Real-life use of ropeg-interferon α2b in polycythemia vera: patient selection and clinical outcomes.

Ropeginterferon-alfa2b (ropegIFNα2b) is a long-acting IFN formulation with broad FDA/EMA approval as a therapy of polycythemia vera (PV) with no symptomatic splenomegaly. There is currently lack of information on the real-world patient selection, including the impact of local reimbursement policies, and drug management, particularly: type/timing of screening and follow-up tests; absolute/relative contraindications to therapy; ropegIFNα2b dose and combinations with hydroxyurea. As a sub-analysis of the PV-ARC retrospective study (NCT06134102), we here report our monocenter experience with ropegIFNα2b in the period from January 2021, corresponding to drug availability outside clinical trial, and December 2023. Among the 149 patients with EMA/FDA indication, only 55 (36.9%) met the local reimbursement criteria and 18 (12.1%) received ropegIFNα2b. Thanks to appropriate screening, relative/absolute contraindications to ropegIFNα2b were detected and managed in a multidisciplinary manner. Efficacy and safety of ropegIFNα2b was confirmed, with 3 cases of early molecular response. General use of low ropegIFNα2b dose, with frequent need for hydroxyurea combinations, was noted. This real-world experience suggests a significant impact of local regulations on drug prescription and the need for greater real-world data collection on ropegIFNα2b in PV patients. Also, it describes appropriate multidisciplinary screening and monitoring procedures during ropegIFNα2b therapy.

The Reference Site Collaborative Network of the European Innovation Partnership on Active and Healthy Ageing.

Seventy four Reference Sites of the European Innovation Partnership on Active and Healthy Ageing (EIP on AHA) have been recognised by the European Commission in 2016 for their commitment to excellence in investing and scaling up innovative solutions for active and healthy ageing. The Reference Site Collaborative Network (RSCN) brings together the EIP on AHA Reference Sites awarded by the European Commission, and Candidate Reference Sites into a single forum. The overarching goals are to promote cooperation, share and transfer good practice and solutions in the development and scaling up of health and care strategies, policies and service delivery models, while at the same time supporting the action groups in their work. The RSCN aspires to be recognized by the EU Commission as the principal forum and authority representing all EIP on AHA Reference Sites. The RSCN will contribute to achieve the goals of the EIP on AHA by improving health and care outcomes for citizens across Europe, and the development of sustainable economic growth and the creation of jobs.

Regione Lombardia: a tool for improving quality in hospitals.

INTRODUCTION: The regional healthcare system of the Lombardy Region pay great attention to monitoring the effectiveness and quality level with which its services. The aim of this paper is to describe the method adopted by the Lombardy Region to create a governance tool for the healthcare system that would be applied within hospitals to create value at financial-economic level, to achieve continuous quality improvement and to increase patient/customer satisfaction levels. It was called: Piano Integrato del Miglioramento dell'Organizzazione (PIMO), i.e. Integrated Plan for Hospital Improvement. METODS: The approach for the definition of the PIMO was based on: the Plan Do Check Act methodology; the management requirements introduced by the UNI EN ISO 9001:2008 and UNI EN ISO 9004:2005 standards; the regulations and indications made for the Public Administration; the Guidelines for planning and monitoring improvement proposed by the CAF (Common Assessment Framework). RESULTS: The evaluation of the scores for all the health structures shows a good level of quality and qualifies PIMO as a strategic tool for hospitals. CONCLUSIONS: It will be necessary to allow this tool to operate for some time in order to make an overall assessment of the results achieved.

Pollen discrimination and classification by Fourier transform infrared (FT-IR) microspectroscopy and machine learning.

The discrimination and classification of allergy-relevant pollen was studied for the first time by mid-infrared Fourier transform infrared (FT-IR) microspectroscopy together with unsupervised and supervised multivariate statistical methods. Pollen samples of 11 different taxa were collected, whose outdoor air concentration during the flowering time is typically measured by aerobiological monitoring networks. Unsupervised hierarchical cluster analysis provided valuable information about the reproducibility of FT-IR spectra of the same taxon acquired either from one pollen grain in a 25 x 25 microm2 area or from a group of grains inside a 100 x 100 microm2 area. As regards the supervised learning method, best results were achieved using a K nearest neighbors classifier and the leave-one-out cross-validation procedure on the dataset composed of single pollen grain spectra (overall accuracy 84%). FT-IR microspectroscopy is therefore a reliable method for discrimination and classification of allergenic pollen. The limits of its practical application to the monitoring performed in the aerobiological stations were also discussed.

Isolation and molecular characterization of porcine calcitonin gene-related peptide (CGRP) and its endocrine effects in the porcine pancreas.

The aim of this study was to investigate the possible role of porcine calcitonin gene-related peptide (CGRP) in the regulation of the endocrine porcine pancreas. Initially, we isolated and purified CGRP from extracts of porcine adrenal glands and pancreases. A single molecular form of the peptide was found in the two tissues. The adrenal peptide was sequenced and found to differ from human alpha-CGRP at six positions and from human beta-CGRP at three positions. By immunohistochemistry, CGRP was found in nerve fibers in the pancreatic ganglia. A synthetic replica of the porcine peptide was infused at different dose levels (10(-10), 10(-9), and 10(-8) M) into isolated perfused porcine pancreata. With 5 mmol/L glucose in the perfusate. CGRP at 10(-10) and 10(-9) M increased insulin and glucagon secretion, whereas significant decreases were observed with 10(-8) M. Somatostatin secretion was increased significantly by 10(-8) M CGRP. In immunoneutralization studies (n = 6) using a high-affinity somatostatin antibody, the inhibitory effect of CGRP at 10(-8) M was reversed to a significant stimulation of insulin and glucagon secretion. Insulin secretion in response to square-wave increases in glucose concentration to 11 mM was inhibited dose dependently by CGRP; at 10(-8) M the insulin output decreased by 72+/-9% (n = 6). The present results indicate that CGRP may be involved in the regulation of insulin and glucagon secretion from the porcine pancreas.

Immunomodulatory effects of occupational exposure to mancozeb.

The effects of occupational exposure to ethylene-bis-dithiocarbamate of manganese and zinc on the immune system were evaluated in a group of mancozeb-exposed manufacturers and controls. The immune system tests revealed the following: (a) lymphocyte proliferative responses triggered by different activators and mitogen-induced IL-2 production were higher in exposed subjects than in controls; (b) production of monocyte/macrophage-derived IL-1 and polyclonal IgG and IgM, by beta-lymphocytes, did not differ between exposed subjects and controls; (c) percentages and absolute numbers of total T-cells, T-helper cells, T-suppressor/cytotoxic cells, activated T-cells, total beta-cells, and natural killer cells were similar in exposed subjects and controls; (d) serum immunoglobulin classes and complement fractions were within the range of normality; and (e) rheumatoid factor and non-organ-specific serum autoantibodies were absent in exposed and control subjects. An increase in T-cell functional response was found in the exposed group, suggesting a slight immunomodulator effect of mancozeb in conditions of low-level, prolonged occupational exposure.

Neuropeptides and morphological changes in cisplatin-induced dorsal root ganglion neuronopathy.

Dorsal root ganglia (DRG) neuronopathy was induced in rats by chronic treatment (2 mg/kg twice a week for nine injections) with the antineoplastic drug cisplatin. Morphological alterations and changes in peptide [calcitonin gene-related peptide (CGRP), substance P, galanin (Gal), and somatostatin] concentration were studied in the DRG, the spinal cord, and the sciatic nerve. Peptide concentration was increased in DRG neurons, with CGRP and Gal showing the highest increase. Conversely, in the sciatic nerve there was a general decrease in peptide content. In DRG a reduction in the nuclear, cytoplasmic, and nucleolar areas of primary sensory neurons was evident and was accompanied by clear-cut aspects of nucleolar structural damage. In peripheral nerves only extensive morphometric determinations could evidence a reduction in nerve conduction velocities and impairment in pain detection and coordination. Some of the nerve fibers presented axonal and adaxonal accumulations, suggesting the presence of an axonopathy. These results confirm that DRG cells are the primary target of cisplatin-induced neurotoxicity. Milder alterations can be detected in peripheral nerves. The increase in peptide concentration in DRG is probably due to cisplatin-related damage to the axonal transport system rather than to an increased synthesis.

Programmes and activities of the International Centre for Pesticide Safety.

The International Centre for Pesticide Safety (ICPS) is a research unit of the National Health Service created by the Government of the Region of Lombardy at the proposal of the World Health Organization-Regional Office for Europe, in cooperation with the University of Milan, and in agreement with the Italian Ministry of Health. ICPS operates in the following areas of activity: information and documentation on pesticide toxicity to man and environment, epidemiological, toxicological and clinical research on effects of pesticides in man; training and education of personnel in public health, assessment of environmental and health impact of pesticides by means of Geographical Information Systems, laboratory research for development and standardisation of methods for residue measurement in environmental and biological media. ICPS is also a centre of international meetings and continuing education courses. A number of projects carried out or underway at ICPS are briefly described in this paper.

Proglucagon processing in porcine and human pancreas.

In the pancreas proglucagon (PG), a peptide precursor of 160 amino acids is cleaved to produce glucagon and a 30-amino acid N-terminal flanking peptide, but the fate of the C-terminal flanking peptide (99 amino acids) is incompletely known. We subjected acid ethanol extracts of human and porcine pancreases to gel filtration and analyzed the fractions with specific radioimmunoassays for the following regions of proglucagon: PG 62-69, PG 72-81, PG 78-87, PG 98-107 amide, PG 126-134, and PG 149-158. Based on these assays and successive purifications by high performance liquid chromatography we isolated and purified to homogeneity three porcine peptides which were subjected to mass spectrometry and sequencing. One peptide was PG 64-69. The second was PG 72-108, as determined by mass spectrometry, N-terminal amino acid sequencing, and specific radioimmunoassays. The third had a molecular size of approximately 10,000, an N-terminal sequence corresponding to PG 72-81, and a C-terminal sequence terminating at PG 158 (specific radioimmunoassay). A similar peptide with an identical N-terminal sequence, a C-terminal sequence corresponding to PG 146-158, and a molecular mass of 9969 (theoretical mass for human PG 72-158 = 9971) was isolated from human pancreas together with small amounts of a peptide corresponding to PG 72-107 amide. Thus, the pancreatic processing of the C-terminal flanking peptide in proglucagon includes the formation of equimolar (to glucagon) amounts of PG 64-69 and PG 72-158 (major proglucagon fragment) and smaller amounts of N-terminally extended glucagon-like peptide-1 (GLP-1) (PG 72-108 in pigs and PG 72-107 amide in humans).

[The role of biotransformation in assessing the toxicological risk from pesticides]. / Ruolo della biotrasformazione nella valutazione del rischio tossicologico da pesticidi.

Pesticides are an extremely heterogeneous group of chemical compound with varying toxicity. The biotransformation and degradation mechanisms are essential factors in the regulation of their toxic effects and of their persistence in the environment and biological tissues. Knowledge of metabolism and measurement of metabolites are fundamental to evaluate the risk from residues in food and water and to assess the toxicological risk for subjects occupationally exposed to pesticides.

Pigs produce only a single form of CGRP, part of which is processed to N- and C-terminal fragments.

Using radioimmunoassays with two different antisera, one directed towards the C-terminal and one towards the mid part of porcine and human alpha-CGRP, respectively, we isolated three immunoreactive peptides from acid/ethanol extracts of porcine spinal cord by means of HPLC. By amino acid sequence analysis and mass spectrometry (PDMS), the most abundant peptide was found to be identical to the 37 residue CGRP previously isolated from porcine adrenal glands and spinal cord. The two remaining peptides were identified as pCGRP(18-37) and pCGRP(19-37). Furthermore, the oxidized forms (oxidized Met in position 22) of all three peptides were isolated. We extracted a large amount of tissue and the extractable peptides were purified without discarding side fractions. The purification steps were monitored by immunochemical methods that are highly sensitive for human alpha- and beta-CGRP. Yet we were unable to detect any second full-length form of CGRP. Thus, we conclude that only a single form of full-length CGRP is found in pigs and that this peptide may be cleaved to produce potentially bioactive N- and C-terminal fragments.

Biological monitoring of human exposure to atrazine.

Atrazine exposure was evaluated in six manufacturing workers by personal and biological monitoring. Total atrazine exposure varied from 10 to 700 mumol per workshift and total urinary atrazine excretion accounted for 1-2% of the external dose. The spectrum of the urinary atrazine metabolites comprises bi-dealkylated (80%), deisopropylated (10%), deethylated (8%) and unmodified atrazine (2%). The metabolites are eliminated in urine in slightly longer than 24 h: 50% of the amount is excreted in the first 8 h following the workshift.

On the effects of human galanin in man.

Human galanin was recently isolated and sequenced and was found to differ from porcine galanin, hitherto used for studies in humans, in several important respects. We therefore synthesized and purified human galanin and infused it i.v. at a rate of 74 pmol.kg-1.min-1 into six healthy volunteers for 60 min during a hyperglycaemic clamp. The clamp was achieved by i.v. infusion of glucose at a rate which in a control experiment had been demonstrated to maintain the plasma glucose level at 12-13 mmol/l for 90 min. Galanin concentrations reached a plateau of approximately 1500 pmol/l throughout the infusion as opposed to pre-infusion and control levels of 20-30 pmol/l. The glucose levels obtained in the two experiments were indistinguishable. Plasma levels of C-peptide and insulin increased significantly in both experiments and the dynamic concentration curves were almost identical. Glucagon concentrations in plasma decreased significantly and similarly. Growth hormone levels, however, increased eight-fold during galanin infusions. Galanin was eliminated from plasma with a half-life of 3.7 +/- 0.4 min, similar to that of porcine galanin. It is concluded that human galanin powerfully stimulates growth hormone secretion in man, but has no effect on pancreatic endocrine secretion or glucose metabolism in the concentrations obtained in this study.

Extrinsic control of the release of galanin and VIP from intrinsic nerves of isolated, perfused, porcine ileum.

By immunohistochemistry galanin-like immunoreactivity and vasoactive intestinal polypeptide (VIP)-like immunoreactivity were found in nerve cell bodies mostly in the submucous plexus and in nerve fibres in the mucosa, submucosa and muscularis including the myenteric plexus of the porcine ileum and were found to co-exist in most of these structures. Using isolated, perfused porcine ileum we studied the release of galanin and VIP in response to electrical stimulation of the mixed periarterial nerves or to intraarterial infusions of different neuroactive agents. Nerve stimulation (4-10 Hz) inhibited the basal release of galanin and VIP from the ileum (to 69 +/- 6 and 62 +/- 6% of basal release). After infusion of the alpha-adrenergic blocker, phentolamine, (10(-6) M) electrical stimulation increased the release of both galanin and VIP (to 140 +/- 12 and 133 +/- 13% of basal output). This increase was abolished by atropine (10(-6) M) and by hexamethonium (3.10(-5) M). Infusion of norepinephrine (10(-6) M) inhibited, whereas acetylcholine (10(-6) M) stimulated the release of both peptides. The effect of the latter was abolished by atropine. The inhibitory effect of nerve stimulation was not influenced by atropine. Our results suggest that the galanin- and VIP-producing intrinsic neurons receive inhibitory signals by noradrenergic nerve fibers and stimulatory signals mediated by cholinergic nerves, possibly via a cholinergic interneuron.

Distribution and characterization of different molecular products of pro-somatostatin in the hypothalamus and posterior pituitary lobe of the Mongolian gerbil (Meriones unguiculatus).

Antisera raised against various synthetic peptide fragments of the pro-somatostatin molecule were used to visualize immunohistochemically the distributions of different pro-somatostatin fragments in the hypothalamus and posterior pituitary of the Mongolian gerbil. To define the nature of the immunoreactive somatostatin-related molecular forms, gel chromatography combined with radioimmunoassays of hypothalamic and posterior pituitary extracts was performed. Within the hypothalamus, only trace amounts of somatostatin-28 and somatostatin-28(1-12) were present, whereas pro-somatostatin(1-76), pro-somatostatin(1-64), and somatostatin-14 peptides were present in equimolar amounts. In contrast, the posterior pituitary lobe contained equal amounts of somatostatin-14, somatostatin-28, and somatostatin-28(1-12) but no pro-somatostatin(1-76), indicating that pro-somatostatin is further processed during the axonal flow to posterior pituitary nerve terminals. The gel chromatographic data were further substantiated by immunohistochemical data. Thus, perikarya containing all of these five immunoreactivities were strictly confined to the periventricular area and parvocellular subset of the paraventricular nucleus. However, the number of somatostatin-28- and somatostatin-28(1-12)-immunoreactive perikarya was approximately 20% of the number of somatostatin-14- and pro-somatostatin(1-64)-immunoreactive cells. In other hypothalamic areas only somatostatin-14 and pro-somatostatin(1-64) immunoreactivities were detectable in cell bodies. These cell bodies were encountered in the organum vasculosum laminae terminalis; the suprachiasmatic, ventromedial, arcuate, perifornical, and posterior hypothalamic nuclei; and the median preoptic and retrochiasmatic areas. In situ hybridization histochemistry revealed that the cellular distribution of pro-somatostatin mRNA corresponds to that of somatostatin-14 and pro-somatostatin immunoreactivity, suggesting that the immunoreactive material observed within the cell bodies is synthetized there and that the differences in density of immunoreactivities may be explained by intracellular processing of pro-somatostatin. Somatostatinergic nerve fibers and terminals in hypothalamic areas and the posterior pituitary lobe were immunoreactive to all of the employed antisera. From the present results, obvious differences between intrahypothalamic and hypothalamo-pituitary somatostatinergic neurons emerge. Within hypothalamic neurons not projecting to the median eminence and the posterior pituitary lobe, pro-somatostatin is posttranslationally processed in the cell body predominantly into pro-somatostatin(1-64) and somatostatin-14. Otherwise, within periventricular neurons projecting to the median eminence and the posterior pituitary lobe, pro-somatostatin is posttranslationally processed during the axonal flow into pro-somatostatin(1-64), somatostatin-14, somatostatin-28, and somatostatin-28(1-12).

Nerve fibers in the rat posterior pituitary lobe contain prosomatostatin (1-64).

The neurohormone and neurotransmitter somatostatin arises from the processing of a larger precursor, prosomatostatin (proSS). An immunohistochemical investigation in the rat, using a well-characterized antiserum raised against a synthetic peptide identical to the 20-36 residues of the proSS molecule, revealed the presence of immunoreactive nerve fibers and nerve terminals in the median eminence, infundibulum, infundibular stalk and posterior pituitary lobe. The largest number of immunoreactive nerve fibers and nerve terminals was observed in apposition to the portal vessels, whereas a moderate number of proSS-immunoreactive fibers was identified in the infundibular stalk and in the proximal part of the posterior pituitary lobe. The proSS-immunoreactive nerves entered the posterior pituitary lobe from the infundibular and pituitary stalks and were followed to rostral and ventral aspects of the organ. In contrast, positive fibers were rarely identified in caudal and posterior parts. Extracts of rat posterior pituitaries subjected to gel chromatography and reversed-phase high-pressure liquid chromatography (HPLC) analysis showed the presence of a single proSS-immunoreactive molecule corresponding to the size of proSS(1-64). The functional significance of the proSS(1-64) in the hypothalamus and pituitary is at present unknown, but its location in the hypothalamo-hypophyseal system suggests that this end product of the processing of proSS is released into the portal and perhaps also the general circulation.

Galanin and galanin extended at the N-terminus with seven and nine amino acids are produced in and secreted from the porcine adrenal medulla in almost equal amounts.

Galanin is present in high concentrations in porcine adrenals, but nothing is known about the processing and secretion of other products of the 123-amino acid precursor preprogalanin. Using, in combination, RIA against galanin, a variety of chromatographic procedures, mass spectrometry, and amino acid sequencing, we studied the processed and the secreted products of preprogalanin. From the tissue extracts we isolated in equimolar amounts and sequenced two major pools of galanin immunoreactive peptides: galanin and two N-terminally extended forms, preprogalanin-(24-61) and preprogalanin-(26-61). The same peptides were identified upon gel chromatography and analytical HPLC in effluents collected during electrical stimulation of the intact splanchnic nerve supply of an isolated perfused preparation of porcine adrenals. The processing of preprogalanin in porcine adrenals thus includes the formation and release of galanin, preprogalanin-(24-61), and preprogalanin-(26-61). The signal peptidase cleaves the preprogalanin at either Gly23 or Gly25.

Substance P and neurokinin A are codistributed and colocalized in the porcine gastrointestinal tract.

Immunoreactive substance P and neurokinin A were measured with radioimmunoassay in extracts of different segments of porcine gastrointestinal tract using C-terminally directed antisera. In all segments, the concentrations of substance P and neurokinin A were similar. The largest concentrations of both peptides were found in the mid-colon. By gel chromatography and reversed-phase high pressure liquid chromatography the immunoreactivity in extracts from ileum eluted as homogenous peptides at the positions of synthetic substance P and neurokinin A, respectively. No neurokinin B was found. By immunohistochemistry of porcine duodenum, jejunum, ileum and mid-colon, identical localization patterns were found for substance P and neurokinin A, and the two peptides demonstrated by double immunofluorescence to be colocalized in the enteric nervous system of the ileum. We conclude that the tachykinins substance P and neurokinin A are codistributed and colocalized in the procine gastrointestinal tract and suggest that the two peptides are produced from a common precursor, beta- and/or gamma-preprotachykinin, in the same neurons.

Identification, characterization and release of GRP gene-associated peptides from the normal porcine and human gastro-intestinal tract.

Using a radioimmunoassay directed towards human proGRP (42-53) on acetic acid extracts, immunoreactivity was measured throughout the porcine GI-tract in concentrations that were parallel to those of GRP (gastrin-releasing peptide or 'mammalian bombesin'). Gel filtration and HPLC studies of human and porcine tissue extracts revealed that the immunoreactivity was mainly due to a peptide with a molecular size of 8-9 kDa. The peptide did not contain the GRP sequence, making it a major fragment of the GRP C-flanking part of proGRP. Furthermore, a peptide of similar size with proGRP (42-53) immunoreactivity was released from isolated, perfused preparations of porcine antral and non-antral stomach and pancreas in parallel with GRP in response to electrical stimulation of the vagus nerves. Our results suggest that a processing of preproGRP occurs in normal, adult human and porcine tissues, that is similar to that previously demonstrated in small cell lung carcinomas and human fetal lungs. The finding that the immunoreactive proGRP fragment is released from the tissues upon appropriate stimulation raises the question of a possible physiological role for proGRP products other than GRP.