A Teledentistry System for the Second Opinion

In this paper we present a teledentistry system aimed to the Second Opinion task. It make use of a particular camera called intra-oral camera, also called dental camera, in order to perform the photo shooting and real-time video of the inner part of the mouth. The pictures acquired by the Operator with such a device are sent to the Oral Medicine Expert (OME) by means of a current File Transfer Protocol (FTP) service and the real-time video is channeled into a video streaming thanks to the VideoLan client/server (VLC) application. It is composed by a HTML5 web-pages generated by PHP and allows to perform the Second Opinion both when Operator and OME are logged and when one of them is offline

Is BRCA1-5083del19, identified in breast cancer patients of Sicilian origin, a Calabrian founder mutation?

Various studies have been published in Italy regarding the different BRCA1 mutations, but only the BRCA1-5083del19 mutation is recurrent and specific to individuals of Italian descent with a founder effect on the Calabrian population. In our previous study, BRCA1-5083del19 mutation carriers were found in four index cases of 106 Sicilian patients selected for familial and/or hereditary breast/ovarian cancers. The high frequency rate of this mutation identified in the Sicilian population led us to perform haplotype analysis in all family carriers. Five highly polymorphic microsatellite markers were used (D17S1320, D17S932, D17S1323, D17S1326, D17S1325) to establish whether or not all these families had a common ancestor. This analysis showed that all mutation carriers of these families had a common allele. None of the non-carriers of the mutation or of the 50 healthy Sicilian controls showed this haplotype. This allelotype analysis highlighted the presence of a common allele (ancestor), thus suggesting the presence of a founder effect in the Sicilian population. Our results are in contrast with other studies but only the allelotype analysis of all the BRCA1-5083del19 mutation carriers of two neighboring regions of the south of Italy (Calabria and Sicily) will make it possible to identify the real ancestor of this mutation

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

Tackling amyloidogenesis in Alzheimer's disease with A2V variants of Amyloid-β

We developed a novel therapeutic strategy for Alzheimer’s disease (AD) exploiting the properties of a natural variant of Amyloid-β (Aβ) carrying the A2V substitution, which protects heterozygous carriers from AD by its ability to interact with wild-type Aβ, hindering conformational changes and assembly thereof. As prototypic compound we designed a six-mer mutated peptide (Aβ1-6A2V), linked to the HIV-related TAT protein, which is widely used for brain delivery and cell membrane penetration of drugs. The resulting molecule [Aβ1-6A2VTAT(D)] revealed strong anti-amyloidogenic effects in vitro and protected human neuroblastoma cells from Aβ toxicity. Preclinical studies in AD mouse models showed that short-term treatment with Aβ1-6A2VTAT(D) inhibits Aβ aggregation and cerebral amyloid deposition, but a long treatment schedule unexpectedly increases amyloid burden, although preventing cognitive deterioration. Our data support the view that the AβA2V-based strategy can be successfully used for the development of treatments for AD, as suggested by the natural protection against the disease in human A2V heterozygous carriers. The undesirable outcome of the prolonged treatment with Aβ1-6A2VTAT(D) was likely due to the TAT intrinsic attitude to increase Aβ production, avidly bind amyloid and boost its seeding activity, warning against the use of the TAT carrier in the design of AD therapeutics

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

Targeting senescence as a novel pharmacological approach in Lymphangioleiomyomatosis

Orphanet Journal of Rare Diseases 2024, 18(1): P11 Background: Lymphangioleiomyomatosis (LAM) is a multisystem ultra-rare disease that affects women[1]. LAM cells, characterized by the constitutive activation of mTOR (mechanistic Target of Rapamycin) due to the impaired expression of its regulatory proteins hamartin or tuberin, invade the lung, leading to the loss of pulmonary function [2]. Rapamycin is the only drug approved for LAM, but new therapies are needed, since LAM relapses to rapamycin discontinuation[3]. mTOR is a driver of senescence[4], a stress-response process characterized by the inhibition of the cell cycle and by the acquisition of a senescence-associated secretory phenotype (SASP) [5]. Through SASP, senescent cells reinforce their phenotype and promote senescence in neighbouring cells[6]. However, dysregulation of senescence might contribute to disease onset, as SASP boost a proinflammatory microenvironment and the cell cycle arrest limits tissue regeneration[7]. We recently demonstrated that primary tuberin-deficient LAM/TSC cells have senescent features depending on mTOR hyperactivation and, in an in vitro model of LAM microenvironment, they have the capability to induce senescence in tuberin-expressing primary lung fibroblasts (PLFs) through their conditioned medium (CM), with increased secretion of Interleukin(IL)-8[8]. Interestingly, the lysine acetyl transferase p300 was recently identified as epigenetic driver of cellular senescence and it is known to interact with mTOR[9, 10]. Materials and methods: Changing in senescent features, as the increased β-galactosidase positivity and p21 expression, were measured in PLFs grown in the presence of IL-8 and in LAM/TSC cells treated with a monoclonal antibody or with the small molecule SB225002 targeting the IL-8 receptor CXCR2. Then SB225002 was added to PLFs in LAM/TSC CM to counteract senescence induction. In addition, phosphorylation of H2A.X histone and cleaved Caspase 3 were assessed in senescent induced lymphoblastoid cell line (SI-LCLs) derived from controls (HD) and individuals with p300 haploinsufficiency due to germline pathogenetic variants (EP300+/). Results: The addition of IL-8 in PLFs CM increased their senescence proportionally to its concentration. Moreover, blocking CXCR2 impaired the capability of LAM/TSC CM to induce senescence in PLFs. At the same time, targeting CXCR2 hampered senescence in LAM/TSC cells. Finally, the impaired phosphorylation of H2A.X in HD SI-LCLs compared to EP300+/- LCLs confirmed the role of p300 in senescence, suggesting possible avenues for targeting its activity in LAM. Conclusions: Our results indicate that the modulation of senescence targeting mTOR downstream effectors and non-mTOR regulated mechanisms is an intriguing novel pharmacological approach to interfere with the pathological communication in LAM microenvironment with possible future application in LAM therapy