Aberrant methylation within RUNX3 CpG island associated with the nuclear and mitochondrial microsatellite instability in sporadic gastric cancers. Results of a GOIM (Gruppo Oncologico dell'Italia Meridionale) prospective study.

Gastric cancer (GC) development is a multistep process, during which numerous alterations accumulate in nuclear and mitochondrial DNA. A deficiency of repair machinery brings about an accumulation of errors introduced within simple repetitive microsatellite sequences during replication of DNA. Aberrant methylation is related to microsatellite instability (MSI) by the silencing of the hMLH1 gene. The aim of this study is to investigate a possible relationship between the RUNX3 promoter methylation, nuclear microsatellite instability (nMSI) and mitochondrial microsatellite instability (mtMSI), in order to clarify its biological role in GC

Laser Pressure Catapulting (LPC): Optimization LPC-System and Genotyping of Colorectal Carcinomas

Genotype analysis is becoming more and more useful in clinical practice, since specific mutations in tumors often correlate with prognosis and/or therapeutic response. Unfortunately, current molecular analytical techniques often require time-consuming and costly steps of analysis, thus making their routine clinical use difficult. Moreover, one of the most difficult problems arising during tumor research is that of their cell heterogeneity, which depends on their clear molecular heterogeneity. SSCP analysis discriminates by means of aberrant electrophoresis migration bands, mutated alleles which may represent as little as 15-20% of their total number. Nevertheless, in order to identify by sequencing the type of alteration revealed by this technique, only the mutated allele must be isolated. The advent of laser microdissection is a procedure which easily solves these problems of accuracy, costs, and time. The aims of this study were to perfect the system of laser pressure catapulting (LPC) laser microdissection for the assessment of the mutational status of p53 and k-ras genes in a consecutive series of 67 patients with colorectal carcinomas (CRC), in order to compare this technique with that involving hand-dissection and to demonstrate that since the LPC system guarantees more accurate biomolecular analyses, it should become part of clinical routine in this field. The LPC-system was perfected with the use of mineral oil and the LPC-membrane. To compare the techniques of hand- and LPC-microdissection, alcohol-fixed, paraffin-embedded tissue from 67 cases of CRC were both hand- and laser-microdissected. In either case, dissected samples were analyzed by SSCP/sequencing and direct sequencing for k-ras and p53 gene mutations. LPC-microdissection made it possible to pick up mutations by direct sequencing or SSCP/sequencing, whereas hand-microdissection mutations were identified only by means of SSCP followed by sequencing; direct sequencing did not reveal any mutation. In the 67 patients examined by either method, 36% (24/67) showed p53 mutations, 32 of which identified. Seventy-eight percent (25/32) were found in the conserved areas of the gene, while 12% (4/32) were in the L2 loop, 50% (16/32) were in the L3 loop, and 12% (4/32) in the LSH motif of the protein. Moreover, of the 67 cases examined, 40% (27/67) showed mutations in k-ras, with a total of 29 mutations identified. Of these, 14 (48%) were found in codon 12 and 15 (52%) in codon 13. The modifications which we brought to the LPC system led to a vast improvement of the technique, making it an ideal substitution for hand-microdissection and guaranteeing a considerable number of advantages regarding facility, accuracy, time, and cost. Furthermore, the data obtained from the mutational analyses performed confirm that the LPC system is more efficient and rapid than hand-microdissection for acquiring useful information regarding molecular profile and can therefore be used with success in clinical routine

Surface mannosylation of dispersion polymerisation derived nanoparticles by copper mediated click chemistry

The synthesis of spherical polymeric nanoparticles containing alkyne surface functionalities for post poly- merisation glycosylation is described. The nanoparticles were obtained by a polymerisation induced self- assembly (PISA) inspired methodology in dispersed media by Cu(0) mediated polymerisation. A water soluble poly(ethylene glycol methacrylate-stat-propargyl methacrylate), poly(PEGMA18-stat-PgMA5), macroinitiator was first synthesised and chain extended with 2-hydroxypropyl methacrylate (HPMA) in water using a copper wire catalyst. It was found that irrespective of the macroinitiator to HPMA ratio and the reaction time the desired spherical morphologies (<100 nm) were obtained while the absence other morphologies suggest a deviation from the classical PISA process due to chain termination in the nano- particle’s core. The obtained nanoparticles contained alkyne functionalities in the shell, which were suc- cessfully reacted by copper mediated click chemistry with fluoresceine azide and mannosides with hydro- phobic and hydrophilic spacers of different lengths. The obtained mannosylated nanoparticles displayed no significant cytotoxicity against human alveolar basal epithelial adenocarcinomic (A549) cells at any dose <0.5 mg mL−1. Preliminary binding studies confirm the ability of the mannosylated nanoparticles to bind to human lectin dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN). The methodology reported here is a convenient route to well-defined spherical and shell- functionalisable nanoparticles to create libraries of bio-active nanomaterials

Significance of P16INK4A hypermethylation gene in primary head/neck and colorectal tumors: it is a specific tissue event? Results of a 3-year GOIM (Gruppo Oncologico dell'Italia Meridionale) prospective study

BACKGROUND: Methylation of the p16 promoter is one of the most frequent mechanisms of gene inactivation; its incidence is extremely variable according to the type of tumor involved. Our purpose was to analyze the hypermethylation of the p16 promoter in laryngeal squamous cell carcinomas (LSCC), salivary gland (SG) tumors and in colorectal cancer (CRC), to detect any possible association with the clinicopathological features and to determine the prognostic significance of the p16 gene in the tumors analyzed. PATIENTS AND METHODS: The hypermethylation of the p16 promoter was prospectively analyzed, by MSP, in a consecutive series of 64 locally advanced LSCC patients, in a consecutive series of 33 SG tumor patients and in a consecutive series of 66 sporadic CRC patients. RESULTS: Hypermethylation was observed in 9% of the LSCC cases, in all cases of SG cancer and in 21% of the CRC cases. No significant association was observed between p16 hypermethylation and clinicopathological variables in all the tissue samples analyzed. Moreover at univariate analysis p16 mutations were not independently related at disease relapse and death in LSCC and CRC. CONCLUSIONS: The results of this study suggest that the lack of p16 function could happen in advanced stage of SG tumors

Grip Force Reveals the Context Sensitivity of Language-Induced Motor Activity during “Action Words

Studies demonstrating the involvement of motor brain structures in language processing typically focus on \ud time windows beyond the latencies of lexical-semantic access. Consequently, such studies remain inconclusive regarding whether motor brain structures are recruited directly in language processing or through post-linguistic conceptual imagery. In the present study, we introduce a grip-force sensor that allows online measurements of language-induced motor activity during sentence listening. We use this tool to investigate whether language-induced motor activity remains constant or is modulated in negative, as opposed to affirmative, linguistic contexts. Our findings demonstrate that this simple experimental paradigm can be used to study the online crosstalk between language and the motor systems in an ecological and economical manner. Our data further confirm that the motor brain structures that can be called upon during action word processing are not mandatorily involved; the crosstalk is asymmetrically\ud governed by the linguistic context and not vice versa

p53 mutations in L3-loop zinc-binding domain, DNA-ploidy, and S phase fraction are independent prognostic indicators in colorectal cancer: A prospective study with a five-year follow-up

p53 gene alterations are among the most common events observed in colorectal cancer, and are accompanied frequently by DNA aneuploidy and high proliferative activity. The prognostic significance of such mutations remains controversial. We prospectively evaluated the prognostic significance of p53 mutations, DNA-ploidy, and S phase fraction (SPF) in a consecutive series of 160 colorectal cancer patients (median follow-up 71 months). Tumor DNA was screened for p53 mutations by PCR/single-strand conformational polymorphism/sequencing. DNA-ploidy and SPF were assessed by DNA flow cytometry. p53 mutations were detected in 68 of 160 (42.5%) cases. In 56% (38 of 68) of these, p53 mutations were found in conserved areas of the gene and in 44% (30 of 68 cases) outside the conserved regions. Eighteen of the 68 cases (26%) had mutations in the L3 loop, 11 of 68 (16%) in the L1 loop-sheet-\u3b1 helix motif, and 39 of 68 (58%) outside L3 and loop-sheet-\u3b1 helix. Seventy-five percent of the cases (120 of 160) showed DNA aneuploidy, whereas 18% of these (22 of 120) were multiclonal. The major independent predictors for both disease relapse and death were advanced Dukes' stage, p53 mutations affecting L3 loop, DNA-aneuploid tumors, and high SPF (>18.5%). Our results show that mutations in L3 functional domain, more than any mutations, are important biological indicators to predict the outcome of patients indicating that these mutations have biological relevance in terms of colorectal cancer disease course

Effects of TMS on Different Stages of Motor and Non-Motor Verb Processing in the Primary Motor Cortex

The embodied cognition hypothesis suggests that motor and premotor areas are automatically and necessarily involved in understanding action language, as word conceptual representations are embodied. This transcranial magnetic stimulation (TMS) study explores the role of the left primary motor cortex in action-verb processing. TMS-induced motor-evoked potentials from right-hand muscles were recorded as a measure of M1 activity, while participants were asked either to judge explicitly whether a verb was action-related (semantic task) or to decide on the number of syllables in a verb (syllabic task). TMS was applied in three different experiments at 170, 350 and 500 ms post-stimulus during both tasks to identify when the enhancement of M1 activity occurred during word processing. The delays between stimulus onset and magnetic stimulation were consistent with electrophysiological studies, suggesting that word recognition can be differentiated into early (within 200 ms) and late (within 400 ms) lexical-semantic stages, and post-conceptual stages. Reaction times and accuracy were recorded to measure the extent to which the participants' linguistic performance was affected by the interference of TMS with M1 activity. No enhancement of M1 activity specific for action verbs was found at 170 and 350 ms post-stimulus, when lexical-semantic processes are presumed to occur (Experiments 1–2). When TMS was applied at 500 ms post-stimulus (Experiment 3), processing action verbs, compared with non-action verbs, increased the M1-activity in the semantic task and decreased it in the syllabic task. This effect was specific for hand-action verbs and was not observed for action-verbs related to other body parts. Neither accuracy nor RTs were affected by TMS. These findings suggest that the lexical-semantic processing of action verbs does not automatically activate the M1. This area seems to be rather involved in post-conceptual processing that follows the retrieval of motor representations, its activity being modulated (facilitated or inhibited), in a top-down manner, by the specific demand of the task

Detection and quantification of mammaglobin in the blood of breast cancer patients: can it be useful as a potential clinical marker? Preliminary results of a GOIM (Gruppo Oncologico dell'Italia Meridionale) prospective study.

BACKGROUND: Mammaglobin is expressed mainly in mammary tissue, overexpressed in breast cancer (BC) and rarely in other tissue. The aim of this study was to assess the sensitivity and specificity of transcript MGB1 detection and to evaluate the role of MGB1 as potential clinical marker for the detection of disseminated cancer cells in the blood of BC patients. PATIENTS AND METHODS: A consecutive series of 23 BC tissues, 36 peripheral blood BC samples and 35 healthy peripheral blood samples was prospectively recruited to investigate MGB1 expression by means of a quantitative Real Time RT-PCR assay. RESULTS: MGB1 overexpression in tissue samples of BC patients is significantly associated only with high level of Ki67 (P <0.05). None of the samples from peripheral blood of 35 healthy female individuals were positive for MGB1 transcript. In contrast MGB1 mRNA expression was detected in three of 36 (8%) peripheral blood of BC patients. CONCLUSIONS: Our preliminary results demonstrate that the detection of MGB1 transcript in peripheral blood of BC patients was specific but with low sensitivity. MGB1 overexpression by itself or in combination with Ki67 might be considered an index of BC progression

Emergence of qualia from brain activity or from an interaction of proto-consciousness with the brain: which one is the weirder? Available evidence and a research agenda

This contribution to the science of consciousness aims at comparing how two different theories can explain the emergence of different qualia experiences, meta-awareness, meta-cognition, the placebo effect, out-of-body experiences, cognitive therapy and meditation-induced brain changes, etc. The first theory postulates that qualia experiences derive from specific neural patterns, the second one, that qualia experiences derive from the interaction of a proto-consciousness with the brain\u2019s neural activity. From this comparison it will be possible to judge which one seems to better explain the different qualia experiences and to offer a more promising research agenda