. In conclusion: 1) the three active fractions of I. laevis showed activity against S. mutans and S. sobrinus, 2) compounds C1 and C2 were present equally in the three active fractions showing activity against the two bacteria, 3) compounds C1 and C2 may be triterpenoid and/or steroidal saponin structures, and 4) the two extraction methods lead equally to obtaining the active fractions.

La caries dental es considerada una enfermedad infecciosa multifactorial, cronica, localizada, poseruptiva y transmisible que conlleva a la destruccion del tejido dental duro. El claro reconocimiento de Streptococcus mutans como la principal especie bacteriana implicada en caries dental, ha conducido a la implementacion de medidas de prevencion y control para la eliminacion o disminucion de este microorganismo en cavidad oral. El objetivo fundamental de la investigacion en plantas medicinales, es la busqueda de sustancias o compuestos con actividad antimicrobiana para ser utilizadas en el control o prevencion de enfermedades infecciosas. En este sentido, en salud bucal muchas sustancias obtenidas de plantas han mostrado actividad antimicrobiana. El objetivo de este estudio fue evaluar la actividad antimicrobiana de fracciones obtenidas de la planta Isertia laevis mediante dos metodologias contra S. mutans y S. sobrinus. El material vegetal se colecto en el municipio de Medina (Cundinamarca- Colombia) situado a una altura de 550 metros sobre el nivel del mar. A partir del extracto etanolico de hojas de I. laevis se obtuvieron fracciones mediante dos metodologias, extraccion por cromatografia en columna al vacio (CCV) y extraccion por fraccionamiento liquido/liquido continuo (FLLC). La evaluacion de la actividad antimicrobiana de las fracciones frente a S. mutans y S. sobrinus se realizo por el metodo de difusion en pozo y bioautografico. De las fracciones obtenidas por CCV, solamente las fracciones metanol y metanol-diclorometano presentaron actividad antimicrobiana sobre S. mutans y S. sobrinus, con una concentracion minima inhibitoria de 2 mg/pozo. De las fracciones obtenidas por FLLC solamente la fraccion diclorometano presento actividad antimicrobiana sobre S. mutans y S. sobrinus, con una concentracion minima inhibitoria de 1 mg/pozo. De las 3 fracciones activas se aislaron los compuestos C1 y C2, que presentaron una concentracion minima inhibitoria de 0.4 mg/pozo tanto para S. mutans como para S. sobrinus con halos de inhibicion, respectivamente, de 6.5 y 6.2 mm. En conclusion, 1. Las fracciones metanol y diclorometano obtenidas por CCV y la fraccion diclorometano obtenida por FLLC de hojas de I. laevis presentaron actividad antimicrobiana sobre S. mutans y S. sobrinus; 2. Los compuestos C1 y C2 presentes por igual en las tres fracciones activas tuvieron accion inhibitoria sobre las dos bacterias en evaluacion; 3. Las pruebas quimicas cualitativas para los compuestos C1 y C2 indican que posiblemente corresponden a estructuras de saponinas triterpenicas y/o esteroidales; y 4. Las dos metodologias de extraccion conducen por igual a la obtencion de las fracciones activas.

Anticariogenic activity of the active fraction from Isertia laevis against S. mutans and S. sobrinus: comparison of two extraction methods.

UNLABELLED: Dental caries is considered a multi-factorial, infectious, chronic, localized, post-eruptive, transmissible disease that leads to the destruction of dental hard tissue. The recognition of Streptococcus mutans as the major bacterial species involved in dental caries has led to the implementation of prevention and control measures for eliminating or reducing it in oral cavity. The main goal of research on medicinal plants is the search for substances or compounds with antimicrobial activity. The aim of this study was to evaluate the antimicrobial activity of fractions obtained by two methods from Isertia laevis against S. mutans and S. sobrinus. The plant material was collected in Medina (Colombia), at an elevation of 550 meters above sea level. From the ethanol extract of leaves of I. laevis, fractions were obtained by two methods: extraction by column vacuum chromatography (CVC) and extraction by continuous liquid/liquid partitioning (CLLP). The evaluation of the antimicrobial activity of fractions against S. mutans and S. sobrinus was performed by well diffusion and bioautography assays. From the CVC technique, only the methanol and methanol-dichloromethane fractions showed activity against S. mutans and S. sobrinus, with a minimum inhibitory concentration of 2 mg/well. From the CLLP technique, only the dichloromethane fraction showed activity against both microorganisms, with a minimum inhibitory concentration of 1 mg/well. Compounds C1 and C2 were isolated from the three active fractions, and showed a minimum inhibitory concentration of 0.4 mg/well for S. mutans and S. sobrinus, with zones of inhibition measuring 6.5 and 6.2 mm, respectively. IN CONCLUSION: 1) the three active fractions of I. laevis showed activity against S. mutans and S. sobrinus, 2) compounds C1 and C2 were presen equally in the three active fractions showing activity against the two bacteria, 3) compounds C1 and C2 may be triterpenoid and/or steroidal saponin structures, and 4) the two extraction methods lead equally to obtaining the active fractions.