Stimulating VAPB-PTPIP51 ER-mitochondria tethering corrects FTD/ALS mutant TDP43 linked Ca2+ and synaptic defects.

Frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are clinically linked major neurodegenerative diseases. Notably, TAR DNA-binding protein-43 (TDP43) accumulations are hallmark pathologies of FTD/ALS and mutations in the gene encoding TDP43 cause familial FTD/ALS. There are no cures for FTD/ALS. FTD/ALS display damage to a broad range of physiological functions, many of which are regulated by signaling between the endoplasmic reticulum (ER) and mitochondria. This signaling is mediated by the VAPB-PTPIP51 tethering proteins that serve to recruit regions of ER to the mitochondrial surface so as to facilitate inter-organelle communications. Several studies have now shown that disrupted ER-mitochondria signaling including breaking of the VAPB-PTPIP51 tethers are features of FTD/ALS and that for TDP43 and other familial genetic FTD/ALS insults, this involves activation of glycogen kinase-3ß (GSK3ß). Such findings have prompted suggestions that correcting damage to ER-mitochondria signaling and the VAPB-PTPIP51 interaction may be broadly therapeutic. Here we provide evidence to support this notion. We show that overexpression of VAPB or PTPIP51 to enhance ER-mitochondria signaling corrects mutant TDP43 induced damage to inositol 1,4,5-trisphosphate (IP3) receptor delivery of Ca2+ to mitochondria which is a primary function of the VAPB-PTPIP51 tethers, and to synaptic function. Moreover, we show that ursodeoxycholic acid (UDCA), an FDA approved drug linked to FTD/ALS and other neurodegenerative diseases therapy and whose precise therapeutic target is unclear, corrects TDP43 linked damage to the VAPB-PTPIP51 interaction. We also show that this effect involves inhibition of TDP43 mediated activation of GSK3ß. Thus, correcting damage to the VAPB-PTPIP51 tethers may have therapeutic value for FTD/ALS and other age-related neurodegenerative diseases.

Using Redox Proteomics to Gain New Insights into Neurodegenerative Disease and Protein Modification.

Antioxidant defenses in biological systems ensure redox homeostasis, regulating baseline levels of reactive oxygen and nitrogen species (ROS and RNS). Oxidative stress (OS), characterized by a lack of antioxidant defenses or an elevation in ROS and RNS, may cause a modification of biomolecules, ROS being primarily absorbed by proteins. As a result of both genome and environment interactions, proteomics provides complete information about a cell's proteome, which changes continuously. Besides measuring protein expression levels, proteomics can also be used to identify protein modifications, localizations, the effects of added agents, and the interactions between proteins. Several oxidative processes are frequently used to modify proteins post-translationally, including carbonylation, oxidation of amino acid side chains, glycation, or lipid peroxidation, which produces highly reactive alkenals. Reactive alkenals, such as 4-hydroxy-2-nonenal, are added to cysteine (Cys), lysine (Lys), or histidine (His) residues by a Michael addition, and tyrosine (Tyr) residues are nitrated and Cys residues are nitrosylated by a Michael addition. Oxidative and nitrosative stress have been implicated in many neurodegenerative diseases as a result of oxidative damage to the brain, which may be especially vulnerable due to the large consumption of dioxygen. Therefore, the current methods applied for the detection, identification, and quantification in redox proteomics are of great interest. This review describes the main protein modifications classified as chemical reactions. Finally, we discuss the importance of redox proteomics to health and describe the analytical methods used in redox proteomics.

A multidimensional evaluation of the effects of sweetener selection and UV-C treatment on orange juice and pectin-based confectionery gels.

BACKGROUND: Consumers are seeking healthier alternatives to traditional confectioneries. They value the use of sugar replacers, more natural ingredients and/or environmentally friendly preservation technologies. UV-C light is considered an emerging alternative to thermal pasteurization that leaves no residue and requires minimal energy. The present study aimed to investigate the effect of novel sweetener combinations and juice UV-C assisted by mild heat treatment (UV-C/H) on the physicochemical, microbiological, morphological, rheological and sensory properties of orange juice pectin-based confectioneries stored at 5 ± 1 °C for 35 days. RESULTS: For orange juice processing, UV-C/H (pilot-scale Dean-flow reactor; 892 mJ cm-2 ; 50 ± 1 °C) and thermal (T-coil, 80 °C; 6 min) treatments were used. Low-calorie confectionery gels were elaborated using the treated juices, low-methoxyl pectin and various sweetener combinations. UV-C/H and T-coil effectively inactivated juice native microbiota. The proposed formulations, derived from a previous Box-Behnken optimization study, included partial (F1: 3%-sucrose-S + 0.019%-rebaudioside-A-RA) or complete sucrose replacement (F2: 5.5%-erythritol-E + 0.019%-RA), and one control (C:10%-S). In general, the microbiota of the gels prepared with the UV-C/H or T-coil treated juices did not recover during storage. The physicochemical and mechanical parameters of the formulations were significantly influenced by the choice of sweetener and the duration of storage. The gel surface got smoother and had fewer holes when the sucrose level dropped, according to a scanning electron microscopy study. The UV-C/H-treated samples did not differ in acceptability, whereas the measured sensory attributes approached ideal levels. F1 and F2 showed distinctive temporal-dominance-of-sensations profiles, mainly dominated by sweetness and orange taste, respectively. However, consumers perceived sourness and astringency in C during consumption. CONCLUSION: The present study provides significant evidence in support of the development of confectionery gels F1 and F2 made from fruit juice treated by UV-C light assisted by mild heat and combinations of sucrose-alternative sweeteners. In terms of the properties investigated, these confectionery gels were comparable to, or even outperformed the full-sucrose option. © 2023 Society of Chemical Industry.

Implementation of Video Blood Pressure Visits in the Veterans Health Administration.

Introduction: Almost half of veterans (44.6%) seen in the U.S. Department of Veterans Affairs outpatient setting are diagnosed with hypertension (HTN). Because of the widespread nature of HTN, use of virtual visits has the potential to improve blood pressure (BP) management. This evaluation assessed the effectiveness of video blood pressure visits (VBPVs) in the management of HTN in veterans enrolled in Veterans Health Administration primary care. Methods: The program was implemented within the existing veteran-centered medical home. VBPVs are scheduled where the nurse observes veterans taking their BP and provides teaching or counseling. A national training curriculum was delivered to local nurse champions through Microsoft Teams. We analyzed improvement in BP over a 2-year period. We also captured actions taken by nurses during the VBPV by searching the electronic notes. Ratings of training and comments were summarized using feedback forms completed after training. Results: In total, 81,476 veterans participated in VBPVs over 2 years. Of those, 44,682 veterans had an existing ICD-10 code related to HTN. Of the 18,078 veterans who had a pre- and post-VBPV BP, the average change to systolic measurement was -10.6 mm Hg (range -82 to 78). Average change to diastolic measurement was -4.61 mm Hg (range -59 to 55). Most interventions addressed medication management (77%). Nurses' evaluations of the program were positive. Conclusions: Video visits provide reliable and convenient veteran-centered care. Such visits enable care when unanticipated interruptions occur such as the coronavirus disease 2019 pandemic. In addition to medication management, nurse-led interventions such as counseling on lifestyle changes can be effective in HTN management.

Model-based estimates of chikungunya epidemiological parameters and outbreak risk from varied data types.

Assessing the factors responsible for differences in outbreak severity for the same pathogen is a challenging task, since outbreak data are often incomplete and may vary in type across outbreaks (e.g., daily case counts, serology, cases per household). We propose that outbreaks described with varied data types can be directly compared by using those data to estimate a common set of epidemiological parameters. To demonstrate this for chikungunya virus (CHIKV), we developed a realistic model of CHIKV transmission, along with a Bayesian inference method that accommodates any type of outbreak data that can be simulated. The inference method makes use of the fact that all data types arise from the same transmission process, which is simulated by the model. We applied these tools to data from three real-world outbreaks of CHIKV in Italy, Cambodia, and Bangladesh to estimate nine model parameters. We found that these populations differed in several parameters, including pre-existing immunity and house-to-house differences in mosquito activity. These differences resulted in posterior predictions of local CHIKV transmission risk that varied nearly fourfold: 16% in Italy, 28% in Cambodia, and 62% in Bangladesh. Our inference method and model can be applied to improve understanding of the epidemiology of CHIKV and other pathogens for which outbreaks are described with varied data types.

Inactivation of Alicyclobacillus acidoterrestris spores, single or composite Escherichia coli and native microbiota in isotonic fruit-flavoured sports drinks processed by UV-C light.

Pasteurized sports drinks and other fruit-based beverages are susceptible to deterioration due to thermal processing ineffectiveness to inactivate certain spoilage microorganisms, like Alicyclobacillus acidoterrestris. This represents a major challenge for the beverage industry. The goals of this study were to: i) investigate the UV-C inactivation (annular thin film unit, actinometrical delivered fluence: 795-1270 mJ/cm2, 10-15 min, 20 °C, 1.8 L/h, Reh = 391-1067, recirculation mode operation) and the evolution during refrigerated storage of A. acidoterrestris ATCC 49025 spores and single or composite Escherichia coli ATCC 25922 in isotonic sports drinks (ISDs) made from orange (orange-ISD, UVT% = 81) or orange-banana-mango-kiwi-strawberry-lemon juices (multi-fruit-ISD, UVT% = 91), compared to a turbid orange-tangerine juice (OT juice, UVT% = 40); ii) assess the effect of pH, °Brix, A254nm, turbidity, colour and particle size of the ISDs and juice on microbial inactivation, iii) evaluate the evolution of native microbiota during cold storage, iv) investigate the Coroller, biphasic, Weibull, and Weibull-plus-tail models' ability to describe microbial inactivation and v) measure 5-hydroxymethylfurfural (HMF) formation. The modified biodosimetry method was used to calculate the germicidal UV-C fluences. Heat pasteurization (T-coil, 80 °C/6 min) was evaluated as the control treatment. UV-C was highly effective at inactivating E. coli as 4.1-5.1 and 4.5-5.6 log reductions were determined in the multi-fruit-ISD and orange-ISD, respectively, barely impacted by the background microbiota. No significant differences were recorded for the inactivation of E. coli in the UV-C and T-coil systems. Whereas, a significantly higher inactivation of A. acidoterrestris spores was achieved by UV-C (3.7-4.0 log reductions), compared to the negligible one achieved by the thermal treatment. Even though E. coli inactivation curves were similar in shape, UV-C was less effective when a cocktail of other E. coli strains was present. In comparison to the OT juice, the ISDs' inactivation kinetics were markedly different in shape, with a rapid decrease in population during the first minutes of treatment. The germicidal fluence (Hd biod) corresponding to A. acidoterrestris (19.1 mJ/cm2) was selected as it was higher than the one obtained for E. coli (11.0 mJ/cm2). UV-C induced 2.8- or 1.3 and 2.3- or 0.8 log-reductions of total aerobes or moulds and yeasts in the multi-fruit-ISD and orange-ISD, respectively. Compared to the other models, the Coroller and biphasic models showed a better fit and more accurate parameter estimates. UV-C-induced HMF production was not significant in the ISDs. The current study found that the UV-C treatment was more effective than typical heat pasteurization for inactivating A. acidoterrestris spores in isotonic drinks, following a similar trend for E. coli and native microbiota.

Disruption of the VAPB-PTPIP51 ER-mitochondria tethering proteins in post-mortem human amyotrophic lateral sclerosis.

Signaling between the endoplasmic reticulum (ER) and mitochondria regulates many neuronal functions that are perturbed in amyotrophic lateral sclerosis (ALS) and perturbation to ER-mitochondria signaling is seen in cell and transgenic models of ALS. However, there is currently little evidence that ER-mitochondria signaling is altered in human ALS. ER-mitochondria signaling is mediated by interactions between the integral ER protein VAPB and the outer mitochondrial membrane protein PTPIP51 which act to recruit and "tether" regions of ER to the mitochondrial surface. The VAPB-PTPI51 tethers are now known to regulate a number of ER-mitochondria signaling functions. These include delivery of Ca2+ from ER stores to mitochondria, mitochondrial ATP production, autophagy and synaptic activity. Here we investigate the VAPB-PTPIP51 tethers in post-mortem control and ALS spinal cords. We show that VAPB protein levels are reduced in ALS. Proximity ligation assays were then used to quantify the VAPB-PTPIP51 interaction in spinal cord motor neurons in control and ALS cases. These studies revealed that the VAPB-PTPIP51 tethers are disrupted in ALS. Thus, we identify a new pathogenic event in post-mortem ALS.

The PTPIP51 coiled-coil domain is important in VAPB binding, formation of ER-mitochondria contacts and IP3 receptor delivery of Ca2+ to mitochondria.

Signaling between the endoplasmic reticulum (ER) and mitochondria regulates a number of fundamental physiological processes. This signaling involves close physical contacts between the two organelles that are mediated by the VAPB-PTPIP51 ″tethering" proteins. The VAPB-PTPIP51 tethers facilitate inositol 1,4,5-trisphosphate (IP3) receptor delivery of Ca2+ from ER to mitochondria. Damage to the tethers is seen in Alzheimer's disease, Parkinson's disease and frontotemporal dementia with related amyotrophic lateral sclerosis (FTD/ALS). Understanding the mechanisms that regulate the VAPB-PTPIP51 interaction thus represents an important area of research. Recent studies suggest that an FFAT motif in PTPIP51 is key to its binding to VAPB but this work relies on in vitro studies with short peptides. Cellular studies to support this notion with full-length proteins are lacking. Here we address this issue. Immunoprecipitation assays from transfected cells revealed that deletion of the PTPIP51 FFAT motif has little effect on VAPB binding. However, mutation and deletion of a nearby coiled-coil domain markedly affect this binding. Using electron microscopy, we then show that deletion of the coiled-coil domain but not the FFAT motif abrogates the effect of PTPIP51 on ER-mitochondria contacts. Finally, we show that deletion of the coiled-coil domain but not the FFAT motif abrogates the effect of PTPIP51 on the IP3 receptor-mediated delivery of Ca2+ to mitochondria. Thus, the coiled-coil domain is essential for PTPIP51 ER-mitochondria signaling functions.

Targeting ER-Mitochondria Signaling as a Therapeutic Target for Frontotemporal Dementia and Related Amyotrophic Lateral Sclerosis.

Frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are two major neurodegenerative diseases. FTD is the second most common cause of dementia and ALS is the most common form of motor neuron disease. These diseases are now known to be linked. There are no cures or effective treatments for FTD or ALS and so new targets for therapeutic intervention are required but this is hampered by the large number of physiological processes that are damaged in FTD/ALS. Many of these damaged functions are now known to be regulated by signaling between the endoplasmic reticulum (ER) and mitochondria. This signaling is mediated by "tethering" proteins that serve to recruit ER to mitochondria. One tether strongly associated with FTD/ALS involves an interaction between the ER protein VAPB and the mitochondrial protein PTPIP51. Recent studies have shown that ER-mitochondria signaling is damaged in FTD/ALS and that this involves breaking of the VAPB-PTPIP51 tethers. Correcting disrupted tethering may therefore correct many other downstream damaged features of FTD/ALS. Here, we review progress on this topic with particular emphasis on targeting of the VAPB-PTPIP51 tethers as a new drug target.

Endoplasmic reticulum-mitochondria signaling in neurons and neurodegenerative diseases.

Recent advances have revealed common pathological changes in neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis with related frontotemporal dementia (ALS/FTD). Many of these changes can be linked to alterations in endoplasmic reticulum (ER)-mitochondria signaling, including dysregulation of Ca2+ signaling, autophagy, lipid metabolism, ATP production, axonal transport, ER stress responses and synaptic dysfunction. ER-mitochondria signaling involves specialized regions of ER, called mitochondria-associated membranes (MAMs). Owing to their role in neurodegenerative processes, MAMs have gained attention as they appear to be associated with all the major neurodegenerative diseases. Furthermore, their specific role within neuronal maintenance is being revealed as mutant genes linked to major neurodegenerative diseases have been associated with damage to these specialized contacts. Several studies have now demonstrated that these specialized contacts regulate neuronal health and synaptic transmission, and that MAMs are damaged in patients with neurodegenerative diseases. This Review will focus on the role of MAMs and ER-mitochondria signaling within neurons and how damage of the ER-mitochondria axis leads to a disruption of vital processes causing eventual neurodegeneration.

Disruption of ER-mitochondria tethering and signalling in C9orf72-associated amyotrophic lateral sclerosis and frontotemporal dementia.

Hexanucleotide repeat expansions in C9orf72 are the most common cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The mechanisms by which the expansions cause disease are not properly understood but a favoured route involves its translation into dipeptide repeat (DPR) polypeptides, some of which are neurotoxic. However, the precise targets for mutant C9orf72 and DPR toxicity are not fully clear, and damage to several neuronal functions has been described. Many of these functions are regulated by signalling between the endoplasmic reticulum (ER) and mitochondria. ER-mitochondria signalling requires close physical contacts between the two organelles that are mediated by the VAPB-PTPIP51 'tethering' proteins. Here, we show that ER-mitochondria signalling and the VAPB-PTPIP51 tethers are disrupted in neurons derived from induced pluripotent stem (iPS) cells from patients carrying ALS/FTD pathogenic C9orf72 expansions and in affected neurons in mutant C9orf72 transgenic mice. In these mice, disruption of the VAPB-PTPIP51 tethers occurs prior to disease onset suggesting that it contributes to the pathogenic process. We also show that neurotoxic DPRs disrupt the VAPB-PTPIP51 interaction and ER-mitochondria contacts and that this may involve activation of glycogen synthase kinases-3ß (GSK3ß), a known negative regulator of VAPB-PTPIP51 binding. Finally, we show that these DPRs disrupt delivery of Ca2+ from ER stores to mitochondria, which is a primary function of the VAPB-PTPIP51 tethers. This delivery regulates a number of key neuronal functions that are damaged in ALS/FTD including bioenergetics, autophagy and synaptic function. Our findings reveal a new molecular target for mutant C9orf72-mediated toxicity.

Active food additive based on encapsulated yerba mate (Ilex paraguariensis) extract: effect of drying methods on the oxidative stability of a real food matrix (mayonnaise).

The drying process used to obtain active food additives is critical to ensure its functionality. In this study, freeze- and spray-drying techniques were evaluated for encapsulation of extracts with antioxidant activity from yerba mate (Ilex paraguariensis), using maltodextrin (MD) as wall material. Additionally, the oxidative stability in a real food matrix (mayonnaise) was assessed. Both MD addition and drying methods affected the physical properties [moisture content, water activity (aW)] and oxidative stability. MD addition diminished moisture content and prevented polyphenol compounds from degradation. The spray-dried powders displayed the lowest moisture content (1.6 ± 0.3% bs), the highest polyphenol content (135.4 mg GAE/g pure extract), and oxidative stability than the freeze-dried samples. The antioxidant capacity of the encapsulated powder subjected to spray-drying increased the oxidative stability of the mayonnaise (258 ± 32 min) more than the other assayed system (165 ± 5 min). Therefore, a natural spray-dried antioxidant food additive was obtained with potential use in the food industry.

His452Tyr polymorphism in the human 5-HT2A receptor affects clozapine-induced signaling networks revealed by quantitative phosphoproteomics.

Antipsychotic drugs remain the current standard for schizophrenia treatment. Although they directly recognize the orthosteric binding site of numerous monoaminergic G protein-coupled receptors (GPCRs), these drugs, and particularly second-generation antipsychotics such as clozapine, all have in common a very high affinity for the serotonin 5-HT2A receptor (5-HT2AR). Using classical pharmacology and targeted signaling pathway assays, previous findings suggest that clozapine and other atypical antipsychotics behave principally as 5-HT2AR neutral antagonists and/or inverse agonists. However, more recent findings showed that antipsychotics may also behave as pathway-specific agonists. Reversible phosphorylation is a common element in multiple signaling networks. Combining a quantitative phosphoproteomic method with signaling network analysis, we tested the effect of clozapine treatment on the overall level of protein phosphorylation and signal transduction cascades in vitro in mammalian cell lines induced to express either the human 5-HT2AR or the H452Y variant of the gene encoding the 5-HT2AR receptor. This naturally occurring variation within the 5-HT2AR gene was selected because it has been repeatedly associated with schizophrenia patients who do not respond to clozapine treatment. Our data show that short time exposure (5 or 10 min) to clozapine (10-5 M) led to phosphorylation of numerous signaling components of pathways involved in processes such as endocytosis, ErbB signaling, insulin signaling or estrogen signaling. Cells induced to express the H452Y variant showed a different basal phosphoproteome, with increases in the phosphorylation of mTOR signaling components as a translationally relevant example. However, the effect of clozapine on the functional landscape of the phosphoproteome was significantly reduced in cells expressing the 5-HT2AR-H452Y construct. Together, these findings suggest that clozapine behaves as an agonist inducing phosphorylation of numerous pathways downstream of the 5-HT2AR, and that the single nucleotide polymorphism encoding 5-HT2AR-H452Y affects these clozapine-induced phosphorylation-dependent signaling networks.

Identification of PARP-1 in cancer stem cells of gastrointestinal cancers: A preliminary study.

Advanced-stage gastrointestinal tumors have high mortality due to chemotherapy limitations. One of the causes of treatment failure is the presence of cancer stem cells (CSCs), which show resistance mechanisms against DNA damage, such as poly (adenosine diphosphate-ribose) polymerase 1 (PARP-1). However, little is known about the relevance of PARP-1 in these tumor cells. Our purpose is to analyze the expression of PARP-1 in cancer cells and CSCs from gastrointestinal tumors, its relationship with the DNA damage repair process and its modulation by cytotoxic and PARP-1 inhibitors. We used pancreatic, liver and colon cancer cell lines and isolated CSCs using Aldefluor technology to analyze PARP-1 expression. In addition, we examined the effect of classic cytotoxic drugs (Doxorubicin, Gemcitabine, Irinotecan and 5-Fluorouracil) and a PARP-1 inhibitor (Olaparib) in cultured cells and 3D tumorspheres. We demonstrated that PARP-1 is highly expressed in pancreatic, liver and colon tumor cells and that this expression was significantly higher in cell populations with CSC characteristics. In addition, Doxorubicin and Gemcitabine increased their cytotoxic effect when administered simultaneously with Olaparib, decreasing the formation of 3D tumorspheres. Our findings suggest that PARP-1 is a common and relevant resistance mechanism in CSCs from gastrointestinal tumors and that the use of PARP-1 inhibitors may be an adjuvant therapy to increase apoptosis in this type of cells which are responsible to cancer recurrence and metastasis.

PARP-1 activation after oxidative insult promotes energy stress-dependent phosphorylation of YAP1 and reduces cell viability.

Poly(ADP-ribose) polymerase 1 (PARP-1) is a nuclear enzyme that catalyze the transfer of ADP-ribose units from NAD+ to several target proteins involved in cellular stress responses. Using WRL68 (HeLa derivate) cells, we previously showed that PARP-1 activation induced by oxidative stress after H2O2 treatment lead to depletion of cellular NAD+ and ATP, which promoted cell death. In this work, LC-MS/MS-based phosphoproteomics in WRL68 cells showed that the oxidative damage induced by H2O2 increased the phosphorylation of YAP1, a transcriptional co-activator involved in cell survival, and modified the phosphorylation of other proteins involved in transcription. Genetic or pharmacological inhibition of PARP-1 in H2O2-treated cells reduced YAP1 phosphorylation and degradation and increased cell viability. YAP1 silencing abrogated the protective effect of PARP-1 inhibition, indicating that YAP1 is important for the survival of WRL68 cells exposed to oxidative damage. Supplementation of NAD+ also reduced YAP1 phosphorylation, suggesting that the loss of cellular NAD+ caused by PARP-1 activation after oxidative treatment is responsible for the phosphorylation of YAP1. Finally, PARP-1 silencing after oxidative treatment diminished the activation of the metabolic sensor AMPK. Since NAD+ supplementation reduced the phosphorylation of some AMPK substrates, we hypothesized that the loss of cellular NAD+ after PARP-1 activation may induce an energy stress that activates AMPK. In summary, we showed a new crucial role of PARP-1 in the response to oxidative stress in which PARP-1 activation reduced cell viability by promoting the phosphorylation and degradation of YAP1 through a mechanism that involves the depletion of NAD+.

Torin 2 Derivative, NCATS-SM3710, Has Potent Multistage Antimalarial Activity through Inhibition of P. falciparum Phosphatidylinositol 4-Kinase (Pf PI4KIIIß).

Drug resistance is a constant threat to malaria control efforts making it important to maintain a good pipeline of new drug candidates. Of particular need are compounds that also block transmission by targeting sexual stage parasites. Mature sexual stages are relatively resistant to all currently used antimalarials except the 8-aminoquinolines that are not commonly used due to potential side effects. Here, we synthesized a new Torin 2 derivative, NCATS-SM3710 with increased aqueous solubility and specificity for Plasmodium and demonstrate potent in vivo activity against all P. berghei life cycle stages. NCATS-SM3710 also has low nanomolar EC50s against in vitro cultured asexual P. falciparum parasites (0.38 ± 0.04 nM) and late stage gametocytes (5.77 ± 1 nM). Two independent NCATS-SM3710/Torin 2 resistant P. falciparum parasite lines produced by growth in sublethal Torin 2 concentrations both had genetic changes in PF3D7_0509800, annotated as a phosphatidylinositol 4 kinase (Pf PI4KIIIß). One line had a point mutation in the putative active site (V1357G), and the other line had a duplication of a locus containing Pf PI4KIIIß. Both lines were also resistant to other Pf PI4K inhibitors. In addition NCATS-SM3710 inhibited purified Pf PI4KIIIß with an IC50 of 2.0 ± 0.30 nM. Together the results demonstrate that Pf PI4KIIIß is the target of Torin 2 and NCATS-SM3710 and provide new options for potent multistage drug development.

Hurdle processing of turbid fruit juices involving encapsulated citral and vanillin addition and UV-C treatment.

The aim of this study was to evaluate a hurdle strategy for orange-tangerine (OT) and orange-banana-mango-kiwi-strawberry (OBMKS) juices processing based on UV-C treatment assisted or not by mild heat and the addition of natural antimicrobials. Vanillin and citral emulsions were successfully encapsulated using maltodextrin and HI-CAP (5,18,3) and characterized. The susceptibility of Lactobacillus plantarum ATCC 8014, Escherichia coli ATCC 25922, and Saccharomyces cerevisiae KE 162 to binary mixtures of the encapsulated agents was examined in culture media according to the Berenbaum experimental design. The boundary between growth and non-growth as a function of vanillin and citral concentrations was predicted by means of the probabilistic model using logistic regression. Microbial inactivation achieved by pilot-scale UV-C light (0-390 mJ/cm2) on its own, assisted by mild heat (50 °C, UV-C/H) and combined with antimicrobials (1000 ppm vanillin plus 100 ppm citral) addition (UV-C + A/UV-C/H + A) was assessed in OT and OBMKS. Yeast induced damage in a model solution treated by UV-C + A was studied by flow cytometry (FC). All the antimicrobial mixtures resulted in additive effects (FICindex = 1), thus offering through the probabilistic models a range of formulation possibilities with antimicrobial capacity encompassing lower vanillin and citral concentrations compared to those required when used alone (Vrange = 0-1875 ppm plus Crange = 392-0 ppm). UV-C led up to 3.7-3.8, 2.4-3.6 and 1.5-1.6 log-reductions of E. coli, L. plantarum and S. cerevisiae in OT and OBMKS, respectively. A significant increase of 1.7-2.2, 2.1-2.7 and 4.1-5.3 log cycles in microbial inactivation was observed after UV-C/H treatment. Additional inactivation of 0.7-3.1 and 0.5-2.7 log reductions were observed for E. coli and S. cerevisiae, respectively, when UV-C + A and UV-C/H + A were applied in both juices. Therefore, the addition of antimicrobials to the UV-C treated juices, showed additive to synergistic effects on E. coli and S. cerevisiae, respectively along refrigerated storage. A shift from yeast cells with intact membrane and esterase activity in control samples to cells with permeabilized membrane in C + A, UV-C and UV-C + A samples were determined by FC. The shift was more noticeable in UV-C + A samples. Sublethally damaged cells were only detected in C + A and UV-C samples. This study demonstrates that combining a pilot-scale UV-C treatment with the addition of chosen binary mixtures of vanillin and citral, can ensure more than 5 log-reductions of E. coli, L. plantarum and S. cerevisiae in OT and OBKMS juice blends.

Effect of pilot-scale UV-C light treatment assisted by mild heat on E. coli, L. plantarum and S. cerevisiae inactivation in clear and turbid fruit juices. Storage study of surviving populations.

Consumer growing demands for high-quality and safe food and beverages have stimulated the interest in alternative preservation technologies. Short-wavelength ultraviolet light (UV-C, 254 nm) has proven to be useful for the decontamination of a great variety of clear juices while improving their quality compared to traditional thermal treatments. Suspended solids and coloured compounds in turbid juices, diminish light transmission. The use of UV-C under a hurdle approach, may be a promising strategy for their treatment. The purpose of this study was to analyse Escherichia coli ATCC 25922, Saccharomyces cerevisiae KE 162 and Lactobacillus plantarum ATCC 8014 inactivation in clear pear juice (PJ), turbid orange-tangerine (OT) and orange-banana-mango-kiwi-strawberry (OBMKS) juices processed by single UV-C (390 mJ/cm2, 20 °C) and UV-C assisted by mild heat (UV-C/H, 50 °C) at pilot-scale in a coiled tubing unit and stored under refrigeration (5 °C). Inactivation studies were also conducted in peptone water (PW) and model solution (MS). The adequacy of the Coroller, Weibull and Biphasic Plus Shoulder models was studied. UV-C was highly effective in PW, MS and PJ, achieving up to 5.5-6.3-4.7, 4.8-5.1-4.6 and 4.4-5.5 log reductions for L. plantarum, E. coli,and S. cerevisiae, respectively. Whereas, a moderate inactivation by single UV-C was recorded in the turbid blends, reducing up to 2.4-3.8-1.6 and 3.6-3.7-1.3 log-cycles in OT and OBMKS, respectively. When the UV-C/H treatment was applied, high bacterial inactivation was observed achieving 5.2-5.6, 6.3-6.6 and 5.5-6.7 log reductions in OT, OBMKS and PJ, respectively, while 4.6-4.9 log reductions were determined for the yeast in OBMKS and OT, respectively. Thus, additive inactivation effects between UV-C and H were observed. All the models tested gave useful information regarding the existence of microbial subpopulations with varying resistances. However, the cumulative Weibull distribution function was the most versatile one, fitting inactivation curves with different shapes. Additionally, the frequency distributions of resistances showed that UV-C/H not only increased the UV-C microbicidal effect but changed the distribution of inactivation times. Principal component analysis revealed that UV-C effectiveness was associated to low particle size, a⃰, turbidity and high UV-C transmittance. An increase on the inactivation of treated bacterial populations was recorded along storage, while no yeast recovery was observed, thus emphasizing the contribution of refrigerated storage to microbial inactivation. Microbial inactivation in clear and turbid juices achieved by UV-C (390 mJ/cm2) assisted by mild heat (50 °C) and subsequent refrigerated storage may represent an useful alternative for multiple applications in the juice industry.

Exploración del contexto y las necesidades de educación en salud en un grupo indígena uitoto en Bogotá / Survey on the context and healthcare education needs of a uitoto indigenous group in Bogota

Introducción: Colombia es un país multiétnico y multilingüe con seis millones (14%) de habitantes que pertenecen a grupos étnicos minoritarios, por lo que la situación actual de las comunidades indígenas ha sido precaria por las limitantes de acceso a los planes de salud. Objetivo: realizar una exploración de la comunidad uitoto ubicada en la ciudad de Bogotá D.C., Colombia, del contexto y las necesidades de educación frente al cáncer del cuello uterino y la salud sexual y reproductiva. Materiales y métodos: estudio cualitativo de enfoque etnográfico (prueba piloto) en la comunidad uitoto con una observación participante, registrado en un diario de campo. Resultados: las personas de la comunidad uitoto manifiestan que muchos de sus integrantes no cuentan con un servicio de salud contributivo, sino subsidiado por condiciones de desplazamiento. Discusión: se reconoce la existencia latente de las barreras de salud en comunidades indígenas en estado de vulnerabilidad en las áreas urbanas. Conclusión: se destaca la importancia de integrar en la medicina occidental y tradicional en los procesos de salud para esta comunidad.

Introduction: Colombia is a multiethnic and multilingual country with six million (14%) of inhabitants belonging to minority ethnic groups. These indigenous communities are currently living in a precarious situation due to their limited access to healthcare plans. Objective: to conduct a survey on a uitoto community living in the city of Bogotá D.C., Colombia, predominantly on their context and their education needs regarding uterine cervix cancer and sexual and reproductive health. Materials and Methods: a qualitative study with a focus on ethnography (pilot study) on a uitoto community, using participant observation recorded in a fieldwork diary. Results: uitoto community members interviewed manifest that many of them do not count with a contributive healthcare service. They only count with a subsidiary service as a result from being displaced people. Discussion: better awareness of the latent existence of barriers to healthcare for vulnerable indigenous communities living in urban areas was achieved. Conclusion: we stress the importance of integrating traditional medicine into Western medicine healthcare processes for this community.

Promoción del lavado de manos en la población con manos en la población con discapacidad auditiva / Promoting handwashing among people with hearing impairment

Introducción: las campañas de promoción y prevención del lavado de manos son estrategias económicas y de alto impacto según la OMS/OPS. Las personas con discapacidad auditiva presentan dificultades para acceder a estas. Objetivo: realizar una reflexión sobre la promoción del lavado de manos en población con discapacidad auditiva mediante la virtualidad. Desarrollo: la experiencia con la aplicación de un OVA en estudiantes con discapacidad auditiva permite comprender que diseñar y aplicar herramientas a partir de las TICs para transmitir educación en salud a la población con discapacidad auditiva, requiere recursos tecnológicos, humanos y financieros que pueden ser financiados por entes gubernamentales y asociaciones que apoyen esta población. Conclusiones: los profesionales del área de la salud deben diseñar programas de promoción y prevención virtual para comunidades con discapacidades o diferentes vulnerabilidades para generar un aporte desde la salud.

ntroduction: According to the WHO/PHO handwashing promotion campaigns are high-impact economic strategies. People with hearing disability have difficulties for accessing these campaigns. Objective: to conduct a discussion exercise on promoting handwashing among people with hearing disability by means of a virtual campaign. Development: the experience applying VLA (virtual learning objects) in students with hearing disability allows understanding that designing and applying information and communication technology (ICT) tools to transmit heath education to people with hearing challenges, requires technological, human and financial resources which may be provided by the state and organizations supporting this population. Conclusions: healthcare professionals must design virtual promotion and prevention programs targeted at communities with disabilities and various vulnerabilities, as a contribution from the health sector.