Desenvolupament de la tecnica de deteccio d'HPV en frotis de pell i bulbs pilosos de la cella per a l'aplicació com a marcador de risc en carcinogènesi cutània

Estudi realitzat a partir d’una estada al Department of Pathology de la Vrije Universiteit Medical Center, Holanda, entre agost del 2006 i gener del 2007. Es parteix de la hipòtesi principal que el virus del papil•loma humà (VPH) està implicat com a cofactor en la carcinogènesi del càncer cutani no-melanoma associat a l'exposició solar i a la immunosupressió. L'objectiu principal era la validació d'una tècnica de detecció del VPH en frotis cutanis per a determinar el paper d'aquest en el càncer cutani en pacients trasplantats renals. Es pretenia desenvolupar l'ús dels frotis cutanis per a la detecció del VPH en pell no tumoral i posteriorment establir quines són les mostres més adequades (en quan a localització i tipus d'extracció) per tal de definir el concepte de "portador de VPH". Es recolliren frotis cutanis de zones exposades (front i mà) i no exposades (part interna del braç) al sol, de la zona perilesional així com pèls de cella. Les mostres pertanyen tant a pacients trasplantats renals (immunodeprimits) com a pacients no trasplantats (immunocompetents), de pell normal i de pell cancerosa. Es van emprar diferents tècniques d'extracció de DNA. El DNA del VPH va ser amplificat amb una tècnica de reacció en cadena de la polimerasa (PCR ) específica de tipus cutanis (Beta-Gamma Cutaneous HPV PCR) i es va tipificar amb una hibridació reversa amb sondes específiques (Reverse Line Blotting). Tots els assajos es van fer per triplicat, per tal de poder avaluar la reproduibilitat dels resultats en aquestes mostres. Com a control de la qualitat i la quantitat del DNA les mostres van ser testades per la PCR del gen de la beta-globina. S’ha dectectat el VPH present tant en pell com en pell cancerosa. La tècnica aporta resultats reproduïbes. S’aprecia una bona correlació tant entre els resultats obtinguts dels frotis i el bulbs pilosos, així com de diferents zones raspades d'un mateix pacient.

Precursor uptake assays and metabolic analyses in isolated tomato fruit chromoplasts

Background Carotenoids are the most widespread group of pigments found in nature. In addition to their role in the physiology of the plant, carotenoids also have nutritional relevance as their incorporation in the human diet provides health benefits. In non-photosynthetic tissues, carotenoids are synthesized and stored in specialized plastids called chromoplasts. At present very little is known about the origin of the metabolic precursors and cofactors required to sustain the high rate of carotenoid biosynthesis in these plastids. Recent proteomic data have revealed a number of biochemical and metabolic processes potentially operating in fruit chromoplasts. However, considering that chloroplast to chromoplast differentiation is a very rapid process during fruit ripening, there is the possibility that some of the proteins identified in the proteomic analysis could represent remnants no longer having a functional role in chromoplasts. Therefore, experimental validation is necessary to prove whether these predicted processes are actually operative in chromoplasts. Results A method has been established for high-yield purification of tomato fruit chromoplasts suitable for metabolic studies. Radiolabeled precursors were efficiently incorporated and further metabolized in isolated chromoplast. Analysis of labeled lipophilic compounds has revealed that lipid biosynthesis is a very efficient process in chromoplasts, while the relatively low incorporation levels found in carotenoids suggest that lipid production may represent a competing pathway for carotenoid biosynthesis. Malate and pyruvate are efficiently converted into acetyl-CoA, in agreement with the active operation of the malic enzyme and the pyruvate dehydrogenase complex in the chromoplast. Our results have also shown that isolated chromoplasts can actively sustain anabolic processes without the exogenous supply of ATP, thus suggesting that these organelles may generate this energetic cofactor in an autonomous way. Conclusions We have set up a method for high yield purification of intact tomato fruit chromoplasts suitable for precursor uptake assays and metabolic analyses. Using targeted radiolabeled precursors we have been able to unravel novel biochemical and metabolic aspects related with carotenoid and lipid biosynthesis in tomato fruit chromoplasts. The reported chromoplast system could represent a valuable platform to address the validation and characterization of functional processes predicted from recent transcriptomic and proteomic data.

Diphenyl urea derivatives as inhibitors of transketolase: a structure-based virtual screening.

Transketolase is an enzyme involved in a critical step of the non-oxidative branch of the pentose phosphate pathway whose inhibition could lead to new anticancer drugs. Here, we report new human transketolase inhibitors, based on the phenyl urea scaffold, found by applying structure-based virtual screening. These inhibitors are designed to cover a hot spot in the dimerization interface of the homodimer of the enzyme, providing for the first time compounds with a suggested novel binding mode not based on mimicking the thiamine pyrophosphate cofactor.

Relationship between oral cancer and implants: clinical cases and systematic literature review

The use of implants for oral rehabilitation of edentulous spaces has recently been on the increase, which has also led to an increase in complications such as peri-implant inflammation or peri-implantitis. Chronic inflammation is a risk factor for developing oral squamous cell carcinoma (OSCC). Objectives: To review the literature of cases that associate implant placement with the development of oral cancer. Study design: We present two clinical cases and a systematic review of literature published on the relationship between oral cancer and implants. Results: We found 13 articles published between the years 1996 and 2009, referencing 18 cases in which the osseointegrated implants are associated with oral squamous cell carcinoma. Of those, 6 articles were excluded because they did not meet the inclusion criteria. Of the 18 cases reported, only 7 cases did not present a previous history of oral cancer or cancer in other parts of the body. Conclusions: Based on the review of these cases, a clear cause-effect relationship cannot be established, although it can be deduced that there is a possibility that implant treatment may constitute an irritant and/or inflammatory cofactor which contributes to the formation and/or development of OSCC.

Concerted changes in N and C primary metabolism in alfalfa (Medicago sativa) under water restriction

Although the mechanisms of nodule N2 fixation in legumes are now well documented, some uncertainty remains on the metabolic consequences of water deficit. In most cases, little consideration is given to other organs and, therefore, the coordinated changes in metabolism in leaves, roots, and nodules are not well known. Here, the effect of water restriction on exclusively N2-fixing alfalfa (Medicago sativa L.) plants was investigated, and proteomic, metabolomic, and physiological analyses were carried out. It is shown that the inhibition of nitrogenase activity caused by water restriction was accompanied by concerted alterations in metabolic pathways in nodules, leaves, and roots. The data suggest that nodule metabolism and metabolic exchange between plant organs nearly reached homeostasis in asparagine synthesis and partitioning, as well as the N demand from leaves. Typically, there was (i) a stimulation of the anaplerotic pathway to sustain the provision of C skeletons for amino acid (e.g. glutamate and proline) synthesis; (ii) re-allocation of glycolytic products to alanine and serine/glycine; and (iii) subtle changes in redox metabolites suggesting the implication of a slight oxidative stress. Furthermore, water restriction caused little change in both photosynthetic efficiency and respiratory cost of N2 fixation by nodules. In other words, the results suggest that under water stress, nodule metabolism follows a compromise between physiological imperatives (N demand, oxidative stress) and the lower input to sustain catabolism.