Construcción de un modelo de colon proximal humano para el estudio de prebióticos

El colon proximal humano es la región del intestino grueso donde se ha reportado e mayor efecto prebiótico, sin embargo, no es un sitio fácilmente accesible para estudios de rutina. Para realizar estudios de ecología microbiana en esta región se han desarrollado modelos de laboratorio que imitan algunas características del sistema digestivo. El objetivo de este trabajo fue construir un modelo de laboratorio con las condiciones de fermentación y absorción del colon proximal humano, así como determinar la respuesta a la adición de inulina, almidón resistente de plátano y fibra dietética de naranja. El modelo se construyó empleando un tubo de vidrio (50 x 5 cm) con una membrana de diálisis en su interior. El sustrato se introdujo en la membrana de diálisis tres veces al día, simulando una alimentación humana típica. La composición del sustrato incluyó 58% de carbohidratos, 35% de proteínas, 3% de fibra, 3% de almidón, y 1% de lípidos en base seca, con el 90% de humedad. Como inóculo se utilizó un cultivo de biomasa fecal propagado en caldo de soya y tripticaseina (TSB). La absorción intestinal fue simulada mediante el flujo continuo de una solución de polietilenglicol (PEG) alrededor de la membrana de diálisis. Se logró el cultivo de biomasa fecal para su empleo como inóculo. El inóculo se obtuvo con una concentración de microorganismos y ácidos grasos de cadena corta (AGCC) semejante a la encontrada en solución fecal. Con la propagación se mejoraron las condiciones de manejo y transporte, se aumentó la concentración de microorganismos anaerobios y se eliminó la interferencia por sustancias desconocidas. En el modelo del colon proximal se obtuvo crecimiento de microorganismos aerobios y anaerobios así como producción de ácidos grasos de cadena corta en concentraciones similares a las reportadas por otros autores para la región del colon. Todos los microorganismos aumentaron su cuenta después de la inoculación del sustrato y se obtuvo el mayor crecimiento después de cada alimentación. La concentración de AGCC dentro y fuera de la membrana fue significativamente diferente debido a la eficacia de extracción de la solución de PEG. La mayor producción se obtuvo a las 48 h. Al principio, el acetato fue el compuesto predominante, pero después de 12 h la proporción de butirato aumentó y el acetato disminuyó. Esta producción de AGCC fue similar a la del colon proximal en los sistemas vivos. El funcionamiento continuo del modelo del colon durante 48 h fue suficiente para obtener un buen desarrollo de microorganismos y la producción de AGCC. Este modelo reproduce las condiciones del ser humano de colon proximal de manera adecuada y puede ser utilizado para estudiar el desarrollo de la microbiota colónica. La adición de inulina en el modelo provocó el aumento en la concentración de anaerobios totales y facultativos, así como en lactobacilos y bifidobacterias. Aunque en la concentración de mesófilos aerobios se presentó un aumento, en coliformes y enterobacterias hubo una disminución en la concentración. La concentración de clostridios se mantuvo igual tanto a las 24 h como a las 48 h; su crecimiento no fue afectado por la adición de inulina. Se demostró la sensibilidad del sistema modelo del colon para mostrar pequeños cambios que ocurren en el metabolismo de la microflora colónica ante un estímulo alimenticio como lo es la adición de inulina. Se obtuvo la caracterización fisicoquímica y funcional de almidón resistente de plátano y fibra dietética de naranja. El almidón de plátano Musa balbisiana Colla contiene más de la mitad de almidón resistente a la digestión. La harina de naranja contiene alrededor de 40% de fibra dietética, de la cual, la mitad corresponde a fibra dietética soluble. Al evaluar estos sustratos en el modelo de colon se obtuvo una disminución en la concentración de enterobacterias y un aumento en lactobacilos y bifidobacterias. Con los almidones se obtuvo una mayor concentración de AGCC y se estimuló la producción de ácido butírico, mientras que con la fibra de naranja y pectina sólo se obtuvo formación de ácido acético. Con el modelo de colon se contribuirá a aumentar el conocimiento sobre la ecología colónica, así como el efecto de diversas sustancias en su crecimiento y metabolismo.The human proximal colon is the region of the large intestine where the greater prebiotic effect has been reported. However, this is not a region easily accessible to routine studies. In order to carry out studies on microbial ecology in this region, laboratory models that imitate some characteristics of the digestive system have been developed. The aim of this research was to build up a laboratory model with the same conditions of fermentation and adsorption than the human proximal colon, as well as to determine the response to the addition of inulin, resistant banana starch and orange dietary fiber. The model was constructed using a glass tube (50 x 5 cm) with a dialysis membrane inside. The substrate was introduced in the dialysis membrane three times a day, simulating a typical human food. The composition of the substrate was 58% carbohydrates, 35% protein, 3% fiber, 3% starch, 1% lipids in dry weight basis with 90% moisture. As inoculum it was used fecal cultivated biomass propagated in trypticasein soy broth (TSB). The intestinal absorption was simulated by means of a continuous flow of a solution of polyethylene glycol (PEG) outside the dialysis membrane. Grow of biomass to be used as fecal inoculum was achieved. The inoculum obtained had a concentration of microorganisms and short-chain fatty acids (SCFA) similar to that found in a fecal solution. The use of a propagated inoculum improves conditions for handling and transportation, increases concentration of anaerobic microorganisms, and avoid interference of unknown substances. In the proximal human colon model the growth of both aerobic and anaerobic microorganisms was obtained, as well as the production of short-chain fatty acids, similarly to those concentrations reported by other authors for this region. All microorganisms increased their count after inoculation of the substrate, and higher growths were obtained after each feeding. The concentration of SCFA inside and outside the membrane was significantly different due to the extraction efficiency of the PEG solution. The highest production was reached at 48 hours. At the beginning acetate was the predominant compound, but after 12 hours the proportion of butyrate increased while acetate decreased. This production of SCFA was similar to that of proximal colon in living systems. The continuous operation of the colon model for 48 hours was enough to obtain a good development of microorganisms and of SCFA production. This model reproduced the conditions of the human proximal colon adequately and can be used to study the development of the colonic microbiota. The addition of inulin in the model led to an increase in the concentration of total and facultative anaerobes, as well as lactobacilli and bifidobacteria. On the other hand, in spite that the aerobic mesophilic count was increased, the concentration of coliforms and enterobacteria decreased. The concentration of clostridia remained the same at 24 hours until 48 hours; its growth was not affected by the addition of inulin. It was demonstrated the susceptibility of the colon model system to small changes in the metabolism of colonic microflora due to food stimulus such as the addition of inulin. The physicochemical and functional characterization o resistant starch of banana and orange dietary fiber was obtained. The starch from banana Musa balbisiana Colla contains more than a half of digestion resistant starch. Orange flour contains about 40% of dietary fiber, from which, a half is soluble dietary fiber. When these substrates were assessed in the colon model, a decrease in the concentration of enterobacteria was obtained, while an increase in lactobacilli and bifidobacteria resulted. With starch a grater concentration of SCFA was obtained and the production of butyric acid was stimulated, while with orange fiber and pectin only the formation of acetic acid was got. With this model it is possible to contribute to the knowledge of the ecology of colonic microbiota, as well as the effect of different substances in its growth and metabolism

Chromogenic in situ hybridization (CISH): a novel alternative in screening archival breast cancer tissue samples for HER-2/neu status

BACKGROUND: Chromogenic in situ hybridization (CISH) is emerging as a practical, cost-effective, and valid alternative to fluorescent in situ hybridization in testing for gene alteration, especially in centers primarily working with immunohistochemistry (IHC). METHODS: We assessed Her-2/neu alteration using CISH on formalin-fixed paraffin-embedded primary invasive ductal carcinoma tumors in which IHC (CB11 antibody) had previously been performed, and we compared the results with IHC. The 160 selected cases were equally stratified randomly into the four IHC categories (scores of 0, 1+, 2+, and 3+). We also compared age at diagnosis and tumor histologic grade with IHC and CISH Her-2/neu. RESULTS: We were able to perform and evaluate CISH successfully on all cases. The agreement between 3+ IHC and CISH-amplified cases as well as between all IHC and CISH Her-2/neu negative cases was 100%, and the concordance on all positive cases was 72.50%, with an overall agreement of 86.25%. All the discordant cases had 2+ IHC scores. Although we noted Her-2/neu positivity more in premenopausal women, the age at diagnosis was not significantly associated with IHC or CISH results. Similarly, although the small group of well-differentiated tumors was apparently Her-2/neu negative in both tests, no significant association was noted between any tumor histologic grade and either IHC or CISH results. CONCLUSIONS: CISH is easily integrated into routine testing in our laboratory. It is a necessary adjunct in determining the subset of non-amplified IHC-positive invasive tumors that will not benefit from trastuzumab therapy. Those cases with 2+ IHC results will be triaged and subjected to CISH. Her-2/neu testing should be done on all breast cancer cases regardless of age at presentation and tumor histologic grade

A Prospective, Multicenter, Real-World Registry of Coronary Lithotripsy in Calcified Coronary Arteries

BACKGROUND Intravascular lithotripsy (IVL) has demonstrated effectiveness in the treatment of calcified lesions in selected patients with stable coronary disease. OBJECTIVES The authors sought to assess the performance of coronary IVL in calcified coronary lesions in a real-life, all comers, setting. METHODS The REPLICA-EPIC18 study prospectively enrolled consecutive patients treated with IVL in 26 centers in Spain. An independent core laboratory performed the angiographic analysis and event adjudication. The primary effectiveness endpoint assessed procedural success (successful IVL delivery, final diameter stenosis <20%, and absence of in- hospital major adverse cardiovascular events [MACE]). The primary safety endpoint measured freedom from MACE at 30 days. A predefined substudy compared outcomes between acute coronary syndrome (ACS) and chronic coronary syndrome (CCS) patients. RESULTS A total of 426 patients (456 lesions) were included, 63% of the patients presenting with ACS. IVL delivery was successful in 99% of cases. Before IVL, 49% of lesions were considered undilatable. The primary effectiveness endpoint was achieved in 66% of patients, with similar rates among CCS patients (68%) and ACS patients (65%). Likewise, there were no significant differences in angiographic success after IVL between CCS and ACS patients. The rate of MACE at 30 days (primary safety endpoint) was 3% (1% in CCS and 5% in ACS patients [P = 0.073]). CONCLUSIONS Coronary IVL proved to be a feasible and safe procedure in a real-life setting, effectively facilitating stent implantation in severely calcified lesions. Patients with ACS on admission showed similar angiographic success rates but showed a trend toward higher 30-day MACE compared with patients with CCS. (REPLICA-EPIC18 study [Registry of Coronary Lithotripsy in Spain]; NCT04298307) (c) 2024 The Authors. Published by Elsevier on behalf of the American College of Cardiology Foundation. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)

Psychometric properties of the Alcohol Use Disorders Identification Test (AUDIT) across cross-cultural subgroups, genders, and sexual orientations: Findings from the International Sex Survey (ISS)

Introduction. Despite being a widely used screening questionnaire, there is no consensus on the most appropriate measurement model for the Alcohol Use Disorders Identification Test (AUDIT). Furthermore, there have been limited studies on its measurement invariance across cross-cultural subgroups, genders, and sexual orientations. Aims. The present study aimed to examine the fit of different measurement models for the AUDIT and its measurement invariance across a wide range of subgroups by country, language, gender, and sexual orientation. Methods. Responses concerning past-year alcohol use from the participants of the cross-sectional International Sex Survey were considered (N = 62,943; Mage: 32.73; SD = 12.59). Confirmatory factor analysis, as well as measurement invariance tests were performed for 21 countries, 14 languages, three genders, and four sexual-orientation subgroups that met the minimum sample size requirement for inclusion in these analyses. Results. A two-factor model with factors describing ‘alcohol use’ (items 1–3) and ‘alcohol problems’ (items 4–10) showed the best model fit across countries, languages, genders, and sexual orientations. For the former two, scalar and latent mean levels of invariance were reached considering different criteria. For gender and sexual orientation, a latent mean level of invariance was reached. Conclusions. In line with the two-factor model, the calculation of separate alcohol-use and alcohol-problem scores is recommended when using the AUDIT. The high levels of measurement invariance achieved for the AUDIT support its use in cross-cultural research, capable also of meaningful comparisons among genders and sexual orientations

Choice of the initial antiretroviral treatment for HIV-positive individuals in the era of integrase inhibitors

BACKGROUND: We aimed to describe the most frequently prescribed initial antiretroviral therapy (ART) regimens in recent years in HIV-positive persons in the Cohort of the Spanish HIV/AIDS Research Network (CoRIS) and to investigate factors associated with the choice of each regimen. METHODS: We analyzed initial ART regimens prescribed in adults participating in CoRIS from 2014 to 2017. Only regimens prescribed in >5% of patients were considered. We used multivariable multinomial regression to estimate Relative Risk Ratios (RRRs) for the association between sociodemographic and clinical characteristics and the choice of the initial regimen. RESULTS: Among 2874 participants, abacavir(ABC)/lamivudine(3TC)/dolutegavir(DTG) was the most frequently prescribed regimen (32.1%), followed by tenofovir disoproxil fumarate (TDF)/emtricitabine (FTC)/elvitegravir(EVG)/cobicistat(COBI) (14.9%), TDF/FTC/rilpivirine (RPV) (14.0%), tenofovir alafenamide (TAF)/FTC/EVG/COBI (13.7%), TDF/FTC+DTG (10.0%), TDF/FTC+darunavir/ritonavir or darunavir/cobicistat (bDRV) (9.8%) and TDF/FTC+raltegravir (RAL) (5.6%). Compared with ABC/3TC/DTG, starting TDF/FTC/RPV was less likely in patients with CD4100.000 copies/mL. TDF/FTC+DTG was more frequent in those with CD4100.000 copies/mL. TDF/FTC+RAL and TDF/FTC+bDRV were also more frequent among patients with CD4<200 cells//muL and with transmission categories other than men who have sex with men. Compared with ABC/3TC/DTG, the prescription of other initial ART regimens decreased from 2014-2015 to 2016-2017 with the exception of TDF/FTC+DTG. Differences in the choice of the initial ART regimen were observed by hospitals' location. CONCLUSIONS: The choice of initial ART regimens is consistent with Spanish guidelines' recommendations, but is also clearly influenced by physician's perception based on patient's clinical and sociodemographic variables and by the prescribing hospital location

Comparative effectiveness and safety of non-vitamin K antagonists for atrial fibrillation in clinical practice: GLORIA-AF Registry

Background and purpose: Prospectively collected data comparing the safety and effectiveness of individual non-vitamin K antagonists (NOACs) are lacking. Our objective was to directly compare the effectiveness and safety of NOACs in patients with newly diagnosed atrial fibrillation (AF). Methods: In GLORIA-AF, a large, prospective, global registry program, consecutive patients with newly diagnosed AF were followed for 3&nbsp;years. The comparative analyses for (1) dabigatran vs rivaroxaban or apixaban and (2) rivaroxaban vs apixaban were performed on propensity score (PS)-matched patient sets. Proportional hazards regression was used to estimate hazard ratios (HRs) for outcomes of interest. Results: The GLORIA-AF Phase III registry enrolled 21,300 patients between January 2014 and December 2016. Of these, 3839 were prescribed dabigatran, 4015 rivaroxaban and 4505 apixaban, with median ages of 71.0, 71.0, and 73.0&nbsp;years, respectively. In the PS-matched set, the adjusted HRs and 95% confidence intervals (CIs) for dabigatran vs rivaroxaban were, for stroke: 1.27 (0.79–2.03), major bleeding 0.59 (0.40–0.88), myocardial infarction 0.68 (0.40–1.16), and all-cause death 0.86 (0.67–1.10). For the comparison of dabigatran vs apixaban, in the PS-matched set, the adjusted HRs were, for stroke 1.16 (0.76–1.78), myocardial infarction 0.84 (0.48–1.46), major bleeding 0.98 (0.63–1.52) and all-cause death 1.01 (0.79–1.29). For the comparison of rivaroxaban vs apixaban, in the PS-matched set, the adjusted HRs were, for stroke 0.78 (0.52–1.19), myocardial infarction 0.96 (0.63–1.45), major bleeding 1.54 (1.14–2.08), and all-cause death 0.97 (0.80–1.19). Conclusions: Patients treated with dabigatran had a 41% lower risk of major bleeding compared with rivaroxaban, but similar risks of stroke, MI, and death. Relative to apixaban, patients treated with dabigatran had similar risks of stroke, major bleeding, MI, and death. Rivaroxaban relative to apixaban had increased risk for major bleeding, but similar risks for stroke, MI, and death. Registration: URL: https://www.clinicaltrials.gov. Unique identifiers: NCT01468701, NCT01671007. Date of registration: September 2013

Anticoagulant selection in relation to the SAMe-TT2R2 score in patients with atrial fibrillation. the GLORIA-AF registry

Aim: The SAMe-TT2R2 score helps identify patients with atrial fibrillation (AF) likely to have poor anticoagulation control during anticoagulation with vitamin K antagonists (VKA) and those with scores &gt;2 might be better managed with a target-specific oral anticoagulant (NOAC). We hypothesized that in clinical practice, VKAs may be prescribed less frequently to patients with AF and SAMe-TT2R2 scores &gt;2 than to patients with lower scores. Methods and results: We analyzed the Phase III dataset of the Global Registry on Long-Term Oral Antithrombotic Treatment in Patients with Atrial Fibrillation (GLORIA-AF), a large, global, prospective global registry of patients with newly diagnosed AF and ≥1 stroke risk factor. We compared baseline clinical characteristics and antithrombotic prescriptions to determine the probability of the VKA prescription among anticoagulated patients with the baseline SAMe-TT2R2 score &gt;2 and ≤ 2. Among 17,465 anticoagulated patients with AF, 4,828 (27.6%) patients were prescribed VKA and 12,637 (72.4%) patients an NOAC: 11,884 (68.0%) patients had SAMe-TT2R2 scores 0-2 and 5,581 (32.0%) patients had scores &gt;2. The proportion of patients prescribed VKA was 28.0% among patients with SAMe-TT2R2 scores &gt;2 and 27.5% in those with scores ≤2. Conclusions: The lack of a clear association between the SAMe-TT2R2 score and anticoagulant selection may be attributed to the relative efficacy and safety profiles between NOACs and VKAs as well as to the absence of trial evidence that an SAMe-TT2R2-guided strategy for the selection of the type of anticoagulation in NVAF patients has an impact on clinical outcomes of efficacy and safety. The latter hypothesis is currently being tested in a randomized controlled trial. Clinical trial registration: URL: https://www.clinicaltrials.gov//Unique identifier: NCT01937377, NCT01468701, and NCT01671007

Cabbage and fermented vegetables : From death rate heterogeneity in countries to candidates for mitigation strategies of severe COVID-19

Large differences in COVID-19 death rates exist between countries and between regions of the same country. Some very low death rate countries such as Eastern Asia, Central Europe, or the Balkans have a common feature of eating large quantities of fermented foods. Although biases exist when examining ecological studies, fermented vegetables or cabbage have been associated with low death rates in European countries. SARS-CoV-2 binds to its receptor, the angiotensin-converting enzyme 2 (ACE2). As a result of SARS-CoV-2 binding, ACE2 downregulation enhances the angiotensin II receptor type 1 (AT(1)R) axis associated with oxidative stress. This leads to insulin resistance as well as lung and endothelial damage, two severe outcomes of COVID-19. The nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is the most potent antioxidant in humans and can block in particular the AT(1)R axis. Cabbage contains precursors of sulforaphane, the most active natural activator of Nrf2. Fermented vegetables contain many lactobacilli, which are also potent Nrf2 activators. Three examples are: kimchi in Korea, westernized foods, and the slum paradox. It is proposed that fermented cabbage is a proof-of-concept of dietary manipulations that may enhance Nrf2-associated antioxidant effects, helpful in mitigating COVID-19 severity.Peer reviewe

Nrf2-interacting nutrients and COVID-19 : time for research to develop adaptation strategies

There are large between- and within-country variations in COVID-19 death rates. Some very low death rate settings such as Eastern Asia, Central Europe, the Balkans and Africa have a common feature of eating large quantities of fermented foods whose intake is associated with the activation of the Nrf2 (Nuclear factor (erythroid-derived 2)-like 2) anti-oxidant transcription factor. There are many Nrf2-interacting nutrients (berberine, curcumin, epigallocatechin gallate, genistein, quercetin, resveratrol, sulforaphane) that all act similarly to reduce insulin resistance, endothelial damage, lung injury and cytokine storm. They also act on the same mechanisms (mTOR: Mammalian target of rapamycin, PPAR gamma:Peroxisome proliferator-activated receptor, NF kappa B: Nuclear factor kappa B, ERK: Extracellular signal-regulated kinases and eIF2 alpha:Elongation initiation factor 2 alpha). They may as a result be important in mitigating the severity of COVID-19, acting through the endoplasmic reticulum stress or ACE-Angiotensin-II-AT(1)R axis (AT(1)R) pathway. Many Nrf2-interacting nutrients are also interacting with TRPA1 and/or TRPV1. Interestingly, geographical areas with very low COVID-19 mortality are those with the lowest prevalence of obesity (Sub-Saharan Africa and Asia). It is tempting to propose that Nrf2-interacting foods and nutrients can re-balance insulin resistance and have a significant effect on COVID-19 severity. It is therefore possible that the intake of these foods may restore an optimal natural balance for the Nrf2 pathway and may be of interest in the mitigation of COVID-19 severity