New perspectives on an old disease: proteomics in cancer research

A report on the American Association for Cancer Research Conference 'Advances in Proteomics in Cancer Research', Amelia Island, USA, 27 February-2 March 2007

Paper de les aldo-ceto reductases en el metabolisme de retinoides : estudi funcional i estructural de l'AKR1B10 /

Consultable des del TDXTítol obtingut de la portada digitalitzadaPer primer cop, el nostre grup va identificar un enzim de la superfamilia de les aldo-ceto reductases (AKR) actiu amb retinoides. Atès que l'AKR1B12 presenta una identitat seqüencial pròxima al 70% amb l'AKR1B1 i l'AKR1B10 humanes, es va decidir ampliar l'estudi de l'activitat retinoide oxidoreductasa entre les AKR. En la present Tesi Doctoral, hem aprofundit en l'estudi de l'activitat retinal reductasa de membres de la superfamilia de les aldo-ceto reductases (AKR). El treball es va iniciar amb la caracterització in vitro de l'activitat amb retinoides d'AKR humanes de la família AKR1. En col·laboració amb el grup de la Dra. Natalia Kedishvili (actualment en el Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, EUA) es va realitzar una anàlisi comparativa de l'activitat retinol oxidoreductasa d'enzims de les superfamílies de les deshidrogenases/reductases de cadena mitjana (MDR), deshidrogenases/reductases de cadena curta (SDR) i AKR. Es va emprar un nou mètode basat en la solubilització de retinoides amb BSA, i anàlisi mitjançant HPLC. Així, es van poder determinar els valors de les constants cinètiques per a enzims humans de les tres superfamílies sense l'efecte d'inhibició del Tween-80, detergent utilitzat tradicionalment per a l'estudi cinètic de les MDR i AKR (Capítol I). Aquests resultats van posar de relleu que el valor de Km per a retinoides és similar (0,1-1 μM) en els enzims estudiats. Les principals diferències es centren en el valor de kcat, el qual va variar fins a tres ordres de magnitud entre els diferents enzims. Un altre resultat destacable, va ser la desmostració que cap dels enzims era capaç d'utilitzar com a substrat retinol, o retinal, unit a la proteïna cel·lular unidora de retinol (CRBPI). Fins el present treball, s'utilitzaven dos arguments per defensar un paper principal de les SDR en l'oxidació de retinol a retinal, primera etapa de síntesi de l'àcid retinoic: 1) Valors de Km inferiors de les SDR per als retinoides (fins a dos ordres de magnitud respecte les ADH). 2) Capacitat de reconèixer l'holoCRBPI com a substrat, una propietat aparentment exclusiva d'algunes SDR. El nostre estudi, a part de descartar aquests dos arguments, ha introduït les AKR com a tercer grup enzimàtic implicat en el metabolisme de retinoides. La segona part de la Tesi es centra en l'estudi estructural i funcional de l'AKR1B10, l'AKR que presenta una major eficiència catalítica per a la reducció del retinal. Aquest enzim és sobrexpressat en diferents tipus de càncers humans, el que va fer augmentar el nostre interès per determinar el seu paper en la ruta de síntesi de l'àcid retinoic. A part de l'estudi in vitro de l'activitat retinoide oxidoreductasa de l'enzim, també es va dur a terme un estudi in vivo. Utilitzant cèl·lules COS-1 com a model, i mitjançant transfecció transitòria, es va poder observar que AKR1B10 també participava en la reducció de retinal in vivo, però que en canvi no era capaç d'oxidar retinol. Finalment, i en col·laboració amb el grup de cristal·lografia de proteïnes del Dr. Ignasi Fita (Parc Científic de Barcelona-CSIC), es va obtenir l'estructura tridimencional del complex ternari AKR1B10-NADP+-tolrestat. El tolrestat és un inhibidor d'AKR1B1 àmpliament estudiat com a fàrmac en el tractament de les complicacions de la diabetis, però que també inhibeix AKR1B10 tant in vitro com in vivo. Tot i l'elevada identitat seqüencial entre AKR1B1 i AKR1B10, aquests enzims presenten constants cinètiques molt diferents per a la reducció del retinal. Així, gràcies a la resolució de l'estructura d'AKR1B10, esperem poder estudiar els determinants estructurals que confereixen a cada enzim les seves propietats cinètiques característiques, així com establir les bases per al disseny de nous inhibidors específics per a cadascun d'ells.Our group was the first to identify an enzyme from the aldo-keto reductase (AKR) superfamily, AKR1B12, active with retinoids (Crosas et al., 2001). Due to the fact that AKR1B12 shows a 70% sequence identity with the human enzymes AKR1B1 and AKR1B10, we decided to study more deeply the retinoid oxidoreductase activity within the AKRs. Here, we extended the study on the retinal reductase activity of AKR superfamily members. The work was initiated with the in vitro characterization of retinoid activities of human AKRs from the AKR1 family. In collaboration with Dr Natalia Kedishvili's group (Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, USA) we carried out a comparative analysis of retinol oxidoreductase activity of the enzymes from the medium chain dehydrogenases/reductases (MDR), short chain dehydrogenases/reductases (SDR) and AKR. A new method based on a retinoid solubilisation with BSA and analysis by HPLC was employed. Kinetic constant values were determined for human enzymes from the three superfamilies, without the inhibitory effect of Tween-80, a detergent traditionally employed for MDR and AKR kinetic studies. The Km values obtained with retinoids (0,1-1 μM) were similar for all the enzymes tested. Main differences were found in the kcat values, which varied up to 10000-fold between the studied enzymes. Furthermore, none of the studied enzymes was able to use retinol, nor retinal, bound to cellular retinol binding protein (CRBPI). So far, a major role for SDR in retinol oxidation was supported by: 1) SDR had lower reported Km values with retinoid (up to 100-fold lower than ADHs). 2) SDR were the only enzymes with reported ability to recognize holoCRBPI as a substrate. Our group refused both arguments and introduced AKRs as the third enzymatic group implicated in retinoid metabolism. The second part of the thesis was focused on the structural and functional study of AKR1B10, which is the AKR with the highest catalytic efficiency reducing retinal. The enzyme is overexpressed in different human cancers, which increased our interest to determine its role in the retinoic acid synthesis pathway. In addition to the study on the retinoid oxidoreductase activity in vitro, we performed an in vivo study. COS-1 cells were used as a model, and transitional transfection experiments were carried out. We observed that AKR1B10 participates in the retinal reduction in vivo, but it was unable to oxidate retinol. Finally, in collaboration with Dr. Ignasi Fita's group (Parc Científic de Barcelona-CSIC), the threedimensional structure of the AKR1B10-NADP+-tolrestat complex was obtained. Tolrestat is an AKR1B1 inhibitor widely studied as a drug for diabetes treatment. It also inhibits AKR1B10 in vitro and in vivo. Despite the high sequential identity shared between AKR1B1 and AKR1B10, the two enzymes showed extremely different kinetic constant values for retinal reduction. The AKR1B10 structure resolution might help us to study structural features for the retinoid specificity showed, and to propose new fundamentals in order to design new specific inhibitors for each enzyme

Livestock management at the Late Iron Age site of Baltarga (eastern Pyrenees): an integrated bio-geoarchaeological approach

Despite the important role of livestock farming amongst Iron Age communities living in mountain regions, there is little information about livestock management, and particularly stabling practises, breeding systems, and grazing/foddering patterns. The study of the ground floor of Building G in Tossal de Baltarga has provided valuable insights into these important issues and has given us a better understanding of the social and economic patterns involved in all these livestock activities. It revealed the existence of a stable from the Late Iron Age, thanks to unique in situ finds of the stabled animals, including four sheep, a goat, and a horse, in addition to a range of organic remains preserved by fire and penning deposits. It is the first documented to date in the northeastern Iberian Peninsula. Through an integrated bio-geoarchaeological approach, combining a range of analytic procedures, including osteology, dental microwear, stable isotopes, phytoliths, dung spherulite analyses, and thin-section micromorphology, for the first time, this study has provided new, high-resolution evidence of livestock management strategies. Specifically, the research shed light on animal penning and feeding practises, revealing variable herbivorous regimes between species, the practise of seasonal movements, and the possible use of fodder as the main dietary regime of the animals stabled there. At the same time, the Baltarga case-study illustrates an indoor production unit that could reveal possible private control of some domestic animals in the Pyrenean Late Iron Age.Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. LC is currently supported by a Ramón y Cajal contract (RYC2019-026732-I-AEI/10.13039/ 501100011033). CM has the financial support of the Secretaria d’Universitats i Recerca de la Generalitat de Catalunya and European Social Fund (ESF) “Investing in your future” (2022 FI_B2 00070). The funding for this research has been partially provided within the framework of the projects “Control, gestión y explotación del territorio en la Hispania romana”, PID2021-122879OB-I00, MICIN, and “PATCA-3”, Generalitat de Catalunya, 9071-55/2022

PyF2F: a robust and simplified fluorophore-to-fluorophore distance measurement tool for Protein interactions from Imaging Complexes after Translocation experiments

Structural knowledge of protein assemblies in their physiological environment is paramount to understand cellular functions at the molecular level. Protein interactions from Imaging Complexes after Translocation (PICT) is a live-cell imaging technique for the structural characterization of macromolecular assemblies in living cells. PICT relies on the measurement of the separation between labelled molecules using fluorescence microscopy and cell engineering. Unfortunately, the required computational tools to extract molecular distances involve a variety of sophisticated software programs that challenge reproducibility and limit their implementation to highly specialized researchers. Here we introduce PyF2F, a Python-based software that provides a workflow for measuring molecular distances from PICT data, with minimal user programming expertise. We used a published dataset to validate PyF2F’s performance

Signalment, Immunological and Parasitological Status and Clinicopathological Findings of Leishmania-Seropositive Apparently Healthy Dogs

Canine leishmaniosis caused by Leishmania infantum is a disease with a wide range of clinical manifestations. Epidemiological serosurveys performed in Europe often lack a thorough assessment of clinical health status of studied dogs. The aim of this study was to evaluate signalment, immunological and parasitological status and clinicopathological findings of L. infantum-seropositive apparently healthy dogs (n = 212) living in endemic areas. Routine laboratory tests, endpoint inhouse ELISA to quantify the anti-Leishmania antibodies, blood Leishmania qPCR and IFN-ELISA were performed. All dogs enrolled were L. infantum-seropositive and were classified as healthy (n = 105) or sick (n = 107) according to LeishVet guidelines. The sick group presented a higher proportion of medium to high antibody levels and positive qPCR and lower IFN-concentration compared to the healthy group. Sick dogs were mostly classified in LeishVet stage IIa. Biochemical alterations (98%) were the most common clinicopathological findings, with fewer urinary tract (46%) and hematological (40%) alterations. Apparently healthy L. infantum-seropositive dogs can be classified between truly healthy dogs and sick dogs with clinicopathological findings. Sick dogs presented medium to high seropositivity and parasitemia and low IFN- concentrations, and their most common clinicopathological abnormalities were serum protein alterations followed by proteinuria and lymphopenia

Highly Specific and Wide Range NO2 sensor with color readout

We present a simple and inexpensive method to implement a Griess-Saltzman-type reaction that combines the advantages of the liquid phase method (high specificity and fast response time) with the benefits of a solid implementation (easy to handle). We demonstrate that the measurements can be carried out using conventional RGB sensors; circumventing all the limitations around the measurement of the samples with spectrometers. We also present a method to optimize the measurement protocol and target a specific range of NO2 concentrations. We demonstrate that it is possible to measure the concentration of NO2 from 50 ppb to 300 ppm with high specificity and without modifying the Griess-Saltzman reagent

Declive cognitivo en la enfermedad de Alzheimer. Seguimiento de más de 3 años de una muestra de pacientes

Introducción. Las tasas de declive cognitivo en los pacientes con enfermedad de Alzheimer (EA) presentan variaciones debido a diversos factores. El objetivo del estudio fue determinar la influencia de la edad, escolaridad, género, actividades de la vida diaria (AVD) e inhibidores de la acetilcolinesterasa (IAChE) y memantina en el ritmo y tasas de declive cognitivo. Pacientes y métodos. Estudio retrospectivo de una muestra de 383 pacientes con EA, con evaluaciones neuropsicológicas durante un período superior a 3 años. Se utilizó como medida cognitiva el Cambridge Cognitive Examination (CAMCOG). Se agruparon los pacientes según su tasa de declive anual (TDA) y se realizaron análisis bivariante y de regresión lineal multivariante utilizando como variable dependiente la diferencia de puntuaciones en el CAMCOG (basal-final). Resultados. La menor edad (β = -0,23; p < 0,001), la mayor escolaridad (β = 0,26; p < 0,001) y el mayor deterioro de las AVD (β = 0,24; p < 0,001) estuvieron asociados a un mayor declive en todos los pacientes. Los fármacos tuvieron un efecto benéfico (β = -0,18; p = 0,011) en el grupo con menor y más lento declive (TDA < 5%). Conclusiones. La menor edad, la mayor escolaridad y el deterioro de las AVD se relacionan con un mayor declive cognitivo. Los IAChE y memantina tuvieron un efecto benéfico, enlenteciendo el declive en el grupo de pacientes con menor TD

Predictors of cognitive decline in Alzheimer"s disease and mild cognitive impairment using the CAMCOG: a five-year follow-up

Background: There are discrepant findings regarding which subscales of the Cambridge Cognitive Examination(CAMCOG) are able to predict cognitive decline. The study aimed to identify the baseline CAMCOG subscales that can discriminate between patients and predict cognitive decline in Alzheimer"s disease (AD)and mild cognitive impairment (MCI). Methods: This was a five-year case-control study of patients with cognitive impairment and a control group.Participants were grouped into AD (n = 121), MCI converted to dementia (MCI-Ad, n = 43), MCI-stable(MCI-St, n = 66), and controls (CTR, n = 112). Differences in the mean scores obtained by the four groupswere examined. Receiver operating characteristic curves were used to compare subscale scores in the AD and MCI-Ad groups with those of controls. The influence of age, gender, schooling, and depression on baseline subscale scores was assessed. Results: Of the CAMCOG subscales, Orientation and Memory (learning and recent) (OR + MEM) showed the highest discriminant capacity in the baseline analysis of the four groups. This baseline analysis indicated that OR + MEM was the best predictor of conversion to AD in the MCI-Ad group (area under the curve, AUC = 0.81), whereas the predictive capacity of the global MMSE and CAMCOG scores was poor (AUC = 0.59 and 0.53, respectively). Conclusions: In the baseline analysis, the Orientation and Memory (learning and recent) subscales showed the highest discriminant and predictive capacity as regards both cognitive decline in the AD group and conversion to AD among MCI-Ad patients. This was not affected by age, gender, schooling, or depression

Needle tract seeding and malignant transformation of hepatocellular adenoma into well-differentiated hepatocellular carcinoma in a dog

Altres ajuts: acords transformatius de la UABAn 11-year-old neutered female Golden Retriever was referred for investigation of marked increases in liver enzyme activities. Abdominal ultrasound revealed a large pedunculated liver mass. Diagnosis of hepatocellular adenoma (HCA) was made when the mass was excised after a first unsuccessful attempt through ultrasound-guided core-needle biopsy. One and a half years after presentation, a nodule embedded between muscles of the abdominal wall appeared. The mass was first diagnosed as a well-differentiated hepatocellular carcinoma (HCC) through cytologic examination, which was later confirmed with histopathology. Ki 67 immunostaining of the abdominal wall nodule showed an increased immunoreactivity compared with the liver mass. Therefore, the present case documents the first needle-tract seeding of a hepatocellular epithelial tumor with possible malignant transformation of HCA into a well-differentiated HCC in a dog