Molecular Aspects and Prognostic Significance of Microcalcifications in Human Pathology: A Narrative Review

The presence of calcium deposits in human lesions is largely used as imaging biomarkers of human diseases such as breast cancer. Indeed, the presence of micro- or macrocalcifications is frequently associated with the development of both benign and malignant lesions. Nevertheless, the molecular mechanisms involved in the formation of these calcium deposits, as well as the prognostic significance of their presence in human tissues, have not been completely elucidated. Therefore, a better characterization of the biological process related to the formation of calcifications in different tissues and organs, as well as the understanding of the prognostic significance of the presence of these calcium deposits into human tissues could significantly improve the management of patients characterized by microcalcifications associated lesions. Starting from these considerations, this narrative review highlights the most recent histopathological and molecular data concerning the formation of calcifications in breast, thyroid, lung, and ovarian diseases. Evidence reported here could deeply change the current point of view concerning the role of ectopic calcifications in the progression of human diseases and also in the patients’ management. In fact, the presence of calcifications can suggest an unfavorable prognosis due to dysregulation of normal tissues homeostasis.     

The Chick Chorioallantoic Membrane Model: A New In Vivo Tool to Evaluate Breast Cancer Stem Cell Activity

The high plasticity of cancer stem-like cells (CSCs) allows them to differentiate and proliferate, specifically when xenotransplanted subcutaneously into immunocompromised mice. CSCs are highly tumorigenic, even when inoculated in small numbers. Thus, in vivo limiting dilution assays (LDA) in mice are the current gold standard method to evaluate CSC enrichment and activity. The chick embryo chorioallantoic membrane (CAM) is a low cost, naturally immune-incompetent and reproducible model widely used to evaluate the spontaneous growth of human tumor cells. Here, we established a CAM-LDA assay able to rapidly reproduce tumor specificities—in particular, the ability of the small population of CSCs to form tumors. We used a panel of organotropic metastatic breast cancer cells, which show an enrichment in a stem cell gene signature, enhanced CD44⁺/CD24^−/low cell surface expression and increased mammosphere-forming efficiency (MFE). The size of CAM-xenografted tumors correlate with the number of inoculated cancer cells, following mice xenograft growth pattern. CAM and mice tumors are histologically comparable, displaying both breast CSC markers CD44 and CD49f. Therefore, we propose a new tool for studying CSC prevalence and function—the chick CAM-LDA—a model with easy handling, accessibility, rapid growth and the absence of ethical and regulatory constraints.     

Curcumin Prevents Cerebellar Hypoplasia and Restores the Behavior in Hyperbilirubinemic Gunn Rat by a Pleiotropic Effect on the Molecular Effectors of Brain Damage

Bilirubin toxicity to the central nervous system (CNS) is responsible for severe and permanent neurologic damage, resulting in hearing loss, cognitive, and movement impairment. Timely and effective management of severe neonatal hyperbilirubinemia by phototherapy or exchange transfusion is crucial for avoiding permanent neurological consequences, but these therapies are not always possible, particularly in low-income countries. To explore alternative options, we investigated a pharmaceutical approach focused on protecting the CNS from pigment toxicity, independently from serum bilirubin level. To this goal, we tested the ability of curcumin, a nutraceutical already used with relevant results in animal models as well as in clinics in other diseases, in the Gunn rat, the spontaneous model of neonatal hyperbilirubinemia. Curcumin treatment fully abolished the landmark cerebellar hypoplasia of Gunn rat, restoring the histological features, and reverting the behavioral abnormalities present in the hyperbilirubinemic rat. The protection was mediated by a multi-target action on the main bilirubin-induced pathological mechanism ongoing CNS damage (inflammation, redox imbalance, and glutamate neurotoxicity). If confirmed by independent studies, the result suggests the potential of curcumin as an alternative/complementary approach to bilirubin-induced brain damage in the clinical scenario.     

Target Score—A Proteomics Data Selection Tool Applied to Esophageal Cancer Identifies GLUT1-Sialyl Tn Glycoforms as Biomarkers of Cancer Aggressiveness

Esophageal cancer (EC) is a life-threatening disease, demanding the discovery of new biomarkers and molecular targets for precision oncology. Aberrantly glycosylated proteins hold tremendous potential towards this objective. In the current study, a series of esophageal squamous cell carcinomas (ESCC) and EC-derived circulating tumor cells (CTCs) were screened by immunoassays for the sialyl-Tn (STn) antigen, a glycan rarely expressed in healthy tissues and widely observed in aggressive gastrointestinal cancers. An ESCC cell model was glycoengineered to express STn and characterized in relation to cell proliferation and invasion in vitro. STn was found to be widely present in ESCC (70% of tumors) and in CTCs in 20% of patients, being associated with general recurrence and reduced survival. Furthermore, STn expression in ESCC cells increased invasion in vitro, while reducing cancer cells proliferation. In parallel, an ESCC mass spectrometry-based proteomics dataset, obtained from the PRIDE database, was comprehensively interrogated for abnormally glycosylated proteins. Data integration with the Target Score, an algorithm developed in-house, pinpointed the glucose transporter type 1 (GLUT1) as a biomarker of poor prognosis. GLUT1-STn glycoproteoforms were latter identified in tumor tissues in patients facing worst prognosis. Furthermore, healthy human tissues analysis suggested that STn glycosylation provided cancer specificity to GLUT1. In conclusion, STn is a biomarker of worst prognosis in EC and GLUT1-STn glycoforms may be used to increase its specificity on the stratification and targeting of aggressive ESCC forms.     

Pharmacological Evidence on Augmented Antiallodynia Following Systemic Co-Treatment with GlyT-1 and GlyT-2 Inhibitors in Rat Neuropathic Pain Model

The limited effect of current medications on neuropathic pain (NP) has initiated large efforts to develop effective treatments. Animal studies showed that glycine transporter (GlyT) inhibitors are promising analgesics in NP, though concerns regarding adverse effects were raised. We aimed to study NFPS and Org-25543, GlyT-1 and GlyT-2 inhibitors, respectively and their combination in rat mononeuropathic pain evoked by partial sciatic nerve ligation. Cerebrospinal fluid (CSF) glycine content was also determined by capillary electrophoresis. Subcutaneous (s.c.) 4 mg/kg NFPS or Org-25543 showed analgesia following acute administration (30–60 min). Small doses of each compound failed to produce antiallodynia up to 180 min after the acute administration. However, NFPS (1 mg/kg) produced antiallodynia after four days of treatment. Co-treatment with subanalgesic doses of NFPS (1 mg/kg) and Org-25543 (2 mg/kg) produced analgesia at 60 min and thereafter meanwhile increased significantly the CSF glycine content. This combination alleviated NP without affecting motor function. Test compounds failed to activate G-proteins in spinal cord. To the best of our knowledge for the first time we demonstrated augmented analgesia by combining GlyT-1 and 2 inhibitors. Increased CSF glycine content supports involvement of glycinergic system. Combining selective GlyT inhibitors or developing non-selective GlyT inhibitors might have therapeutic value in NP.     

Integration of Data from Liquid–Liquid Phase Separation Databases Highlights Concentration and Dosage Sensitivity of LLPS Drivers

Liquid–liquid phase separation (LLPS) is a molecular process that leads to the formation of membraneless organelles, representing functionally specialized liquid-like cellular condensates formed by proteins and nucleic acids. Integrating the data on LLPS-associated proteins from dedicated databases revealed only modest agreement between them and yielded a high-confidence dataset of 89 human LLPS drivers. Analysis of the supporting evidence for our dataset uncovered a systematic and potentially concerning difference between protein concentrations used in a good fraction of the in vitro LLPS experiments, a key parameter that governs the phase behavior, and the proteomics-derived cellular abundance levels of the corresponding proteins. Closer scrutiny of the underlying experimental data enabled us to offer a sound rationale for this systematic difference, which draws on our current understanding of the cellular organization of the proteome and the LLPS process. In support of this rationale, we find that genes coding for our human LLPS drivers tend to be dosage-sensitive, suggesting that their cellular availability is tightly regulated to preserve their functional role in direct or indirect relation to condensate formation. Our analysis offers guideposts for increasing agreement between in vitro and in vivo studies, probing the roles of proteins in LLPS.     

Polymerization of isoprene in n-heptane with triethylaluminum–titanium tetrachloride catalyst. I. Effect of Al/Ti ratio on stereoregularity,conversion, molecular weight,and molecular weight distribution

The influence of the Al/Ti ratio of the heterogeneous Ziegler–Natta catalyst system AlEt₃–TiCl₄ on the isoprene polymerization in n-heptane at 30°C in a bench scale reactor was investigated. Each batch run consisted of 50 mL isoprene, 200 mL n-heptane with 0.45 and 0.5g catalyst. Conversion of isoprene and molecular weights of polyisoprene increased with increasing Al/Ti ratio, reaching their maxima values at 1.0 and 1.2, respectively. Conversion and molecular weights decreased remarkably at a higher ratio of Al/Ti. As this ratio decreases from the value of 1.2 to 0.4, the cis1,4 content in polyisoprene decreased from 97 to 25%. When the ratio of Al/Ti increased from 1.2 to 2.2 the cis-1,4 structure of polyisoprene decreased from 97 to 85%.     

Catalysts based in cerium oxide for wet oxidation of acrylic acid in the prevention of environmental risks

Acrylic acid is a refractory compound for the non-catalytic wet oxidation (WO) process and can seriously damage the environment when released in industrial effluents. Oxidation of acrylic acid by catalytic wet oxidation (CWO) was studied in slurry conditions in a high-pressure batch reactor at 200 °C and 15 bar of oxygen partial pressure. Several solid cerium-based catalysts prepared in our laboratory were used (Ag/Ce, Co/Ce, Mn/Ce, CeO, MnO) and evaluated in terms of activity, selectivity and stability. Mn/Ce shows the higher activity in 2 h with 97.7% reduction of total organic carbon (TOC) followed by: MnO(95.5%)>Ag/Ce(85.0%)>Co/Ce(65.1%)>CeO(61.2%). Attempts were also carried out to analyze the influence of different Mn/Ce molar ratios. High percentages of Mn lead to practically total organic carbon concentration (TOC) abatements while low ratios lead to the formation of non-oxidizable compounds. Acrylic acid was readily degraded by all the catalysts pointing out the high importance of using a catalytic process. pH was an indicator of the reaction pathway and acetic acid was found as the major reaction intermediate compound; however it is completely oxidized after 2 h with exception for Co/Ce, CeO and MnO. Carbon adsorption and leaching of metals were poorly found for Mn/Ce indicating high stability. The catalyst microstructure after the reaction was analyzed and formation of whiskers of β-MnO₂ (or less probably MnOOH) were observed at the catalyst surface. Therefore, Mn/Ce revealed to be a promising catalyst for the treatment of effluents containing acrylic acid; nevertheless, its commercialization depends on further research.     

Produkte der Dimerisierung ungesättigter Fettsäuren IX: Kinetische Untersuchungen zur Dimerisierung von Linolsäure

Products of the Dimerisation of Unsaturated Fatty Acids IX: Kinetic Studies about the Dimerisation of Linoleic Acid To clarify the dimerisation process of linoleic acid, we investigated samples taken in different time intervals. The first reaction step is a water addition at double bonds of the starting material and not double bond isomerisation as previously assumed. The resulting unsaturated monohydroxy fatty acids can cyclize with the second double bond in an intramolecular reaction forming 2,5-disubstituted tetrahydrofuran respectively 2,6-disubstituted tetrahydropyran derivates. Linolenic acid, present nearly always in small amounts in linoleic acid, reacts to first dimerisation products with linoleic acid by formation of a C-C-bond. The aliphatic dimers cyclize in an intramolecular reaction to mono-cyclic compounds. No dimeric acids, which would result from a Diels-Alder-reaction, could be identified. Bicyclic and aromatic dimeric acids can also be found in the early phase of the dimerisation. In the further progress of the reaction isomerisations, hydrogenations and dehydrogenations of the primary reaction products occur, thus the content of aromatic substances increases steadily.