Fluorescence Sensing Using DNA Aptamers in Cancer Research and Clinical Diagnostics

Among the various advantages of aptamers over antibodies, remarkable is their ability to tolerate a large number of chemical modifications within their backbone or at the termini without losing significant activity. Indeed, aptamers can be easily equipped with a wide variety of reporter groups or coupled to different carriers, nanoparticles, or other biomolecules, thus producing valuable molecular recognition tools effective for diagnostic and therapeutic purposes. This review reports an updated overview on fluorescent DNA aptamers, designed to recognize significant cancer biomarkers both in soluble or membrane-bound form. In many examples, the aptamer secondary structure switches induced by target recognition are suitably translated in a detectable fluorescent signal using either fluorescently-labelled or label-free aptamers. The fluorescence emission changes, producing an enhancement ("signal-on") or a quenching ("signal-off") effect, directly reflect the extent of the binding, thereby allowing for quantitative determination of the target in bioanalytical assays. Furthermore, several aptamers conjugated to fluorescent probes proved to be effective for applications in tumour diagnosis and intraoperative surgery, producing tumour-type specific, non-invasive in vivo imaging tools for cancer pre- and post-treatment assessment

Double-ended Pseudocomedones in Hidradenitis Suppurativa: Clinical, Dermoscopic, and Histopatho­logical Correlation

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Trifunctionalized Naphthalene Diimides and Dimeric Analogues as G-Quadruplex-Targeting Anticancer Agents Selected by Affinity Chromatography

A focused library of newly designed monomeric and dimeric naphthalene diimides (NDIs) was analyzed in its ability to recognize specific G-quadruplex (G4) structures discriminating duplex DNA. The best G4 ligands—according to an affinity chromatography-based screening method named G4-CPG—were tested on human cancer and healthy cells, inducing DNA damage at telomeres, and in parallel, showing selective antiproliferative activity on HeLa cancer cells with IC50 values in the low nanomolar range. CD and fluorescence spectroscopy studies allowed detailed investigation of the interaction in solution with different G4 and duplex DNA models of the most promising NDI of the series, as determined by combining the biophysical and biological assays’ data

The Effects of Exercise and Kinesio Tape on Physical Limitations in Patients with Knee Osteoarthritis

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Blood film examination for vacuolated and PASpositive lymphocytes as diagnostic screening test for patients with late onset Pompe disease (LOPD)

Pompe disease is an inherited metabolic multisystemic disorder resulting in glycogen storage in different tissues, caused by a deficiency of the lysosomal enzyme acid alfa-glucosidase. Glycogen storage is often a morphological marker in muscle biopsy of Pompe patients but it could be also present in other tissues. Abnormal cytoplasmic vacuolation of lymphocytes, detectable on routine blood film examination, has been recently proposed as possible screening in these patients. We examined blood smear of 16 LOPD patients, aged 14-71 years. The cohort phenotype encompasses 3 patients with presymptomatic hyperckemia, 2 with myalgia and faticability, and 11 with proximal muscle weakness. Among those, 6 patients are on ERT treatment. We collected also peripheral blood films from 20 healthy controls and from 12 patients affected by other muscle glycogenoses. Blood sample was followed by preparation of four blood films: two of them were stained by May-Grunwald/ Giemsa (MGG) end the other two by PAS. . To investigate the diagnostic value of the test, we quantified the percentage of vacuolated lymphocytes and the percentage of PAS-positive lymphocytes. The mean values of PAS-positive lymphocytes in LOPD patients (25.6%) were significantly higher than those of healthy controls (4.6%) and of patients with other muscle glycogenoses (4.8%). In this group of patients, we have shown that PAS-positive cytoplasmic vacuolation of lymphocytes in peripheral blood films could be considered as a reliable screening tool to support an early diagnosis of Pompe disease

Risk stratification in hypertrophic cardiomyopathy. Insights from genetic analysis and cardiopulmonary exercise testing

The role of genetic testing over the clinical and functional variables, including data from the cardiopulmonary exercise test (CPET), in the hypertrophic cardiomyopathy (HCM) risk stratification remains unclear. A retrospective genotype–phenotype correlation was performed to analyze possible differences between patients with and without likely pathogenic/pathogenic (LP/P) variants. A total of 371 HCM patients were screened at least for the main sarcomeric genes MYBPC3 (myosin binding protein C), MYH7 (β-myosin heavy chain), TNNI3 (cardiac troponin I) and TNNT2 (cardiac troponin T): 203 patients had at least an LP/P variant, 23 patients had a unique variant of uncertain significance (VUS) and 145 did not show any LP/P variant or VUS. During a median 5.4 years follow-up, 51 and 14 patients developed heart failure (HF) and sudden cardiac death (SCD) or SCD-equivalents events, respectively. The LP/P variant was associated with a more aggressive HCM phenotype. However, left atrial diameter (LAd), circulatory power (peak oxygen uptake*peak systolic blood pressure, CP%) and ventilatory efficiency (C-index = 0.839) were the only independent predictors of HF whereas only LAd and CP% were predictors of the SCD end-point (C-index = 0.738). The present study reaffirms the pivotal role of the clinical variables and, particularly of those CPET-derived, in the HCM risk stratification.publishedVersio

Next Generation Molecular Diagnosis of Hereditary Spastic Paraplegias: An Italian Cross-Sectional Study

Hereditary spastic paraplegia (HSP) refers to a group of genetically heterogeneous neurodegenerative motor neuron disorders characterized by progressive age-dependent loss of corticospinal motor tract function, lower limb spasticity, and weakness. Recent clinical use of next generation sequencing (NGS) methodologies suggests that they facilitate the diagnostic approach to HSP, but the power of NGS as a first-tier diagnostic procedure is unclear. The larger-than-expected genetic heterogeneity-there are over 80 potential disease-associated genes-and frequent overlap with other clinical conditions affecting the motor system make a molecular diagnosis in HSP cumbersome and time consuming. In a single-center, cross-sectional study, spanning 4 years, 239 subjects with a clinical diagnosis of HSP underwent molecular screening of a large set of genes, using two different customized NGS panels. The latest version of our targeted sequencing panel (SpastiSure3.0) comprises 118 genes known to be associated with HSP. Using an in-house validated bioinformatics pipeline and several in silico tools to predict mutation pathogenicity, we obtained a positive diagnostic yield of 29% (70/239), whereas variants of unknown significance (VUS) were found in 86 patients (36%), and 83 cases remained unsolved. This study is among the largest screenings of consecutive HSP index cases enrolled in real-life clinical-diagnostic settings. Its results corroborate NGS as a modern, first-step procedure for molecular diagnosis of HSP. It also disclosed a significant number of new mutations in ultra-rare genes, expanding the clinical spectrum, and genetic landscape of HSP, at least in Italy

Candidate biomarkers from the integration of methylation and gene expression in discordant autistic sibling pairs

While the genetics of autism spectrum disorders (ASD) has been intensively studied, resulting in the identification of over 100 putative risk genes, the epigenetics of ASD has received less attention, and results have been inconsistent across studies. We aimed to investigate the contribution of DNA methylation (DNAm) to the risk of ASD and identify candidate biomarkers arising from the interaction of epigenetic mechanisms with genotype, gene expression, and cellular proportions. We performed DNAm differential analysis using whole blood samples from 75 discordant sibling pairs of the Italian Autism Network collection and estimated their cellular composition. We studied the correlation between DNAm and gene expression accounting for the potential effects of different genotypes on DNAm. We showed that the proportion of NK cells was significantly reduced in ASD siblings suggesting an imbalance in their immune system. We identified differentially methylated regions (DMRs) involved in neurogenesis and synaptic organization. Among candidate loci for ASD, we detected a DMR mapping to CLEC11A (neighboring SHANK1) where DNAm and gene expression were significantly and negatively correlated, independently from genotype effects. As reported in previous studies, we confirmed the involvement of immune functions in the pathophysiology of ASD. Notwithstanding the complexity of the disorder, suitable biomarkers such as CLEC11A and its neighbor SHANK1 can be discovered using integrative analyses even with peripheral tissues

How future surgery will benefit from SARS-COV-2-related measures: a SPIGC survey conveying the perspective of Italian surgeons

COVID-19 negatively affected surgical activity, but the potential benefits resulting from adopted measures remain unclear. The aim of this study was to evaluate the change in surgical activity and potential benefit from COVID-19 measures in perspective of Italian surgeons on behalf of SPIGC. A nationwide online survey on surgical practice before, during, and after COVID-19 pandemic was conducted in March-April 2022 (NCT:05323851). Effects of COVID-19 hospital-related measures on surgical patients' management and personal professional development across surgical specialties were explored. Data on demographics, pre-operative/peri-operative/post-operative management, and professional development were collected. Outcomes were matched with the corresponding volume. Four hundred and seventy-three respondents were included in final analysis across 14 surgical specialties. Since SARS-CoV-2 pandemic, application of telematic consultations (4.1% vs. 21.6%; p < 0.0001) and diagnostic evaluations (16.4% vs. 42.2%; p < 0.0001) increased. Elective surgical activities significantly reduced and surgeons opted more frequently for conservative management with a possible indication for elective (26.3% vs. 35.7%; p < 0.0001) or urgent (20.4% vs. 38.5%; p < 0.0001) surgery. All new COVID-related measures are perceived to be maintained in the future. Surgeons' personal education online increased from 12.6% (pre-COVID) to 86.6% (post-COVID; p < 0.0001). Online educational activities are considered a beneficial effect from COVID pandemic (56.4%). COVID-19 had a great impact on surgical specialties, with significant reduction of operation volume. However, some forced changes turned out to be benefits. Isolation measures pushed the use of telemedicine and telemetric devices for outpatient practice and favored communication for educational purposes and surgeon-patient/family communication. From the Italian surgeons' perspective, COVID-related measures will continue to influence future surgical clinical practice