Erythrocyte Plasma Membrane Lipid Composition Mirrors That of Neurons and Glial Cells in Murine Experimental In Vitro and In Vivo Inflammation.

Lipid membrane turnover and myelin repair play a central role in diseases and lesions of the central nervous system (CNS). The aim of the present study was to analyze lipid composition changes due to inflammatory conditions. We measured the fatty acid (FA) composition in erythrocytes (RBCs) and spinal cord tissue (gas chromatography) derived from mice affected by experimental allergic encephalomyelitis (EAE) in acute and remission phases; cholesterol membrane content (Filipin) and GM1 membrane assembly (CT-B) in EAE mouse RBCs, and in cultured neurons, oligodendroglial cells and macrophages exposed to inflammatory challenges. During the EAE acute phase, the RBC membrane showed a reduction in polyunsaturated FAs (PUFAs) and an increase in saturated FAs (SFAs) and the omega-6/omega-3 ratios, followed by a restoration to control levels in the remission phase in parallel with an increase in monounsaturated fatty acid residues. A decrease in PUFAs was also shown in the spinal cord. CT-B staining decreased and Filipin staining increased in RBCs during acute EAE, as well as in cultured macrophages, neurons and oligodendrocyte precursor cells exposed to inflammatory challenges. This regulation in lipid content suggests an increased cell membrane rigidity during the inflammatory phase of EAE and supports the investigation of peripheral cell membrane lipids as possible biomarkers for CNS lipid membrane concentration and assembly.

Optical tissue clearing associated with 3D imaging: application in preclinical and clinical studies.

Understanding the inner morphology of intact tissues is one of the most competitive challenges in modern biology. Since the beginning of the twentieth century, optical tissue clearing (OTC) has provided solutions for volumetric imaging, allowing the microscopic visualization of thick sections of tissue, organoids, up to whole organs and organisms (for example, mouse or rat). Recently, tissue clearing has also been introduced in clinical settings to achieve a more accurate diagnosis with the support of 3D imaging. This review aims to give an overview of the most recent developments in OTC and 3D imaging and to illustrate their role in the field of medical diagnosis, with a specific focus on clinical applications.

MicroRNAs Patterns as Potential Tools for Diagnostic and Prognostic Follow-Up in Cancer Survivorship.

Advances in screening methods and pharmacological treatments are increasing the life expectancy of cancer patients. During recent decades, the community of long-term disease-free cancer survivors (LCS) has grown exponentially, raising the issues related to cancer follow-up. Cancer relapse and other cancer-related diseases, as well as lifestyle, influence cancer survival. Recently, the regulatory role of microRNAs (miRNAs) in gene expression and their involvement in human diseases, including cancer, has been identified. Extracellular circulating miRNAs (ECmiRNAs) have been found in biological fluids and specific ECmiRNAs have been associated with cancer development and progression or with a therapy response. Here, we focus on the pivotal role of ECmiRNAs as biomarkers in cancer diagnosis and prognosis. Then, we discuss the relevance of ECmiRNAs expression in cancer survivors for the identification of specific ECmiRNAs profiles as potential tools to assess cancer outcome and to control LCS follow-up.

The Role of Extracellular Vesicles as Shuttles of RNA and Their Clinical Significance as Biomarkers in Hepatocellular Carcinoma.

Extracellular vesicles (EVs) have attracted interest as mediators of intercellular communication following the discovery that EVs contain RNA molecules, including non-coding RNA (ncRNA). Growing evidence for the enrichment of peculiar RNA species in specific EV subtypes has been demonstrated. ncRNAs, transferred from donor cells to recipient cells, confer to EVs the feature to regulate the expression of genes involved in differentiation, proliferation, apoptosis, and other biological processes. These multiple actions require accuracy in the isolation of RNA content from EVs and the methodologies used play a relevant role. In liver, EVs play a crucial role in regulating cell-cell communications and several pathophysiological events in the heterogeneous liver class of cells via horizontal transfer of their cargo. This review aims to discuss the rising role of EVs and their ncRNAs content in regulating specific aspects of hepatocellular carcinoma development, including tumorigenesis, angiogenesis, and tumor metastasis. We analyze the progress in EV-ncRNAs' potential clinical applications as important diagnostic and prognostic biomarkers for liver conditions.

Legislation to limit the environmental plastic and microplastic pollution and their influence on human exposure.

Plastic pollution is an emerging problem and is a consequence of the post-consumer plastic waste accumulation in the environment coupled to mismanaged waste programmes. Countries are counteracting the continuous growth of plastic litter with different strategies: introducing bans and limits on both plastic items and materials, promoting plastic recycling and recovery strategies and encouraging voluntary clean up actions, as well as raising public awareness. However, the toxicity of plastics to the environment and organisms is not only related to their polymer chains, but also to the fact that plastic materials contain hazardous additives and can adsorb environmental pollutants (i.e. heavy metals and persistent organic contaminants, respectively). The plastic/additives/pollutants combination may be ingested by marine organisms and then enter in the food chain. Therefore, legislation for additives and contaminants is crucial both to reduce environmental pollution and their toxic effects on organisms, which of course includes humans. In this review, the current policies on plastics and related contaminants are described focusing on current laws. Moreover, recommendations for seafood consumption are suggested, since each fish or mollusc eaten may potentially result in plastic particles, additives or contaminants ingestion.

The Complexity of the Tumor Microenvironment and Its Role in Acute Lymphoblastic Leukemia: Implications for Therapies.

The microenvironment that surrounds a tumor, in addition to the tumor itself, plays an important role in the onset of resistance to molecularly targeted therapies. Cancer cells and their microenvironment interact closely between them by means of a molecular communication that mutually influences their biological characteristics and behavior. Leukemia cells regulate the recruitment, activation and program of the cells of the surrounding microenvironment, including those of the immune system. Studies on the interactions between the bone marrow (BM) microenvironment and Acute Lymphoblastic Leukemia (ALL) cells have opened a scenario of potential therapeutic targets which include cytokines and their receptors, signal transduction networks, and hypoxia-related proteins. Hypoxia also enhances the formation of new blood vessels, and several studies show how angiogenesis could have a key role in the pathogenesis of ALL. Knowledge of the molecular mechanisms underlying tumor-microenvironment communication and angiogenesis could contribute to the early diagnosis of leukemia and to personalized molecular therapies. This article is part of a Special Issue entitled: Innovative Multi-Disciplinary Approaches for Precision Studies in Leukemia edited by Sandra Marmiroli (University of Modena and Reggio Emilia, Modena, Italy) and Xu Huang (University of Glasgow, Glasgow, United Kingdom).

Two neuroendocrine G protein-coupled receptor molecules, somatostatin and melatonin: Physiology of signal transduction and therapeutic perspectives.

Recent studies have shown that G protein-coupled receptors (GPCRs), the largest signal-conveying receptor family, are targets for mutations occurring frequently in different cancer types. GPCR alterations associated with cancer development represent significant challenges for the discovery and the advancement of targeted therapeutics. Among the different molecules that can activate GPCRs, we focused on two molecules that exert their biological actions regulating many typical features of tumorigenesis such as cellular proliferation, survival, and invasion: somatostatin and melatonin. The modulation of signaling pathways, that involves these two molecules, opens an interesting scenario for cancer therapy, with the opportunity to act at different molecular levels. Therefore, the aim of this review is the analysis of the biological activity and the therapeutic potential of somatostatin and melatonin, displaying a high affinity for GPCRs, that interfere with cancer development and maintenance.

Application of ethyl cinnamate based optical tissue clearing and expansion microscopy combined with retrograde perfusion for 3D lung imaging.

PURPOSE: 3 D imaging of the lung is not a trivial undertaking as during preparation the lung may collapse. Also serial sections and scans followed by 3 D reconstruction may lead to artifacts. The present study aims to figure out the best way to perform 3 D imaging in lung research. MATERIALS AND METHODS: We applied an optical tissue clearing (OTC) method, which uses ethyl cinnamate (ECi) as a fast, nontoxic and cheap clearing solvent, for 3 D imaging of retrograde perfused lungs by laser confocal fluorescence microscopy and light sheet fluorescence microscopy. We also introduced expansion microscopy (ExM), a cutting-edge technique, in 3 D imaging of lungs. We examined and compared the usefulness of these techniques for 3 D lung imaging. The ExM protocol was further extended to paraffin-embedded lung metastases blocks. RESULTS: The MHI148-PEI labeled lung vasculature was visualized by retrograde perfusion combined with trachea ligation and ECi based OTC. As compared with trans-cardiac perfusion, the retrograde perfusion results in a better maintenance of lung morphology. 3 D structure of alveoli, vascular branches and cilia in lung were revealed by immunofluorescence staining after ExM. 3 D distribution of microvasculature and neutrophil cells in 10 years old paraffin-embedded lung metastases were analyzed by ExM. CONCLUSIONS: The retrograde perfusion combined with trachea ligation technique could be applied in the lung research in mice. 3 D structure of lung vasculature can be visualized by MHI148-PEI perfusion and ECi based OTC in an efficient way. ExM and immunofluorescence staining protocol is highly recommended to perform 3 D imaging of fresh fixed lung as well as paraffin-embedded lung blocks.

New technical approaches for 3D morphological imaging and quantification of measurements.

3D imaging is becoming more and more popular, as it allows us to identify interactions between structures in organs. Furthermore, it gives the possibility to quantify and size these structures. To allow 3D imaging, the tissue sample has to be transparent. This is usually achieved by using optical tissue clearing protocols. Although using optical tissue clearing often results in perfect 3D images, these protocols have some pitfalls, like long duration of sample preparation (up to several weeks), use of toxic substances, damage to antibody staining, fluorescent proteins or dyes, high refractive indices, and high costs of sample processing.Recently we described [Huang et al., Scientific Reports 9(1): 521 (2019)] a fast, safe, and inexpensive ethyl cinnamate (ECi) based optical tissue clearing protocol. Here, we present extensions of our protocol with respect to the deparaffinization of old paraffin-embedded samples allowing 3D imaging of the blocks. In addition, we learned to remove ECi from the samples allowing the use of routine immunolabeling protocols. Furthermore, we demonstrate new pictures of lungs after expansion microscopy and adaptation of already existing protocols. The aim of our work is, in summary, to describe the advances in these methodologies, focusing on the morphological imaging of kidneys and lungs.

A cationic near infrared fluorescent agent and ethyl-cinnamate tissue clearing protocol for vascular staining and imaging.

Understanding vascular structures and dysfunction is a fundamental challenge. This task has been approached by using traditional methodologies such as microscopic computed tomography and magnetic resonance imaging. Both techniques are not only expensive but also time-consuming. Here, we present a new method for visualizing vascular structures in different organs in an efficient manner. A cationic near infrared (NIR) fluorescent dye was developed with attractive features to specifically stain blood vessels. Furthermore, we refined the process of organ staining and harvesting by retrograde perfusion and optimized the subsequent dehydration and clearing process by the use of an automatic tissue processor and a non-toxic substance, ethyl-cinnamate. Using this approach, the time interval between organ harvesting and microscopic analysis can be reduced from day(s) or weeks to 4 hours. Finally, we have demonstrated that the new NIR fluorescent agent in combination with confocal or light-sheet microscopy can be efficiently used for visualization of vascular structures, such as the blood vessels in different organs e.g. glomeruli in kidneys, with an extremely high resolution. Our approach facilitates the development of automatic image processing and the quantitative analysis to study vascular and kidney diseases.

Stem/Stromal Cells for Treatment of Kidney Injuries With Focus on Preclinical Models.

Within the last years, the use of stem cells (embryonic, induced pluripotent stem cells, or hematopoietic stem cells), Progenitor cells (e.g., endothelial progenitor cells), and most intensely mesenchymal stromal cells (MSC) has emerged as a promising cell-based therapy for several diseases including nephropathy. For patients with end-stage renal disease (ESRD), dialysis or finally organ transplantation are the only therapeutic modalities available. Since ESRD is associated with a high healthcare expenditure, MSC therapy represents an innovative approach. In a variety of preclinical and clinical studies, MSC have shown to exert renoprotective properties, mediated mainly by paracrine effects, immunomodulation, regulation of inflammation, secretion of several trophic factors, and possibly differentiation to renal precursors. However, studies are highly diverse; thus, knowledge is still limited regarding the exact mode of action, source of MSC in comparison to other stem cell types, administration route and dose, tracking of cells and documentation of therapeutic efficacy by new imaging techniques and tissue visualization. The aim of this review is to provide a summary of published studies of stem cell therapy in acute and chronic kidney injury, diabetic nephropathy, polycystic kidney disease, and kidney transplantation. Preclinical studies with allogeneic or xenogeneic cell therapy were first addressed, followed by a summary of clinical trials carried out with autologous or allogeneic hMSC. Studies were analyzed with respect to source of cell type, mechanism of action etc.