MISURAZIONE DIRETTA DEL TRAFFICO NERVOSO SIMPATICO IN DIFFERENTI FASI DELL'ACROMEGALIA

Sympathovagal imbalance has been shown in acromegaly by indirect measurements of adrenergic tone. Data regarding direct measurement of sympathetic activity are lacking as yet. Aim of this study was to assess the adrenergic tone through direct recording of muscle sympathetic nerve activity (MSNA) in acromegalic patients. Skin sympathetic nerve traffic (SSNA) was also recordered. Study: After evaluating anthropometric and echocardiographic parameters, anterior pituitary function, glucose and lipid metabolism, and measuring plasma leptin, direct recording of sympathetic outflow via the microneurographic technique was performed in the following groups of subjects: 15 newly diagnosed acromegalics without hyperprolactinaemia, pituitary hormone deficiencies, obstructive sleep apnoea and cardiac hypertrophy (GROUP 1); 22 patients on somatostatin analogues (SSA), 11 of whom attaining biochemical control according to the currently accepted criteria and 11 not attaining biochemical control (GROUP 3); 2 acromegalic patients affected by OSAS (GROUP 4); 10 patients cured from acromegaly after neurosurgery (GROUP 5). The 15 newly diagnosed acromegalic patients were also studied for SSNA (GROUP 2). Fiften normalweight healthy subjects serving as controls. Results For similar anthropometric and metabolic parameters in patients and contros the group 1 displayed insulin resistance and a marked sympathetic inhibition (MSNA 18\u20223 \ub1 8\u202210) vs controls (37\u20223 \ub1 6\u202248 bursts/ min). A significant reduction in plasma leptin (1\u20226 \ub1 1\u202204 vs 6\u20225 \ub1 2\u202201 lg/l, P < 0\u20220001) was also recorded in patients. Patients on SSA (GROUP 3), either with controlled or uncontrolled disease, displayed mean MSNA values (27.4 \ub1 8.24 and 31.6 \ub1 3.27 bursts/min, respectively) significantly lower than those shown by controls (p < 0.01) but significantly higher than those found in untreated acromegalics (p < 0.05). Mean MSNA values were not significantly different between controlled and uncontrolled SSA-treated patients. GROUP 5 showed mean MSNA values (30.17 \ub1 3.2 bursts/min) significantly higher than those shown by GROUP 1 (p< 0.01) and than patients non controlled by SSA (p<0.05), but superimposable to that of patients controlled by SSA and controls. Mean leptin levels of cured were significantly higher than those shown by active patients (p<0.01). MSNA levels registered in two patients affected by OSAS (GROUP 4) was higher than those found in acromegalic patients without OSAS. There were no significant differences in SSNA between GROUP 2 and controls. Considering the whole population IGF-1 levels were negative correlated with MSNA and leptin levels ( p<0.05 and p<0.0005, respectively) and MSNA was positive correlated with leptin levels (p<0.05). Comment. Our study demonstrates that recently diagnosed acromegalic patients, in spite of insulin resistance (a condition known to increase MSNA), display a decreased sympathetic outflow. This finding, together with the tendency to normalization of adrenergic tone in pharmacologically treated and cured acromegalic patients and the negative correlations found between IGF-1 on the one hand and leptin and MSNA on the other hand, and the positive correlation between leptin and MSNA, suggests a relevant influence of the GH/IGF-I axis on the activity of the sympathetic nervous system through leptin modification

Standardized ultrasound report for thyroid nodules : the endocrinologist&apos;s viewpoint

BACKGROUND: Ultrasonography (US) plays a crucial role in the diagnostic management of thyroid nodules, but its widespread use in clinical practice might generate heterogeneity in ultrasound reports. OBJECTIVES: The aims of the study were to propose (a) a standardized lexicon for description of thyroid nodules in order to reduce US reports of interobserver variability and (b) a US classification system of suspicion for thyroid nodules in order to promote a uniform management of thyroid nodules. METHODS: RELEVANT PUBLISHED ARTICLES WERE IDENTIFIED BY SEARCHING MEDLINE AT PUBMED COMBINING THE FOLLOWING SEARCH TERMS: ultrasonography, thyroid, nodule, malignancy, carcinoma, and classification system. Results were supplemented with our data and experience. RESULTS: A STANDARDIZED US REPORT SHOULD ALWAYS DOCUMENT POSITION, EXTRACAPSULAR RELATIONSHIPS, NUMBER, AND THE FOLLOWING CHARACTERISTICS OF EACH THYROID LESION: shape, internal content, echogenicity, echotexture, presence of calcifications, margins, vascularity, and size. Combining the previous US features, each thyroid nodule can be tentatively classified as: malignant, suspicious for malignancy, borderline, probably benign, and benign. CONCLUSIONS: We propose a standardized US report and a tentative US classification system that may become helpful for endocrinologists dealing with thyroid nodules in their clinical practice. The proposed classification does not allow to bypass the required cytological confirmation, but may become useful in identifying the lesions with a lower risk of neoplasm

Femtosecond laser preparation of resin embedded samples for correlative microscopy workflows in life sciences

Correlative multimodal imaging is a useful approach to investigate complex structural relations in life sciences across multiple scales. For these experiments, sample preparation workflows that are compatible with multiple imaging techniques must be established. In one such implementation, a fluorescently labeled region of interest in a biological soft tissue sample can be imaged with light microscopy before staining the specimen with heavy metals, enabling follow-up higher resolution structural imaging at the targeted location, bringing context where it is required. Alternatively, or in addition to fluorescence imaging, other microscopy methods, such as synchrotron x-ray computed tomography with propagation-based phase contrast or serial blockface scanning electron microscopy, might also be applied. When combining imaging techniques across scales, it is common that a volumetric region of interest (ROI) needs to be carved from the total sample volume before high resolution imaging with a subsequent technique can be performed. In these situations, the overall success of the correlative workflow depends on the precise targeting of the ROI and the trimming of the sample down to a suitable dimension and geometry for downstream imaging. Here, we showcase the utility of a femtosecond laser (fs laser) device to prepare microscopic samples (1) of an optimized geometry for synchrotron x-ray tomography as well as (2) for volume electron microscopy applications and compatible with correlative multimodal imaging workflows that link both imaging modalities

Intermittent C1-Inhibitor Deficiency Associated with Recessive Inheritance: Functional and Structural Insight

C1-inhibitor is a serine protease inhibitor (serpin) controlling complement and contact system activation. Gene mutations result in reduced C1-inhibitor functional plasma level causing hereditary angioedema, a life-threatening disorder. Despite a stable defect, the clinical expression of hereditary angioedema is unpredictable, and the molecular mechanism underlying this variability remains undisclosed. Here we report functional and structural studies on the Arg378Cys C1-inhibitor mutant found in a patient presenting reduced C1-inhibitor levels, episodically undergoing normalization. Expression studies resulted in a drop in mutant C1-innhibitor secretion compared to wild-type. Notwithstanding, the purified proteins had similar features. Thermal denaturation experiments showed a comparable denaturation profile, but the mutant thermal stability decays when tested in conditions reproducing intracellular crowding.Our findings suggest that once correctly folded, the Arg378Cys C1-inhibitor is secreted as an active, although quite unstable, monomer. However, it could bear a folding defect, occasionally promoting protein oligomerization and interfering with the secretion process, thus accounting for its plasma level variability. This defect is exacerbated by the nature of the mutation since the acquired cysteine leads to the formation of non-functional homodimers through inter-molecular disulphide bonding. All the proposed phenomena could be modulated by specific environmental conditions, rendering this mutant exceptionally vulnerable to mild stress

3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy

The processes of life take place in multiple dimensions, but imaging these processes in even three dimensions is challenging. Here, we describe a workflow for 3D correlative light and electron microscopy (CLEM) of cell monolayers using fluorescence microscopy to identify and follow biological events, combined with serial blockface scanning electron microscopy to analyse the underlying ultrastructure. The workflow encompasses all steps from cell culture to sample processing, imaging strategy, and 3D image processing and analysis. We demonstrate successful application of the workflow to three studies, each aiming to better understand complex and dynamic biological processes, including bacterial and viral infections of cultured cells and formation of entotic cell-in-cell structures commonly observed in tumours. Our workflow revealed new insight into the replicative niche of Mycobacterium tuberculosis in primary human lymphatic endothelial cells, HIV-1 in human monocytederived macrophages, and the composition of the entotic vacuole. The broad application of this 3D CLEM technique will make it a useful addition to the correlative imaging toolbox for biomedical research

Trafficking of storage proteins in developing grain of wheat

The processing properties of the wheat flour are largely determined by the structures and interactions of the grain storage proteins (also called gluten proteins) which form a continuous visco-elastic network in dough. Wheat gluten proteins are classically divided into two groups, the monomeric gliadins and the polymeric glutenins, with the latter being further classified into low molecular weight (LMW) and high molecular weight (HMW) subunits. The synthesis, folding and deposition of the gluten proteins take place within the endomembrane system of the plant cell. However, determination of the precise routes of trafficking and deposition of individual gluten proteins in developing wheat grain has been limited in the past by the difficulty of developing monospecific antibodies. To overcome this limitation, a single gluten protein (a LMW subunit) was expressed in transgenic wheat with a C-terminal epitope tag, allowing the protein to be located in the cells of the developing grain using highly specific antibodies. This approach was also combined with the use of wider specificity antibodies to compare the trafficking and deposition of different gluten protein groups within the same endosperm cells. These studies are in agreement with previous suggestions that two trafficking pathways occur in wheat, with the proteins either being transported via the Golgi apparatus into the vacuole or accumulating directly within the lumen of the ER. They also suggest that the same individual protein could be trafficked by either pathway, possibly depending on the stage of development, and that segregation of gluten proteins both between and within protein bodies may occur

A facility for radiation hardness studies based on a medical cyclotron

The development of instrumentation for operation in high-radiation environments represents a challenge in various research fields, particularly in particle physics experiments and space missions, and drives an ever-increasing demand for irradiation facilities dedicated to radiation hardness studies. Depending on the application, different needs arise in terms of particle type, energy and dose rate. In this article, we present a versatile installation based on a medical cyclotron located at the Bern University Hospital (Inselspital), which is used as a controlled 18-MeV proton source. This accelerator is used for daily production of medical radioisotopes, as well as for multidisciplinary research, thanks to a 6.5-meter long beam transfer line that terminates in an independent bunker, dedicated only to scientific activities. The facility offers a wide range of proton fluxes, due to an adjustable beam current from approximately 10 pA to the micro-ampere range, together with a series of steering and focusing magnets along the beamline that allow for the beam spot to be focused down to a few mm^2. The beamline can be instrumented with a variety of beam monitoring detectors, collimators, and beam current measurement devices to precisely control the irradiation conditions. The facility also hosts a well equipped laboratory dedicated to the characterisation of samples after irradiation. An experimental validation of the irradiation setup, with proton fluxes ranging from 5×10^9 cm^-2s^-1 to 4×10^11 cm^-2s^-1, is reported

VID22 counteracts G-quadruplex-induced genome instability

Genome instability is a condition characterized by the accumulation of genetic alterations and is a hallmark of cancer cells. To uncover new genes and cellular pathways affecting endogenous DNA damage and genome integrity, we exploited a Synthetic Genetic Array (SGA)-based screen in yeast. Among the positive genes, we identified VID22, reported to be involved in DNA double-strand break repair. vid22Δ cells exhibit increased levels of endogenous DNA damage, chronic DNA damage response activation and accumulate DNA aberrations in sequences displaying high probabilities of forming G-quadruplexes (G4-DNA). If not resolved, these DNA secondary structures can block the progression of both DNA and RNA polymerases and correlate with chromosome fragile sites. Vid22 binds to and protects DNA at G4-containing regions both in vitro and in vivo. Loss of VID22 causes an increase in gross chromosomal rearrangement (GCR) events dependent on G-quadruplex forming sequences. Moreover, the absence of Vid22 causes defects in the correct maintenance of G4-DNA rich elements, such as telomeres and mtDNA, and hypersensitivity to the G4-stabilizing ligand TMPyP4. We thus propose that Vid22 is directly involved in genome integrity maintenance as a novel regulator of G4 metabolism.Associazione Italiana per la Ricerca sul Cancro (AIRC) 15631, 21806MIUR PRIN 2015- 2015SJLMB9, PRIN 2017-2017KSZZJW, PRIN2017-2017Z55KCMinisterio de Economía y Competitividad BFU2016- 75058-PCanadian Institutes of Health Research FDN-15991

VID22 counteracts G-quadruplex-induced genome instability

Genome instability is a condition characterized by the accumulation of genetic alterations and is a hallmark of cancer cells. To uncover new genes and cellular pathways affecting endogenous DNA damage and genome integrity, we exploited a Synthetic Genetic Array (SGA)-based screen in yeast. Among the positive genes, we identified VID22, reported to be involved in DNA double-strand break repair. vid22Δ cells exhibit increased levels of endogenous DNA damage, chronic DNA damage response activation and accumulate DNA aberrations in sequences displaying high probabilities of forming G-quadruplexes (G4-DNA). If not resolved, these DNA secondary structures can block the progression of both DNA and RNA polymerases and correlate with chromosome fragile sites. Vid22 binds to and protects DNA at G4-containing regions both in vitro and in vivo. Loss of VID22 causes an increase in gross chromosomal rearrangement (GCR) events dependent on G-quadruplex forming sequences. Moreover, the absence of Vid22 causes defects in the correct maintenance of G4-DNA rich elements, such as telomeres and mtDNA, and hypersensitivity to the G4-stabilizing ligand TMPyP4. We thus propose that Vid22 is directly involved in genome integrity maintenance as a novel regulator of G4 metabolism