Diagnostic and prognostic role of liquid biopsy in non-small cell lung cancer: evaluation of circulating biomarkers

Lung cancer is still one of the main causes of cancer-related death, together with prostate and colorectal cancers in males and breast and colorectal cancers in females. The prognosis for non-small cell lung cancer (NSCLC) is strictly dependent on feasibility of a complete surgical resection of the tumor at diagnosis. Since surgery is indicated only in early stages tumors, it is necessary to anticipate the timing of diagnosis in clinical practice. In the diagnostic and therapeutic pathway for NSCLC, sampling of neoplastic tissue is usually obtained using invasive methods that are not free from disadvantages and complications. A valid alternative to the standard biopsy is the liquid biopsy (LB), that is, the analysis of samples from peripheral blood, urine, and other biological fluids, with a simple and non-invasive collection. In particular, it is possible to detect in the blood different tumor derivatives, such as cell-free DNA (cfDNA) with its subtype circulating tumor DNA (ctDNA), cell-free RNA (cfRNA), and circulating tumor cells (CTCs). Plasma-based testing seems to have several advantages over tumor tissue biopsy; firstly, it reduces medical costs, risk of complications related to invasive procedures, and turnaround times; moreover, the analysis of genes alteration, such as EGFR, ALK, ROS1, and BRAF is faster and safer with this method, compared to tissue biopsy. Despite all these advantages, the evidences in literatures indicate that assays performed on liquid biopsies have a low sensitivity, making them unsuitable for screening in lung cancer at the current state. This is caused by lack of standardization in sampling and preparation of specimen and by the low concentration of biomarkers in the bloodstream. Instead, routinely use of LB should be preferred in revaluation of patients with advanced NSCLC resistant to chemotherapy, due to onset of new mutations

Interpretation of the optical transfer function: Significance for image scanning microscopy

The optical transfer function (OTF) is widely used to compare the performance of different optical systems. Conventionally, the OTF is normalized to unity for zero spatial frequency, but in some cases it is better to consider the unnormalized OTF, which gives the absolute value of the image signal. Examples are in confocal microscopy and image scanning microscopy, where the signal level increases with pinhole or array size. Comparison of the respective unnormalized OTFs gives useful insight into their relative performance. The significance of other properties of the general OTF is discussed

On the Advent of Super-Resolution Microscopy in the Realm of Polycomb Proteins

Simple Summary The genomes of metazoans are organized at multiple spatial scales, ranging from the double helix of DNA to whole chromosomes. The intermediate genomic scale of kilobases to megabases, which corresponds to the 50-300 nm spatial scale, is particularly interesting because the tridimensional arrangement of chromatin is implicated in multiple regulatory mechanisms. Indeed, a crucial hallmark of cellular life is the widespread ordering of many biological processes in nano-/mesoscopic domains (10-200 nm), which now may be revealed by an imaging toolbox referred to as super-resolution microscopy. In this context, polycomb proteins stand as major epigenetic modulators of chromatin function, acting prevalently as repressors of gene transcription. This work reviews the current state-of-the-art super-resolution microscopy applied to polycomb proteins. Of note, super-resolution data have complemented cutting-edge molecular biology methods in providing a rational framework for understanding how polycomb proteins may shape 3D chromatin topologies and functions. The genomes of metazoans are organized at multiple spatial scales, ranging from the double helix of DNA to whole chromosomes. The intermediate genomic scale of kilobases to megabases, which corresponds to the 50-300 nm spatial scale, is particularly interesting, as the 3D arrangement of chromatin is implicated in multiple regulatory mechanisms. In this context, polycomb group (PcG) proteins stand as major epigenetic modulators of chromatin function, acting prevalently as repressors of gene transcription by combining chemical modifications of target histones with physical crosslinking of distal genomic regions and phase separation. The recent development of super-resolution microscopy (SRM) has strongly contributed to improving our comprehension of several aspects of nano-/mesoscale (10-200 nm) chromatin domains. Here, we review the current state-of-the-art SRM applied to PcG proteins, showing that the application of SRM to PcG activity and organization is still quite limited and mainly focused on the 3D assembly of PcG-controlled genomic loci. In this context, SRM approaches have mostly been applied to multilabel fluorescence in situ hybridization (FISH). However, SRM data have complemented the maps obtained from chromosome capture experiments and have opened a new window to observe how 3D chromatin topology is modulated by PcGs

Muscular cystic hydatidosis: case report

BACKGROUND: Hydatidosis is a zoonosis caused by Echinococcus granulosus, and ingesting eggs released through the faeces from infected dogs infects humans. The location of the hydatid cysts is mostly hepatic and/or pulmonary, whereas musculoskeletal hydatidosis is very rare. CASE PRESENTATION: We report an unusual case of primary muscular hydatidosis in proximity of the big adductor in a young Sicilian man. The patient, 34 years old, was admitted to the Department of Infectious and Tropical Diseases for ultrasonographic detection, with successive confirmation by magnetic resonance imaging, of an ovular mass (13 × 8 cm) in the big adductor of the left thigh, cyst-like, and containing several small cystic formations. Serological tests for hydatidosis gave negative results. A second drawing of blood was done 10 days after the first one and showed an increase in the antibody titer for hydatidosis. The patient was submitted to surgical excision of the lesion with perioperatory prophylaxis with albendazole. The histopathological examination of the bioptic material was not diriment in the diagnosis, therefore further tests were performed: additional serological tests for hydatidosis for the evaluation of IgE and IgG serotype (Western Blot and REAST), and molecular analysis of the excised material. These more specific serological tests gave positive results for hydatidosis, and the sequencing of the polymerase chain reaction products from the cyst evidenced E. granulosus DNA, genotype G1. Any post-surgery complications was observed during 6 following months. CONCLUSION: Cystic hydatidosis should always be considered in the differential diagnosis of any cystic mass, regardless of its location, also in epidemiological contests less suggestive of the disease. The diagnosis should be achieved by taking into consideration the clinical aspects, the epidemiology of the disease, the imaging and immunological tests but, as demonstrated in this case, without neglecting the numerous possibilities offered by new serological devices and modern day molecular biology techniques

Intensity Weighted Subtraction Microscopy Approach for Image Contrast and Resolution Enhancement

We propose and demonstrate a novel subtraction microscopy algorithm, exploiting fluorescence emission difference or switching laser mode and their derivatives for image enhancement. The key novelty of the proposed approach lies in the weighted subtraction coefficient, adjusted pixel-by-pixel with respect to the intensity distributions of initial images. This method produces significant resolution enhancement and minimizes image distortions. Our theoretical and experimental studies demonstrate that this approach can be applied to any optical microscopy techniques, including label free and non-linear methods, where common super-resolution techniques cannot be used

Two-photon image-scanning microscopy with SPAD array and blind image reconstruction

Two-photon excitation (2PE) laser scanning microscopy is the imaging modality of choice when one desires to work with thick biological samples. However, its spatial resolution is poor, below confocal laser scanning microscopy. Here, we propose a straightforward implementation of 2PE image scanning microscopy (2PE-ISM) that, by leveraging our recently introduced single-photon avalanche diode (SPAD) array detector and a novel blind image reconstruction method, is shown to enhance the effective resolution, as well as the overall image quality of 2PE microscopy. With our adaptive pixel reassignment procedure similar to 1.6 times resolution increase is maintained deep into thick semi-transparent samples. The integration of Fourier ring correlation based semi-blind deconvolution is shown to further enhance the effective resolution by a factor of similar to 2 - and automatic background correction is shown to boost the image quality especially in noisy images. Most importantly, our 2PE-ISM implementation requires no calibration measurements or other input from the user, which is an important aspect in terms of day-to-day usability of the technique. (C) 2020 Optical Society of America under the terms of the OSA Open Access Publishing Agreemen

High quality RNA isolation from Aedes aegypti midguts using laser microdissection microscopy

Background: Laser microdissection microscopy (LMM) has potential as a research tool because it allows precise excision of target tissues or cells from a complex biological specimen, and facilitates tissue-specific sample preparation. However, this method has not been used in mosquito vectors to date. To this end, we have developed an LMM method to isolate midgut RNA using Aedes aegypti

Global Gene Expression Profiling Of Human Pleural Mesotheliomas: Identification of Matrix Metalloproteinase 14 (MMP-14) as Potential Tumour Target

BACKGROUND:The goal of our study was to molecularly dissect mesothelioma tumour pathways by mean of microarray technologies in order to identify new tumour biomarkers that could be used as early diagnostic markers and possibly as specific molecular therapeutic targets. METHODOLOGY:We performed Affymetrix HGU133A plus 2.0 microarray analysis, containing probes for about 39,000 human transcripts, comparing 9 human pleural mesotheliomas with 4 normal pleural specimens. Stringent statistical feature selection detected a set of differentially expressed genes that have been further evaluated to identify potential biomarkers to be used in early diagnostics. Selected genes were confirmed by RT-PCR. As reported by other mesothelioma profiling studies, most of genes are involved in G2/M transition. Our list contains several genes previously described as prognostic classifier. Furthermore, we found novel genes, never associated before to mesotheliom that could be involved in tumour progression. Notable is the identification of MMP-14, a member of matrix metalloproteinase family. In a cohort of 70 mesothelioma patients, we found by a multivariate Cox regression analysis, that the only parameter influencing overall survival was expression of MMP14. The calculated relative risk of death in MM patients with low MMP14 expression was significantly lower than patients with high MMp14 expression (P = 0.002). CONCLUSIONS:Based on the results provided, this molecule could be viewed as a new and effective therapeutic target to test for the cure of mesothelioma

Genomics in neurodevelopmental disorders: an avenue to personalized medicine

Despite the remarkable number of scientific breakthroughs of the last 100 years, the treatment of neurodevelopmental disorders (e.g., autism spectrum disorder, intellectual disability) remains a great challenge. Recent advancements in genomics, such as whole-exome or whole-genome sequencing, have enabled scientists to identify numerous mutations underlying neurodevelopmental disorders. Given the few hundred risk genes that have been discovered, the etiological variability and the heterogeneous clinical presentation, the need for genotype — along with phenotype- based diagnosis of individual patients has become a requisite. In this review we look at recent advancements in genomic analysis and their translation into clinical practice