Levels of Small Extracellular Vesicles Containing hERG-1 and Hsp47 as Potential Biomarkers for Cardiovascular Diseases.

The diagnosis of cardiovascular disease (CVD) is still limited. Therefore, this study demonstrates the presence of human ether-a-go-go-related gene 1 (hERG1) and heat shock protein 47 (Hsp47) on the surface of small extracellular vesicles (sEVs) in human peripheral blood and their association with CVD. In this research, 20 individuals with heart failure and 26 participants subjected to cardiac stress tests were enrolled. The associations between hERG1 and/or Hsp47 in sEVs and CVD were established using Western blot, flow cytometry, electron microscopy, ELISA, and nanoparticle tracking analysis. The results show that hERG1 and Hsp47 were present in sEV membranes, extravesicularly exposing the sequences 430AFLLKETEEGPPATE445 for hERG1 and 169ALQSINEWAAQTT- DGKLPEVTKDVERTD196 for Hsp47. In addition, upon exposure to hypoxia, rat primary cardiomyocytes released sEVs into the media, and human cardiomyocytes in culture also released sEVs containing hERG1 (EV-hERG1) and/or Hsp47 (EV-Hsp47). Moreover, the levels of sEVs increased in the blood when cardiac ischemia was induced during the stress test, as well as the concentrations of EV-hERG1 and EV-Hsp47. Additionally, the plasma levels of EV-hERG1 and EV-Hsp47 decreased in patients with decompensated heart failure (DHF). Our data provide the first evidence that hERG1 and Hsp47 are present in the membranes of sEVs derived from the human cardiomyocyte cell line, and also in those isolated from human peripheral blood. Total sEVs, EV-hERG1, and EV-Hsp47 may be explored as biomarkers for heart diseases such as heart failure and cardiac ischemia.

NCoA3 upregulation in breast cancer­associated adipocytes elicits an inflammatory profile.

Nuclear receptor coactivator 3 (NCoA3) is a transcriptional coactivator of NF­κB and other factors, which is expressed at relatively low levels in normal cells and is amplified or overexpressed in several types of cancer, including breast tumors. NCoA3 levels have been shown to be decreased during adipogenesis; however, its role in tumor­surrounding adipose tissue (AT) remains unknown. Therefore, the present study assessed the modulation of NCoA3 in breast cancer­associated adipocytes and evaluated its association with the expression of inflammatory markers. 3T3­L1 adipocytes were stimulated with conditioned medium from human breast cancer cell lines and the expression levels of NCoA3 were evaluated by reverse transcription­quantitative (q)PCR. NF­κB activation was measured by immunofluorescence, and tumor necrosis factor and monocyte chemoattractant protein 1 levels were analyzed by qPCR and dot blot assays. The results obtained from the in vitro model were supported using mammary AT (MAT) from female mice, MAT adjacent to tumors from patients with breast cancer and bioinformatics analysis. The results revealed that adipocytes expressing high levels of NCoA3 were mainly associated with a pro­inflammatory profile. In 3T3­L1 adipocytes, NCoA3 downregulation or NF­κB inhibition reversed the expression of inflammatory molecules. In addition, MAT from patients with a worse prognosis exhibited high levels of this coactivator. Notably, adipocyte NCoA3 levels could be modulated by inflammatory signals from tumors. The modulation of NCoA3 levels in synergy with NF­κB activity in MAT in a tumor context could be factors required to establish breast cancer­associated inflammation. As adipocytes are involved in the development and progression of breast cancer, this signaling network deserves to be further investigated to improve future tumor treatments.

A Short-Term Sucrose Diet Impacts Cell Proliferation of Neural Precursors in the Adult Hypothalamus.

Radial glia-like cells in the hypothalamus and dorsal vagal complex are neural precursors (NPs) located near subventricular organs: median eminence and area postrema, respectively. Their strategic position can detect blood-borne nutrients, hormones, and mitogenic signals. Hypothalamic NPs increase their proliferation with a mechanism that involves hemichannel (HC) activity. NPs can originate new neurons in response to a short-term high-fat diet as a compensatory mechanism. The effects of high carbohydrate Western diets on adult neurogenesis are unknown. Although sugars are usually consumed as sucrose, more free fructose is now incorporated into food items. Here, we studied the proliferation of both types of NPs in Sprague Dawley rats exposed to a short-term high sucrose diet (HSD) and a control diet. In tanycyte cultures, we evaluated the effects of glucose and fructose and a mix of both hexoses on HC activity. In rats fed an HSD, we observed an increase in the proliferative state of both precursors. Glucose, either in the presence or absence of fructose, but not fructose alone, induced in vitro HC activity. These results should broaden the understanding of the nutrient monitoring capacity of NPs in reacting to changes in feeding behavior, specifically to high sugar western diets.

Medicina paliativa hospitalaria: modelo de atención implementado en un hospital universitario / Hospital palliative medicine. Revision of a care model implemented at a university hospital

BACKGROUND: Palliative Medicine (PM) is a specialty whose objective is to prevent and alleviate suffering associated with advanced diseases. Hospital palliative medicine has benefits in symptom control, quality of life and cost containment. Hospital PM support teams that serve as referral specialists are in charge of a PM care model. AIM: To describe the clinical experience of a PM support team in a tertiary hospital in Chile. MATERIAL AND METHODS: Review of clinical records of patients referred to a hospital PM support team between March 2015 and July 2018. Administrative data of referrals, sociodemographic and clinical characteristics of patients, their investigated problems and the interventions proposed by the PM team were described. RESULTS: During the study period, 790 referrals were registered, most of them from the internal medicine department (31%) or critical care (24%). During the study period, the number of annual referrals increased from 177 to 237 and the time lapse after hospital admission decreased from five to three days. The mean age of patients was 65.8 years and their main diagnosis was an oncological disease in 81%. The most frequently identified symptoms were fatigue in 71% of patients, depression in 68% and pain in 60%. The main interventions proposed by the PM team were communication support in 64% of patients, analgesia in 62% and education for family caregivers in 49%. Conclusions: The hospital PM team proposes a care model that allows the evaluation and a therapeutic approach for patients suffering from advanced diseases, using a multidimensional perspective including their families.

Decision making process in multiple sclerosis: An Argentine pilot study.

BACKGROUND: There is scarce information regarding the decision-making process (DMP) in people with MS (PwMS) from Latin America. OBJECTIVE: To evaluate the DMP in Argentinean PwMS and to assess its relationship with patient preferences, and clinical-demographic characteristics. METHODS: PwMS from the patient organization Esclerosis Múltiple Argentina (n = 1275) were invited to participate in a self-administered web-based survey. Participants were asked to provide clinical-demographic information and to complete a questionnaire on their perceptions about the information provided by their physician, the Control Preference Scale, and the Satisfaction with the Decisions and Care Questionnaire, and were inquired on their preferred sources of information about MS. RESULTS: The survey was completed by 379 PwMS. Most were females (67%); mean age: 40.3 (SD = 11.1) years; mean disease duration: 7.9 (SD = 7.2) years. Patients' decisional control preferred role was active in 47%, shared in 27%, and passive in 26%. A moderate concordance (weighted kappa 0.55) was observed between patients' preferences and self-reported DMP. Seventy-two percent participated in the DMP according to their preferences (concordance rates: active 66%, shared 87%, passive 51%). Most (83%) declared receiving information from their neurologists, matching their preferences (94%). CONCLUSIONS: Argentinian PwMS have distinctive preferences regarding information management and decision making.

The Effect of Dynasore Upon the Negative Interaction Between ENaC and CFTR Channels in Xenopus laevis Oocytes.

Shroom is a family of related proteins linked to the actin cytoskeleton, and one of them, xShroom1, is constitutively expressed in Xenopus laevis oocytes which is required for the expression of the epithelial sodium channel (ENaC). On the other hand, ENaC and the cystic fibrosis transmembrane regulator (CFTR) are co-expressed in many types of cells with a negative or positive interaction depending on the studied tissues. Here, we measured the amiloride-sensitive ENaC currents (INaamil) and CFTR currents (ICFTR) with voltage clamp techniques in oocytes co-injected with ENaC and/or CFTR and xShroom1 antisense oligonucleotides. The objective was to study the mechanism of regulation of ENaC by CFTR when xShroom1 was suppressed and the endocytic traffic of CFTR was blocked. CFTR activation had a measurable negative effect on ENaC and this activation resulted in a greater inhibition of INaamil than with xShroom1 antisense alone. Our results with Dynasore, a drug that acts as an inhibitor of endocytic pathways, suggest that the changes in INaamil by xShroom1 downregulation were probably due to an increment in channel endocytosis. An opposite effect was observed when ICFTR was measured. Thus, when xShroom1 was downregulated, the ICFTR was larger than in the control experiments and this effect is not observed with Dynasore. A speculative explanation could be that xShroom1 exerts a dual effect on the endocytic traffic of ENaC and CFTR and these actions were canceled with Dynasore. In the presence of Dynasore, no difference in either INaamil or ICFTR was observed when xShroom1 was downregulated.

An Intensive Care Unit Team Reflects on End-of-Life Experiences With Patients and Families in Chile.

BACKGROUND: Deaths in the intensive care unit (ICU) represent an experience of suffering for patients, their families, and professionals. End-of-life (EOL) care has been added to the responsibilities of the ICU team, but the evidence supporting EOL care is scarce, and there are many barriers to implementing the clinical recommendations that do exist. OBJECTIVES: To explore the experiences and perspectives of the various members of an ICU care team in Chile regarding the EOL care of their patients. METHODS: A qualitative study was performed in the ICU of a high-complexity academic urban hospital. The study used purposive sampling with focus groups as a data collection method. A narrative analysis based on grounded theory was done. RESULTS: Four discipline-specific focus groups were conducted; participants included 8 nurses, 6 nursing assistants, 8 junior physicians, and 6 senior physicians. The main themes that emerged in the analysis were emotional impact and barriers to carrying out EOL care. The main barriers identified were cultural difficulties related to decision-making, lack of interprofessional clinical practice, and lack of effective communication. Communication difficulties within the team were described along with lack of self-efficacy for family-centered communication. CONCLUSION: These qualitative findings expose gaps in care that must be filled to achieve high-quality EOL care in the ICU. Significant emotional impact, barriers related to EOL decision-making, limited interprofessional clinical practice, and communication difficulties were the main findings cross-referenced.

[Hospital palliative medicine. Revision of a care model implemented at a university hospital]. / Medicina paliativa hospitalaria: modelo de atención implementado en un hospital universitario.

BACKGROUND: Palliative Medicine (PM) is a specialty whose objective is to prevent and alleviate suffering associated with advanced diseases. Hospital palliative medicine has benefits in symptom control, quality of life and cost containment. Hospital PM support teams that serve as referral specialists are in charge of a PM care model. AIM: To describe the clinical experience of a PM support team in a tertiary hospital in Chile. MATERIAL AND METHODS: Review of clinical records of patients referred to a hospital PM support team between March 2015 and July 2018. Administrative data of referrals, sociodemographic and clinical characteristics of patients, their investigated problems and the interventions proposed by the PM team were described. RESULTS: During the study period, 790 referrals were registered, most of them from the internal medicine department (31%) or critical care (24%). During the study period, the number of annual referrals increased from 177 to 237 and the time lapse after hospital admission decreased from five to three days. The mean age of patients was 65.8 years and their main diagnosis was an oncological disease in 81%. The most frequently identified symptoms were fatigue in 71% of patients, depression in 68% and pain in 60%. The main interventions proposed by the PM team were communication support in 64% of patients, analgesia in 62% and education for family caregivers in 49%. CONCLUSIONS: The hospital PM team proposes a care model that allows the evaluation and a therapeutic approach for patients suffering from advanced diseases, using a multidimensional perspective including their families.

Implementation of a Palliative Hospital-Centered Spiritual and Psychological Telehealth System During COVID-19 Pandemic.

BACKGROUND: The severity of the COVID-19 pandemic has resulted in limited provision of palliative care and hospital teams have had to rise to the challenge of how to deliver care safely to people with palliative needs. Telehealth interventions have been seen as a useful resource with potential to improve clinical effectiveness. OBJECTIVE: To describe the implementation of a spiritual and psychological palliative telehealth system during the pandemic. METHODS: Pilot study based on the implementation of a telehealth system designed to support hospitalized patients referred to a mobile palliative care team, through synchronic videoconferences, and including patients' relatives. The implementation included protocol development, physical infrastructure, and training. The intervention consisted of spiritual and psychological telehealth sessions performed remotely by the chaplain and psychologist of a palliative care team. RESULTS: During the study period 59 patients were recruited, median age of 70 years, 57.6% females. The primary diagnosis was severe COVID-19 (50.8%), advanced cancer (32.2%) and advanced chronic illness (16.9%). A total of 211 telehealth sessions were carried out, 82% psychological and 18% spiritual. The main criteria for psychological sessions were being related to seriously ill patients with withdrawal or withholding of life-support treatment (60.1%). The main criteria for spiritual sessions were being a patient with spiritual suffering or requesting spiritual assistance (73.6%). An electronic user satisfaction survey indicated high satisfaction rates. CONCLUSION: This report demonstrates that it is possible to provide spiritual and psychological palliative care to hospitalized patients and families during pandemic restrictions through interdisciplinary telehealth delivery.

Association between Spirituality, Religiosity, Spiritual Pain, Symptom Distress, and Quality of Life among Latin American Patients with Advanced Cancer: A Multicenter Study.

Objectives: The purpose of this multicenter study was to characterize the association between spirituality, religiosity, spiritual pain, symptom distress, coping, and quality of life (QOL) among Latin American advanced cancer patients. Methods: Three hundred twenty-five advanced cancer patients from palliative care clinics in Chile, Guatemala, and the United States completed validated assessments: Faith, Importance and Influence, Community, and Address (FICA) (spirituality/religiosity), Edmonton Symptom Assessment Scale-Financial/Spiritual (ESAS-FS), including spiritual pain, Penn State Worry Questionnaire-Abbreviated (PSWQ-A), Center for Epidemiologic Studies Depression Scale (CES-D), Brief-coping strategies (COPE) and Brief religious coping (RCOPE) and RCOPE, respectively, and Functional Assessment of Chronic Illness Therapy-Spiritual Well-Being, Expanded version (FACIT-Sp-Ex). Results: Median age: 58 years (range: 19-85); 60% female; and 62% Catholic and 30% Christian, but not Catholic. Three hundred fifteen patients (97%) considered themselves spiritual and 89% religious, with median intensities of 7 (interquartile range [IQR]: 5-10) and 7 (5-9), respectively (0-10 scale, 10 = "very much"). Median importance of spirituality/religiosity was 10 (IQR: 8-10). The frequency and associations between spirituality/religiosity and various items were as follows: helps to cope with illness (98%; r = 0.66303; p < 0.0001), positive effect on physical symptoms (81%; r = 0.42067; p < 0.0001), and emotional symptoms (84%; r = 0.16577; p < 0.0001). One hundred ninety-five patients (60%) reported that their spiritual/religious needs had not been supported by the medical team. Spiritual pain was reported in 162/311 patients (52%), with median intensity of 6 (IQR: 5-8). Spiritual pain was associated with pain (p = 0.0225), depression (p < 0.0001), anxiety (p < 0.0001), worry (p < 0.001), behavioral disengagement (p = 0.0148), FACIT-Sp-Ex score (p = 0.0002), and negative RCOPE (p < 0.0001). Significance of Results: Spirituality and religiosity are frequent, intense, and rarely addressed among Latin American patients. Spirituality/religiosity was associated with positive COPE and higher QOL. Spiritual pain was also frequent and associated with physical and psychosocial distress. These patients need increased spiritual/religious support.

Functional relationship between CFTR and RAC3 expression for maintaining cancer cell stemness in human colorectal cancer.

PURPOSE: CFTR mutations not only cause cystic fibrosis, but also increase the risk of colorectal cancer. A putative role of CFTR in colorectal cancer patients without cystic fibrosis has so far, however, not been investigated. RAC3 is a nuclear receptor coactivator that has been found to be overexpressed in several human tumors, and to be required for maintaining cancer stemness. Here, we investigated the functional relationship between CFTR and RAC3 for maintaining cancer stemness in human colorectal cancer. METHODS: Cancer stemness was investigated by analysing the expression of stem cell markers, clonogenic growth and selective retention of fluorochrome, using stable transfection of shCFTR or shRAC3 in HCT116 colorectal cancer cells. In addition, we performed pathway enrichment and network analyses in both primary human colorectal cancer samples (TCGA, Xena platform) and Caco-2 colorectal cancer cells including (1) CD133+ or CD133- side populations and (2) CFTRwt or CFTRmut cells (ConsensusPathDB, STRING, Cytoscape, GeneMANIA). RESULTS: We found that the CD133+ side population expresses higher levels of RAC3 and CFTR than the CD133- side population. RAC3 overexpression increased CFTR expression, whereas CFTR downregulation inhibited the cancer stem phenotype. CFTR mRNA levels were found to be increased in colorectal cancer samples from patients without cystic fibrosis compared to those with CFTR mutations, and this correlated with an increased expression of RAC3. The expression pattern of a gene set involved in inflammatory response and nuclear receptor modulation in CD133+ Caco-2 cells was found to be shared with that in CFTRwt Caco-2 cells. These genes may contribute to colorectal cancer development. CONCLUSIONS: CFTR may play a non-tumor suppressor role in colorectal cancer development and maintenance involving enhancement of the expression of a set of genes related to cancer stemness and development in patients without CFTR mutations.

The FGF2-induced tanycyte proliferation involves a connexin 43 hemichannel/purinergic-dependent pathway.

In the adult hypothalamus, the neuronal precursor role is attributed to the radial glia-like cells that line the third-ventricle (3V) wall called tanycytes. Under nutritional cues, including hypercaloric diets, tanycytes proliferate and differentiate into mature neurons that moderate body weight, suggesting that hypothalamic neurogenesis is an adaptive mechanism in response to metabolic changes. Previous studies have shown that the tanycyte glucosensing mechanism depends on connexin-43 hemichannels (Cx43 HCs), purine release, and increased intracellular free calcium ion concentration [(Ca2+ )i ] mediated by purinergic P2Y receptors. Since, Fibroblast Growth Factor 2 (FGF2) causes similar purinergic events in other cell types, we hypothesize that this pathway can be also activated by FGF2 in tanycytes to promote their proliferation. Here, we used bromodeoxyuridine (BrdU) incorporation to evaluate if FGF2-induced tanycyte cell division is sensitive to Cx43 HC inhibition in vitro and in vivo. Immunocytochemical analyses showed that cultured tanycytes maintain the expression of in situ markers. After FGF2 exposure, tanycytic Cx43 HCs opened, enabling release of ATP to the extracellular milieu. Moreover, application of external ATP was enough to induce their cell division, which could be suppressed by Cx43 HC or P2Y1-receptor inhibitors. Similarly, in vivo experiments performed on rats by continuous infusion of FGF2 and a Cx43 HC inhibitor into the 3V, demonstrated that FGF2-induced ß-tanycyte proliferation is sensitive to Cx43 HC blockade. Thus, FGF2 induced Cx43 HC opening, triggered purinergic signaling, and increased ß-tanycytes proliferation, highlighting some of the molecular mechanisms involved in the cell division response of tanycyte. This article has an Editorial Highlight see https://doi.org/10.1111/jnc.15218.

Extract from Aloysia polystachya Induces the Cell Death of Colorectal Cancer Stem Cells.

Cancer stem cells (CSCs) are an important player in the resistance of cancers to therapy. In this work, we determined the flavonoids composition and biological action of Aloysia polystachya (AP) extracts in colorectal cancer. The chemical characterization of extracts was performed by HPLC. Assays of cytotoxicity, apoptosis, migration and invasion, metalloproteases activity, clonogenic growth, tumorspheres formation, Hoechts efflux, pluripotency marker expression and sensitization to chemotherapeutic drugs were performed in vitro in human HCT116 and murine CT26 colorectal cancer cells. The AP toxicity and effect in tumor growth administered alone or in combination with 5- Fluorouracile was analyzed in vivo, including histopathological studies. We found that AP extracts induced in vitro the apoptosis of colorectal cancer cell lines decreasing the CSC proportion. Moreover, they were capable to kill 5-Fluorouracile resistant side population cells. At not toxic doses in vivo, AP extracts inhibited tumor growth. Regarding the ability to reduce the CSC population, AP extracts deserves to be investigated as a useful therapy for colorectal cancer treatment.

Connexin-43 Gap Junctions Are Responsible for the Hypothalamic Tanycyte-Coupled Network.

Tanycytes are hypothalamic radial glia-like cells that form the basal wall of the third ventricle (3V) where they sense glucose and modulate neighboring neuronal activity to control feeding behavior. This role requires the coupling of hypothalamic cells since transient decreased hypothalamic Cx43 expression inhibits the increase of brain glucose-induced insulin secretion. Tanycytes have been postulated as possible hypothalamic neuronal precursors due to their privileged position in the hypothalamus that allows them to detect mitogenic signals and because they share the markers and characteristics of neuronal precursors located in other neurogenic niches, including the formation of coupled networks through connexins. Using wild-type (WT), Cx30-/- and Cx30-/-, Cx43fl/fl:glial fibrillary acidic protein (GFAP)-Cre (double knockout, dKO) mouse lines, we demonstrated that tanycytes are highly coupled to each other and also give rise to a panglial network specifically through Cx43. Using the human GFAP (hGFAP)-enhanced green fluorescent protein (EGFP) transgenic mouse line, we provided evidence that the main parenchymal-coupled cells were astrocytes. In addition, electrophysiological parameters, such as membrane resistance, were altered when Cx43 was genetically absent or pharmacologically inhibited. Finally, in the dKO mouse line, we detected a significant decrease in the number of hypothalamic proliferative parenchymal cells. Our results demonstrate the importance of Cx43 in tanycyte homotypic and panglial coupling and show that Cx43 function influences the proliferative potential of hypothalamic cells.

CFTR channel in oocytes from Xenopus laevis and its regulation by xShroom1 protein.

Shroom is a family of related proteins linked to the actin cytoskeleton. xShroom1 is constitutively expressed in Xenopus laevis oocytes, and it is required for the expression of the epithelial sodium channel (ENaC). As there is a close relationship between ENaC and the cystic fibrosis transmembrane regulator (CFTR), we examined the action of xShroom1 on CFTR expression and activity. Biotinylation was used to measure CFTR surface expression, and currents were registered with voltage clamp when stimulated with forskolin and 3-isobutyl-1-methylxanthine. Oocytes were coinjected with CFTR complementary RNAs (cRNAs) and xShroom1 sense or antisense oligonucleotides. We observed an increment in CFTR currents and CFTR surface expression in oocytes coinjected with CFTR and xShroom1 antisense oligonucleotides. MG-132, a proteasome inhibitor, did not prevent the increment in currents when xShroom1 was suppressed by antisense oligonucleotides. In addition, we inhibited the delivery of newly synthesized proteins to the plasma membrane with BFA and we found that the half-life of plasma membrane CFTR was prolonged when coinjected with the xShroom1 antisense oligonucleotides. Chloroquine, an inhibitor of the late endosome/lysosome, did not significantly increase CFTR currents when xShroom1 expression was inhibited. The higher expression of CFTR when xShroom1 is suppressed is in concordance with the functional studies suggesting that the suppression of the xShroom1 protein resulted in an increment in CFTR currents by promoting the increase of the half-life of CFTR in the plasma membrane. The role of xShroom1 in regulating CFTR expression could be relevant in the understanding of the channel malfunction in several diseases.

[Information disclosure and decision making preferences of patients with advanced cancer in a Pain and Palliative Care Unit in Chile]. / ¿Cuánta información desean recibir y cómo prefieren tomar sus decisiones pacientes con cáncer avanzado atendidos en una Unidad del Programa Nacional de Dolor y Cuidados Paliativos en Chile?

UNLABELLED: Information disclosure and decision making process are important steps in advanced cancer patients management; however, there is no research done in this area in Chile. AIMS: To know the preferences of patients with advanced cancer related to information disclosure and style of decision making process. METHODS: Prospective observational study with patients in the Palliative Care Unit of Sótero del Río Hospital, in Santiago, Chile. The preferences were evaluated with a Disclosure Information and a Decision Making Preferences Questionnaire. RESULTS: 100 patients were recruited, 52% males, average age 63 years; 90% wanted to receive complete information about diagnosis and 89% complete information about prognosis. The preferences related to decision making process style were: 60% shared, 27% passive and 13% active. The expressed satisfaction with the information received was 89% and 87% with the way decisions were actually made. CONCLUSIONS: A majority of patients preferred to receive complete information about diagnosis and prognosis and to make shared decisions. The satisfaction with information disclosure and decision making process was very high. The data of this study supports the need of an adequate information disclosure and of exploring the individual preferences of our patients, with the goal of promoting an informed decision making process that respects the preferences of our patients.