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The analysis of sterol profile consists in the qualitative and quantitative determination of cholesterol and non-cholesterol sterols. Non-cholesterol sterols include the cholesterol precursors in de novo biosynthesis pathway, cholesterol catabolites and plant sterols absorbed from diet. The sterol profile analysis can be performed on different biological matrices, i.e., plasma/serum, whole blood, erythrocyte membranes, tissues and cells, depending on the clinical and/or experimental purpose. The reference method for the sterol profile analysis is the gas-chromatography coupled with mass spectrometry, although new methods based on liquid-chromatography or direct mass spectrometry have been developed. The sterol profile analysis represents a fundamental tool for the biochemical diagnosis of the congenital defects of cholesterol metabolism. In addition, some non-cholesterol sterols have been validated as surrogate markers of de novo synthesis and intestinal absorption of cholesterol. Therefore, the sterol profile analysis has been used for the evaluation of cholesterol homeostasis in different diseases. This review focuses on these applications and our clinical and experimental findings. In the last nine years, we identified 18 new cases of the cholesterol metabolism defects and we found that cholesterol metabolism is impaired in patients with cystic fibrosis.

Spectroscopic and MS techniques were used to characterize the pigments and the composition of polar and nonpolar binders of a stray wall painting fragment from Liternum (Italy) archaeological excavation. X-ray fluorescence and diffraction analysis of the decorations indicated mainly the presence of calcite, quartz, hematite, cinnabar, and cuprorivaite. Infrared spectroscopy, GC coupled to flame-ionization detector, and MS analysis of the polar and nonpolar components extracted from paint layers from three different color regions revealed the presence of free amino acids, sugars, and fatty acids. Interestingly, LC-MS shotgun analysis of the red painting region showed the presence of αS1-casein of buffalo origin. Compared to our previous results from Pompeii's wall paintings, even though the Liternum painting mixture contained also binders of animal origin, the data strongly suggest that in both cases a tempera painting technique was utilized. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

In recent months, Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread throughout the world. COVID-19 patients show mild, moderate or severe symptoms with the latter ones requiring access to specialized intensive care. SARS-CoV-2 infections, pathogenesis and progression have not been clearly elucidated yet, thus forcing the development of many complementary approaches to identify candidate cellular pathways involved in disease progression. Host lipids play a critical role in the virus life, being the double-membrane vesicles a key factor in coronavirus replication. Moreover, lipid biogenesis pathways affect receptor-mediated virus entry at the endosomal cell surface and modulate virus propagation. In this study, targeted lipidomic analysis coupled with proinflammatory cytokines and alarmins measurement were carried out in serum of COVID-19 patients characterized by different severity degree. Serum IL-26, a cytokine involved in IL-17 pathway, TSLP and adiponectin were measured and correlated to lipid COVID-19 patient profiles. These results could be important for the classification of the COVID-19 disease and the identification of therapeutic targets.

Background Coronavirus disease 2019 (COVID-19) pandemic had a 1st wave in Europe from March to May 2020 and a 2nd wave since September 2020. We previously studied 35 hospitalized COVID-19 patients of the 1st wave demonstrating a cytokine storm and the exhaustion of most lymphocyte subpopulations. Herein, we describe the results obtained from COVID-19 patients of the 2nd wave. Methods We analyzed interleukin (IL)-6 by human-specific enzyme-linked immunosorbent assay and a large set of lymphocyte subpopulations by flow cytometry in 274 COVID-19 patients hospitalized from September 2020 to May 2021. Results Patients of 2nd wave compared with those of 1st wave showed lower serum IL-6 levels and a higher number of B and most T lymphocyte subpopulations in advanced stages, in relation with the age and the gender. On the other hand, we observed in 2nd wave patients: (i) a reduction of most lymphocyte subpopulations at mild and moderate stages; (ii) a reduction of natural killer cells and T regulatory cells together with a higher number of activated T helper (TH) 17 lymphocytes in all stages, which were mainly related to steroid and azithromycin therapies before hospitalization. Conclusions COVID-19 had a less severe impact in patients of the 2nd wave in advanced stages, while the impact appeared more severe in patients of mild and moderate stages, as compared with 1st wave patients. This finding suggests that in COVID-19 patients with milder expression at diagnosis, steroid and azithromycin therapies appear to worsen the immune response against the virus. Furthermore, the cytometric profile may help to drive targeted therapies by monoclonal antibodies to modulate specific IL/lymphocyte inhibition or activation in COVID-19 patients.

Oxylipins are signaling molecules originated by fatty acids that modulate vascular and bronchial tone, bronchial secretion, cytokine production and immune cell activity. The unbalanced production of pro-inflammatory and pro-resolving (i.e., anti-inflammatory) oxylipins has a relevant role in the pathogenesis of pulmonary inflammation like in cystic fibrosis (CF). We analyzed by LC-MRM/MS 65 oxylipins and 4 fatty acids in resting saliva from 69 patients with CF and 50 healthy subjects (controls). The salivary levels of 48/65 oxylipins were significantly different between CF patients and controls. Among these, EpETE, DHET, 6ketoPGE1 and HDHA were significantly higher in saliva from CF patients than in controls. All these molecules display anti-inflammatory effects, i.e., releasing of bronchial and vascular tone, modulation of cytokine release. While 20-hydroxyPGF2A, PGB2, EpDPE, 9 K-12-ELA, bicyclo-PGE2, oleic acid, LTC4, linoleic acid, 15oxoEDE, 20 hydroxyPGE2 and DHK-PGD2/PGE2 (mostly associated to pro-inflammatory effects) resulted significantly lower in CF patients than in controls. Our data suggest that the salivary oxylipins profile in CF patients is addressed toward a global anti-inflammatory effect. Although these findings need be confirmed on larger populations in prospective studies, they will contribute to better understand the pathogenesis of CF chronic inflammation and to drive targeted therapies based on the modulation of oxylipins synthesis and degradation.

The molecular basis of the wide clinical heterogeneity of Coronavirus disease 2019 (COVID-19) is still unknown. Matrix metalloproteinases (MMPs) may have a role in the lung damage and regeneration that occur in severe patients. We studied serum MMP3 and MMP9 as potential biomarkers of COVID-19 severity, in 108 hospitalized patients with different World Health Organization (WHO) severity stage and in 48 controls. At hospital admission, serum MMP3 was increased in COVID-19 patients with a significant trend along the progression of the WHO stage, while serum levels of MMP9 were significantly increased in COVID-19 patients with no correlation with disease severity. At 1 week from hospitalization, MMP3 was reduced, suggesting an early pathogenic role of the protein in lung inflammation, while MMP9 levels were further increased, indicating a late role of the protein in the inflammatory process, specifically during the repairing phase. Furthermore, serum MMP9 was positively correlated with serum interleukin-6, myeloperoxidase, and circulating neutrophils and monocytes number. In conclusion, serum MMP3 may help to early predict the severity of COVID-19 and both proteins, MMP3 and MMP9, may contribute to define severe COVID-19 patients that may benefit from a targeted therapy on MMPs.

Endothelial hyperinflammation and vasculitis are known hallmarks of acute COVID-19 and multisystem inflammatory syndrome in children (MIS-C). They are due to the direct effect of the virus on endothelial cells enhanced by pro-inflammatory modulators and may cause venous/arterial thrombosis. Therefore, it is essential to identify patients with endothelial damage early in order to establish specific therapies. We studied the monocyte chemoattractant protein 1 (MCP-1), the perinuclear anti-neutrophil cytoplasmic antibodies (pANCA), and the vascular endothelial growth factor A (VEGF-A) in serum from 45 MIS-C patients at hospital admission and 24 healthy controls (HC). For 13/45 MIS-C patients, we measured the three serum biomarkers also after one week from hospitalization. At admission, MIS-C patients had significantly higher levels of MCP-1 and VEGF-A than the HC, but no significant differences were observed for pANCA. While after one week, MCP-1 was significantly lower, pANCA was higher and VEGF-A levels were not significantly different from the admission values. These findings suggest an involvement of epithelium in MIS-C with an acute phase, showing high MCP-1 and VEGF-A, followed by an increase in pANCA that suggests a vasculitis development. The serum biomarker levels may help to drive personalized therapies in these phases with anticoagulant prophylaxis, immunomodulators, and/or anti-angiogenic drugs.

Profound clinical differences between the first and second waves of COVID-19 were observed in Europe. Nitric oxide (NO) may positively impact patients with Severe Acute Respiratory Syndrome CoronaVirus-2 (SARS-CoV-2) infection. It is mainly generated by inducible nitric oxide synthase (iNOS). We studied serum iNOS levels together with serum interleukin (IL)-6 and IL-10 in patients with SARS-CoV-2 infection in the first wave (n = 35) and second wave (n = 153). In the first wave, serum iNOS, IL-6, IL-10 levels increased significantly, in line with the World Health Organization (WHO) score severity, while in the second wave, iNOS did not change with the severity. The patients of the second wave showed lower levels of iNOS, IL-6, and IL-10, as compared to the corresponding subgroup of the first wave, suggesting a less severe outcome of COVID-19 in these patients. However, in the severe patients of the second wave, iNOS levels were significantly lower in patients treated with steroids or azithromycin before the hospitalization, as compared to the untreated patients. This suggests an impairment of the defense mechanism against the virus and NO-based therapies as a potential therapy in patients with low iNOS levels.

Metabolomics helps identify metabolites to characterize/refine perturbations of biological pathways in living organisms. Pre-analytical, analytical, and post-analytical limitations that have hampered a wide implementation of metabolomics have been addressed. Several potential biomarkers originating from current targeted metabolomics-based approaches have been discovered. Precision medicine argues for algorithms to classify individuals based on susceptibility to disease, and/or by response to specific treatments. It also argues for a prevention-based health system. Because of its ability to explore gene–environment interactions, metabolomics is expected to be critical to personalize diagnosis and treatment. Stringent guidelines have been applied from the very beginning to design studies to acquire the information currently employed in precision medicine and precision prevention approaches. Large, prospective, expensive and time-consuming studies are now mandatory to validate old, and discover new, metabolomics-based biomarkers with high chances of translation into precision medicine. Metabolites from studies on saliva, sweat, breath, semen, feces, amniotic, cerebrospinal, and broncho-alveolar fluid are predicted to be needed to refine information from plasma and serum metabolome. In addition, a multi-omics data analysis system is predicted to be needed for omics-based precision medicine approaches. Omics-based approaches for the progress of precision medicine and prevention are expected to raise ethical issues.

COVID-19 is a global threat that has spread since the end of 2019, causing severe clinical sequelae and deaths, in the context of a world pandemic. The infection of the highly pathogenetic and infectious SARS-CoV-2 coronavirus has been proven to exert systemic effects impacting the metabolism. Yet, the metabolic pathways involved in the pathophysiology and progression of COVID-19 are still unclear. Here, we present the results of a mass spectrometry-based targeted metabolomic analysis on a cohort of 52 hospitalized COVID-19 patients, classified according to disease severity as mild, moderate, and severe. Our analysis defines a clear signature of COVID-19 that includes increased serum levels of lactic acid in all the forms of the disease. Pathway analysis revealed dysregulation of energy production and amino acid metabolism. Globally, the variations found in the serum metabolome of COVID-19 patients may reflect a more complex systemic perturbation induced by SARS-CoV-2, possibly affecting carbon and nitrogen liver metabolism.

The last ten years have been characterized by an enormous step forward in the therapy and management of patients with Cystic Fibrosis (CF), thanks to the development and combination of Cystic Fibrosis Transmembrane Receptor (CFTR) correctors and potentiators. Specifically, the last approved triple combination elexacaftor/tezacaftor/ivacaftor has been demonstrated to improve lung function in CF patients with both homozygous Phe508del and Phe508del/minimal function genotypes. Here we have assessed the effect of elexacaftor/tezacaftor/ivacaftor in patients carrying the Phe508del/minimal function genotype (n = 20) after one year of treatments on liver function and nutrient absorption with a focus on lipid metabolism. We show that weight, BMI, and albumin significantly increase, suggesting a positive impact of the treatment on nutrient absorption. Furthermore, cholesterol levels as a biomarker of lipid metabolism increased significantly after one year of treatment. Most importantly, we suggest that these results were not dependent on the diet composition, possibly indicating that the drug improves the hepatic synthesis and secretion of proteins and cholesterol.

Background and objectives: ischemic stroke (IS) is among the most frequent causes of death worldwide, thus, it is of paramount relevance to know predisposing factors that may help to identify and treat the high-risk subjects. Materials and Methods:we tested nine variants in genes involved in thrombotic pathway in 282 patients that experienced IS and 87 that had transient ischemic attacks (TIA) in comparison to 430 subjects from the general population (GP) of the same geographic area (southern Italy). We included cases of young and child IS to evaluate the eventual differences in the role of the analyzed variants. Results: we did not observe significant differences between TIA and the GP for any of the variants, while the allele frequencies of methylene-tetrahydrofolate reductase (MTHFR) C677T, beta-fibrinogen -455G&gt, A and factor (FXIII) V34L were significantly higher in patients with IS than in the subjects from the GP. No significant interaction was observed with sex. Conclusions: the present data argue that some gene variants have a role in IS and this appears to be an interesting possibility to be pursued in large population studies to help design specific strategies for IS prevention.

BackgroundWe report the case of a woman with non-Hodgkin lymphoma who remained positive on the molecular assay for SARS-CoV-2 for six months: she has never experienced a severe form of COVID-19 although in absence of seroconversion.MethodsThe whole SARS-CoV-2 genome analysis was performed by the CleanPlex SARS-CoV-2 Research and Surveillance NGS Panel (PARAGON GENOMICS, Hayward, USA).ResultsWe found twenty-two mutations in SARS-CoV-2 genome and a novel deleterious ORF3a frameshift c.766_769del corresponding to a unique and novel lineage. The region affected by this frameshift variant is reported as being important in determining SARS-CoV-2 immunogenicity. Patient’s immunophenotype showed the absence of B lymphocytes and significantly reduced T-cell count. Only after the treatment with hyperimmune plasma she finally became negative on the swab.ConclusionsOur findings could be helpful in the management of patients with immunodeficiency, particularly when novel variants, potentially altering the virus immune response, are present.

COVID-19 may appear with a widely heterogeneous clinical expression. Thus, predictive markers of the outcome/progression are of paramount relevance. The neutrophil/lymphocyte ratio (NLR) has been suggested as a good predictive marker of disease severity and mortality. Accordingly, we found that NLR significantly increased in parallel with the WHO severity stage in COVID-19 patients during the Ist wave (March-May 2020; n = 49), due to the significant reduction of lymphocyte and the significant increase of neutrophil in severe COVID-19 patients. While, we did not observe significant differences of NLR between the WHO severity stage among COVID-19 patients of the IInd wave (September 2020-April 2021; n = 242). In these patients, the number of lymphocytes and neutrophils did not change significantly between patients of different severity subgroups. This difference likely depends on the steroids therapy that the patients of the IInd wave performed before hospitalization while most patients of the Ist wave were hospitalized soon after diagnosis. This is also confirmed by serum interleukin (IL)-6 and myeloperoxidase (MPO) that gradually increased with the disease stage in patients of the Ist wave, while such biomarkers (whose production is inhibited by steroids) did not show differences among patients of the IInd wave in different stages. Thus, the NLR could be tested at diagnosis in naïve patients before starting therapies.

We previously observed an increase of serum interleukins (IL) and a reduction of most lymphocyte subpopulations in hospitalized COVID-19 patients. Herein, we aimed to evaluate the changes in serum IL-6, IL-10, and IL-17A levels and cytometric lymphocyte profiles in 144 COVID-19 patients at admission and after one week, also in relation to steroid treatment before hospitalization. After one week of hospitalization, we found that: (i) total lymphocytes were increased in all patients; (ii) neutrophils and IL-6 were reduced in mild/moderate patients; (iii) B lymphocytes were increased in severe patients; (iv) T lymphocyte populations increased in mild/moderate patients. In the eight patients that died during hospitalization, total leukocytes increased while T, T helper, T cytotoxic, T regulatory, and NK lymphocytes showed a reducing trend in five of the eight patients. Even if seven days are too few to evaluate the adaptive immunity of patients, we found that the steroid therapy was associated with a reduced COVID-19 inflammation and cytokine activation only in patients with severe disease, while in patients with less severe disease, the steroid therapy seems to have immunosuppressive effects on lymphocyte populations, and this could hamper the antiviral response. A better knowledge of cytokine and lymphocyte alterations in each COVID-19 patient could be useful to plan better treatment with steroids or cytokine targeting.

Since the beginning of 2020, the new pandemic caused by SARS-CoV-2 and named coronavirus disease 19 (COVID 19) has changed our socio-economic life. In just a few months, SARS-CoV-2 was able to spread worldwide at an unprecedented speed, causing hundreds of thousands of deaths, especially among the weakest part of the population. Indeed, especially at the beginning of this pandemic, many reports highlighted how people, suffering from other pathologies, such as hypertension, cardiovascular diseases, and diabetes, are more at risk of severe outcomes if infected. Although this pandemic has put the entire academic world to the test, it has also been a year of intense research and many important contributions have advanced our understanding of SARS-CoV-2 origin, its molecular structure and its mechanism of infection. Unfortunately, despite this great effort, we are still a long way from fully understanding how SARS-CoV-2 dysregulates organismal physiology and whether the current vaccines will be able to protect us from possible future pandemics. Here, we discuss the knowledge we have gained during this year and which questions future research should address.

Background ZFYVE19 (Zinc Finger FYVE-Type Containing 19) mutations have most recently been associated to a novel type of high gamma-glutamyl transpeptidase (GGT), non-syndromic, neonatal-onset intrahepatic chronic cholestasis possibly associated to cilia dysfunction. Herein, we report a new case with further studies of whole exome sequencing (WES) and immunofluorescence in primary cilia of her cultured fibroblasts which confirm the observation. Results A now 5-year-old girl born to clinically healthy consanguineous Moroccan parents was assessed at 59 days of life due to severe cholestatic jaundice with increased serum bile acids and GGT, and preserved hepatocellular synthetic function. Despite fibrosis/cirrhosis and biliary ducts proliferation on liver biopsy suggested an extrahepatic biliary obstacle, normal intra-operatory cholangiography excluded biliary atresia. Under choleretic treatment, she maintained a clinically stable anicteric cholestasis but developped hyperlipidemia. After exclusion of the main causes of cholestasis by multiple tests, abnormal concentrations of sterols and WES led to a diagnosis of hereditary sitosterolemia (OMIM #618666), likely unrelated to her cholestasis. Further sequencing investigation revealed a homozygous non-sense mutation (p.Arg223Ter) in ZFYVE19 leading to a 222 aa truncated protein and present in both heterozygous parents. Immunofluorescence analysis of primary cilia on cultured skin fibroblasts showed a ciliary phenotype mainly defined by fragmented cilia and centrioles abnormalities. Conclusions Our findings are consistent with and expands the recent evidence linking ZFYVE19 to a novel, likely non-syndromic, high GGT-PFIC phenotype with neonatal onset. Due to the possible role of ZFYVE19 in cilia function and the unprecedented coexistence of a coincidental hereditary sterol disorder in our case, continuous monitoring will be necessary to substantiate type of liver disease progression and/or possible emergence of a multisystemic involvement. What mentioned above confirms that the application of WES in children with undiagnosed cholestasis may lead to the identification of new causative genes, widening the knowledge on the pathophysiology.

A sample of Roman glass found in Regio I, Insula 14, during the 1950’s Pompeii excavation was examined by Raman and Fourier transformed infrared spectroscopy, scanning electron microscopy, and inductively coupled plasma mass spectrometry. The analyzed specimen was selected based on its intense blue color and its well-preserved aspect. The purpose of the work was the chemical characterization of Pompeii’s glass in correlation to the actual knowledge of Roman glassmaking technology from the Mediterranean area. The results suggested that the Pompeii’s glass was a soda-lime-silica glass, but with a higher calcium content that, given the low content of lead, was used to stabilize the glass. The sample was in origin produced most likely as non-decolorized primary raw materials from eastern Mediterranean sites. Moreover, the intense blue color was related to the use mainly of cobalt, present in a weighty amount, and likely used as important coloring agents in the ancient secondary glass-making workshop.

A relationship is emerging between SARS-CoV-2 infections and ANCA-associated vasculitis (AAV) because: (i) the pulmonary involvement of COVID-19 may mimic that observed in patients with AAV; (ii) the two diseases may occur together; (iii) COVID-19 may trigger AAV. However, few cases of AAV have been identified so far in COVID-19 patients. To define the frequency of ANCA autoimmunity in patients with SARS-CoV-2 infection, we analyzed the serum ANCAs and the serum PR3 and MPO antigens by immunoassays in 124 adult patients with a diagnosis of SARS-CoV-2 infection (16 were asymptomatic and 108 were hospitalized) and 48 control subjects. The serum ANCAs were significantly higher in the hospitalized patients compared with either the controls or the asymptomatic patients and increased with the progression of the COVID-19 severity. After one week of hospitalization, the values were significantly lower. In contrast, no differences emerged among the controls, asymptomatic and hospitalized patients for the PR3 and MPO serum levels. None of the patients had clinical signs of AAV with the exception of a severe pulmonary involvement. Further studies are necessary to define whether the increase in the serum ANCAs might mask subclinical vasculitis in a percentage of patients with SARS-CoV-2 infection or it is an epiphenomenon of SARS-CoV-2 infection with no clinical manifestations.

Background: Cystic fibrosis (CF) is one of the most common inherited diseases. It is characterised by a severe decline in pulmonary function associated with metabolic perturbations and an increased production of inflammatory cytokines. The key role of physical activity (PA) in improving the health status of CF patients and reducing lung function decline has recently been demonstrated. This study evaluated interleukin-6 (IL-6) and tumour necrosis factor α (TNFα) expression in two subgroups of CF patients classified based on PA. Methods: We selected 85 CF patients, half of them regularly undertook supervised PA in the three years leading up to the study and half of them were not physically active. Patients were analysed for serum IL-6 and TNFα levels using enzyme-linked immunosorbent assays. Results: We found that the expression levels of IL-6 and TNFα differed in terms of their regulation by PA. In particular, TNFα levels negatively correlated with FEV1% decrease/year and FEV1% decrease (p = 0.023 and p = 0.02, respectively), and positively correlated with serum fasting glucose (p = 0.019) in PA CF patients. In contrast, in the NPA subgroup, TNFα levels were positively correlated with IL-6 (p = 0.001) and negatively correlated with adiponectin (p = 0.000). In addition, multiple logistic regression analysis confirmed that PA is an independent modulator of the inflammatory state. Conclusions: PA modulates inflammatory processes in CF patients by regulating the secretion of pro-inflammatory cytokines and thus ameliorating lung function. Our data show that PA is a useful complementary strategy in the management of CF and that TNFα may be a marker of these effects of PA.

The defective mucociliary clearance due to CFTR malfunctioning causes predisposition to the colonization of pathogens responsible for the recurrent inflammation and rapid deterioration of lung function in patients with cystic fibrosis (CF). This has also a profound effect on the lung microbiome composition, causing a progressive reduction in its diversity, which has become a common characteristic of patients affected by CF. Although we know that the lung microbiome plays an essential role in maintaining lung physiology, our comprehension of how the microbial components interact with each other and the lung, as well as how these interactions change during the disease’s course, is still at an early stage. Many challenges exist and many questions still to be answered, but there is no doubt that manipulation of the lung microbiome could help to develop better therapies for people affected by CF.

Introduction Congenital chloride diarrhea (CLD) is a rare autosomal recessive disorder characterized by watery diarrhea with a high level of fecal Cl-, metabolic alkalosis, and electrolyte alterations. Several intestinal and extraintestinal complications and even death can occur. An optimal knowledge of the clinical features and best therapeutic strategies is mandatory for an effective management. Methods Articles published between 1 January 1965 and 31 December 2019, reported in PUBMED and EMBASE, were evaluated for a systematic review analyzing four categories: anamnestic features, clinical features, management, and follow-up strategies. Results Fifty-seven papers reporting information on 193 CLD patients were included. The most common anamnestic features were positive family anamnesis for chronic diarrhea (44.4%), consanguinity (75%), polyhydramnios (98.3%), preterm delivery (78.6%), and failure to pass meconium (60.7%). Mean age at diarrhea onset was 6.63 days. Median diagnostic delay was 60 days. Prenatal diagnosis, based on molecular analysis, was described in 40/172 (23.3%). All patients received NaCl/KCl-substitutive therapy. An improvement of diarrhea during adulthood was reported in 91.3% of cases. Failure to thrive (21.6%) and chronic kidney disease (17.7%) were the most common complications. Conclusions This analysis of a large population suggests the necessity of better strategies for the management of CLD. A close follow-up and a multidisciplinary approach is mandatory to manage this condition characterized by heterogeneous and multisystemic complications. Impact In this systematic review, we describe data regarding anamnestic features, clinical features, management, and follow-up of CLD patients obtained from the largest population of patients ever described to date. The results of our investigation could provide useful insights for the diagnostic approach and the management of this condition.

About 50% of patients with cystic fibrosis (CF) have sinonasal complications, which include inferior turbinate hypertrophy (NTH) and/or nasal polyposis (NP), and different degrees of lung disease, which represents the main cause of mortality. Monitoring of sinonasal disease requires complex instrumental procedures, while monitoring of lung inflammation requires invasive collection of bronchoalveolar lavage fluid. The aim of this study was to investigate the associations between salivary cytokines levels and CF-related airway diseases. Salivary biochemical parameters and cytokines, i.e., interleukin-6 (IL-6), IL-8, and tumor necrosis factor alpha (TNF-&alpha, ), were analyzed in resting saliva from healthy subjects and patients with CF. Patients with CF showed significantly higher levels of salivary chloride, IL-6, IL-8, and TNF-&alpha, and lower calcium levels than healthy subjects. Among patients with CF, IL-6 and IL-8 were significantly higher in patients with NTH, while TNF-&alpha, was significantly lower in patients with NP. A decreasing trend of TNF-&alpha, in patients with severe lung disease was also observed. On the other hand, we did not find significant correlation between cytokine levels and Pseudomonas aeruginosa or Stenotrophomonas maltophilia colonization. These preliminary results suggest that salivary IL-6 and IL-8 levels increase during the acute phase of sinonasal disease (i.e., NTH), while the end stages of pulmonary disease and sinonasal disease (i.e., NP) show decreased TNF-&alpha, levels

The clinical manifestation of cystic fibrosis (CF) is heterogeneous also in patients with the same cystic fibrosis transmembrane regulator (CFTR) genotype and in affected sibling pairs. Other genes, inherited independently of CFTR, may modulate the clinical manifestation and complications of patients with CF, including the severity of chronic sinonasal disease and the occurrence of chronic Pseudomonas aeruginosa colonization. The T2R38 gene encodes a taste receptor and recently its functionality was related to the occurrence of sinonasal diseases and upper respiratory infections. We assessed the T2R38 genotype in 210 patients with CF and in 95 controls, relating the genotype to the severity of sinonasal disease and to the occurrence of P. aeruginosa pulmonary colonization. The frequency of the PAV allele i.e., the allele associated with the high functionality of the T2R38 protein, was significantly lower in i) CF patients with nasal polyposis requiring surgery, especially in patients who developed the complication before 14 years of age; and ii) in CF patients with chronic pulmonary colonization by P. aeruginosa, especially in patients who were colonized before 14 years of age, than in control subjects. These data suggest a role for T2R38 as a novel modifier gene of sinonasal disease severity and of pulmonary P. aeruginosa colonization in patients with CF.

Because of the progression of genetics and genomics, the demand for prenatal diagnosis (PD) for inherited genetic diseases has increased. However, several incidental findings may emerge during PD, like misattributed paternity, the evidence of disease in a parent, and the possible misinterpretation of the results because of complex alleles or de novo mutations that have several implications. In a retrospective observational study on all the couples referred to our Medical School (1993&ndash, 2018) for PD of genetic inherited diseases (n = 1502), we selected the cases of PD for cystic fibrosis (CF, n = 239) and hemophilia A and B (HA, HB, n = 47), revising all incidental findings previously mentioned. We found one case in which a technical error led to PD of carrier in two siblings that were born affected by CF, four cases of misattributed paternity, eight cases of asymptomatic parents revealed as affected by CF transmembrane regulator (CFTR)-related disorders, a case of a novel complex allele that could have caused the diagnosis of CF in a carrier fetus, and a case of a de novo mutation in a mother (already a carrier) that caused hemophilia in a child that PD had revealed as healthy. We present these conditions as clinical cases and discuss the technical, clinical, ethical, and legal aspects to be considered.

Systemic vascular damage with micro/macro-thrombosis is a typical feature of severe COVID-19. However, the pathogenesis of this damage and its predictive biomarkers remain poorly defined. For this reason, in this study, serum monocyte chemotactic protein (MCP)-2 and P- and E-selectin levels were analyzed in 204 patients with COVID-19. Serum MCP-2 and P-selectin were significantly higher in hospitalized patients compared with asymptomatic patients. Furthermore, MCP-2 increased with the WHO stage in hospitalized patients. After 1 week of hospitalization, MCP-2 levels were significantly reduced, while P-selectin increased in patients in WHO stage 3 and decreased in patients in WHO stages 5–7. Serum E-selectin was not significantly different between asymptomatic and hospitalized patients. The lower MCP-2 levels after 1 week suggest that endothelial damage triggered by monocytes occurs early in COVID-19 disease progression. MCP-2 may also predict COVID-19 severity. The increase in P-selectin levels, which further increased in mild patients and reduced in severe patients after 1 week of hospitalization, suggests that the inactive form of the protein produced by the cleavage of the active protein from the platelet membrane is present. This may be used to identify a subset of patients that would benefit from targeted therapies. The unchanged levels of E-selectin in these patients suggest that endothelial damage is less relevant.

Background Cystic fibrosis (CF) patients have reduced intestinal absorption of sterols and, despite enhanced endogenous synthesis, low plasma cholesterol. Lumacaftor/ivacaftor CFTR protein modulator therapy is used to improve the clinical outcome of CF patients homozygous for F508del mutation (homo-deltaF508). Aim of the study is to evaluate the cholesterol metabolism and hepatobiliary injury/function in adult homo-deltaF508 patients, before and after lumacaftor/ivacaftor treatment. Baseline parameters in homo-deltaF508 patients were compared to those in CF patients compound heterozygous for F508del mutation and another severe mutation (hetero-deltaF508). Methods Cholesterol metabolism was evaluated measuring plasma phytosterols and cholestanol, as intestinal absorption markers, and lathosterol, as liver biosynthesis marker. We quantified serum vitamin E, as nutritional marker. We evaluated liver injury by aspartate aminotransferase (AST) and alanine transaminase (ALT), biliary injury by γ-glutamyltransferase (γGT) and AP, and the liver function by bilirubin and albumin. Results Before the treatment, homo-deltaF508 patients (n = 20) had significantly lower cholesterol and vitamin E compared to hetero-deltaF508 (n = 20). Lumacaftor/ivacaftor treatment caused: 1) further reduction of cholesterol; 2) lathosterol reduction, suggesting a normalization of endogenous synthesis; 3) cholestanol and vitamin E increment, indicating an improvement of lipid digestion/absorption. Vitamin E difference (after-before treatment) was positively associated to treatment months. Alkaline phosphatase was also reduced. Conclusions These data suggest an effect of lumacaftor/ivacaftor on cholesterol metabolism and enterohepatic flux in CF patients. However, lumacaftor/ivacaftor does not promote the increase of cholesterol serum concentration that on the contrary declines. Further studies are needed to research the real mechanism causing this reduction.

Background Patients with cystic fibrosis (CF) have a reduced intestinal absorption of cholesterol and in a preliminary study we observed differences in plasma sterol profile between patients with pancreatic sufficiency (PS) and those with pancreatic insufficiency (PI). Therefore, we hypothesized that the sterol analysis may contribute to study the digestion and absorption state of lipids in patients with CF. To this aim we evaluated plasma sterols in a significant number of adult patients with CF in relation to the pancreatic status. Methods Beside cholesterol, we measured phytosterols and lathosterol as markers of intestinal absorption and hepatic biosynthesis, respectively, by gas-chromatography in plasma of adult CF patients with pancreatic sufficiency (PS-CF, n = 57), insufficiency (PI-CF, n = 97) and healthy subjects (control group, CT, n = 71). Results PI-CF patients had cholesterol and phytosterols levels significantly lower than PS-CF and CT (p −10) suggesting a reduced intestinal absorption of sterols related to PI. Instead, lathosterol was significantly higher in PI-CF patients than PS-CF and CT (p Conclusions A plasma sterols profile may be useful to evaluate the metabolic status of lipids in adult patients with CF and could help to manage the pancreatic enzyme supplementation therapy.

Cerebrotendinous Xanthomatosis (CTX) is an autosomal recessive defect of the alternative pathway of bile acid biosynthesis, due to the deficiency of mitochondrial cytochrome P450 sterol 27-hydroxylase enzyme encoded by CYP27A1. The deficit of sterol 27-hydroxylase raises cholestanol in plasma and tissues of affected patients. Although there is a marked variability of signs, symptoms, severity and age of onset, the main clinical manifestations of CTX include chronic diarrhea, bilateral cataract, tendon xanthomas and neurological dysfunction. Herein, we report the clinical, biochemical and molecular characterization of a Caucasian female affected by CTX diagnosed at 28years. The patient’s clinical history revealed neurological and behavioral manifestations already at fifth year of life, following by bilateral cataract and chronic diarrhea without xanthomas. At diagnosis, an involvement of the cervical spinal cord was also observed on MRI. Sterols profile analysis in plasma and red blood cell membranes showed very high cholestanol levels. CYP27A1 sequencing revealed a new variant (e.g., c.850_854delinsCTC) at homozygous status. The follow-up after 5months of chenodeoxycholic acid treatment showed a decrease of plasma cholestanol of 64%. After 1 year, the patient showed normalization of bowel function, reduction of risk of falls, improvement of cognitive function although brain and spine MRI and other instrumental examinations remained unchanged. This case highlights the variability of the CTX phenotype that makes it difficult to reach an early diagnosis. Biochemical and/or molecular screening of CTX should be taken into account to early start the pharmacological treatment limiting neurological damages.

Cystic fibrosis (CF) is a genetic disease characterized by progressive decline of lung function and chronic airway inflammation. Adipose tissue, through adiponectin and leptin, exerts several effects on energy metabolism and inflammatory processes. This study evaluated the levels of adiponectin and leptin in adult healthy subjects, in patients with CF and their correlation with long-term physical activity. CF patients were divided into two groups (sedentary versus active) based on their regular physical activity over 3 years. Anthropometric and serum biochemical profiles of CF patients and controls were evaluated and compared. Total serum adiponectin and leptin levels were measured by ELISA; adiponectin oligomeric profiles were analysed by western blot. Adiponectin levels were significantly higher while leptin levels were lower in patients with CF than in healthy controls. Furthermore, adiponectin was significantly lower in active compared to sedentary CF (p=0.047), while leptin was slightly increased in active compared to sedentary CF. In addition, C-reactive protein levels were significantly lower in active than in sedentary CF patients (p=0.048). Interestingly, only in the active group adiponectin levels were inversely correlated with forced expiratory volume (FEV) 1% decrease/year and FEV1% decrease. Moreover, adiponectin levels negatively correlated with lipid profiles. Our findings indicated that regular, long-term physical activity in CF improves respiratory function, metabolism, and inflammation status. These improvements in patients’ conditions are associated with immunometabolic processes involving adiponectin, leptin, and C-reactive protein. Therefore, we propose that both adipokines may be a useful biomarker in the evaluation of metabolic and inflammatory status in patients with CF.

Quantitation of paint powders of ancient wall paintings is often hindered by the calcite contamination during samples withdrawal. To overcome this problem, a new approach was explored based on the mechanical pulverization of the paint powder followed by the evaluation of its true concentration, namely binders, pigments, and decaying compounds, from the comparison of the calcite FT-IR peak area at 2510 cm−1 with that of the corresponding underlying calcite used as calibrator. After extraction of the pulverized paint powder with polar and nonpolar solvents, liquid chromatography, gas chromatography with flame ionization detection, and gas chromatography–mass spectrometry were used to estimate the free amino acids, and fatty acids profiles. Compared to our previous investigation, the results obtained showed a better yield of the extracted organic materials as mg/kg of powder and also a qualitative improvement of the lipids profile.

This study aims to investigate cholesterol metabolism in a mouse model with cystic fibrosis (CF) by the comparison of affected homozygous versuswild type(WT) mice. In particular, we evaluated the effects of a diet enriched with cholesterol in both mice groups in comparison with the normal diet. To this purpose, beyond serum and liver cholesterol, we analyzed serum phytosterols as indirect markers of intestinal absorption of cholesterol, liver lathosterol as indirect marker ofde novocholesterol synthesis, liver cholestanol (a catabolite of bile salts synthesis) and the liver mRNA levels ofLDL receptor(LDLR),3-hydroxy-3-methylglutaryl-CoA reductase(HMG-CoAR),acyl CoA:cholesterol acyl transferase 2(ACAT2),cytochrome P450 7A1(CYP7A1) andtumor necrosis factor alpha(TNFα). CF mice showed lower intestinal absorption and higher liver synthesis of cholesterol than WT mice. In WT mice, the cholesterol supplementation inhibits the synthesis of liver cholesterol and enhances its catabolism, while in CF mice we did not observe a reduction ofLDLRandHMG-CoARexpression (probably due to an altered feed-back), causing an increase of intracellular cholesterol. In addition, we observed a further increase (5-fold) inTNFαmRNA levels. This preliminary study suggests that in CF mice there is a vicious circle in which the altered synthesis/secretion of bile salts may reduce the digestion/absorption of cholesterol. As a result, the liver increases the biosynthesis of cholesterol that accumulates in the cells, triggering inflammation and further compromising the metabolism of bile salts.

Purpose: The mechanisms involved in the coordination of muscle activity are not completely known: to investigate adaptive changes in human motor cortex Transcranial magnetic stimulation (TMS) was often used. The sport models are frequently used to study how the training may affect the corticospinal system excitability: Karate represents a valuable sport model for this kind of investigations for its high levels of coordination required to athletes. This study was aimed at examining possible changes in the resting motor threshold (rMT) and in the corticospinal response in karate athletes, and at determining whether athletes are characterized by a specific value of rMT. Methods: We recruited 25 right-handed young karate athletes and 25 matched non-athletes. TMS was applied to primary motor cortex (M1). Motor evoked potential (MEP) were recorded by two electrodes placed above the first dorsal interosseous (FDI) muscle. We considered MEP latencies and amplitudes at rMT, 110% of rMT, and 120% of rMT. Results: The two groups were similar for age (p > 0.05), height (p > 0.05) and body mass (p > 0.05). The TMS had a 70-mm figure-of-eight coil and a maximum output of 2.2 T, placed over the left motor cortex. During the stimulation, a mechanical arm kept the coil tangential to the scalp, with the handle at 45° respect to the midline. The SofTaxic navigator system (E.M.S. Italy, www.emsmedical.net) was used in order to correctly identifying and repeating the stimulation for every subject. Compared to non-athletes, athletes showed a lower resting motor threshold (p < 0.001). Furthermore, athletes had a lower MEP latency (p < 0.001) and a higher MEP amplitude (p < 0.001) compared to non-athletes. Moreover, a ROC curve for rMT was found significant (area: 0.907; sensitivity 84%, specificity 76%). Conclusions: As the main finding, the present study showed significant differences in cortical excitability between athletes and non-athletes. The training can improve cortical excitability inducing athletes' modifications, as demonstrated in rMT and MEP values. These finding support the hypothesis that the sport practice determines specific brain organizations in relationship with the sport challenges.

In recent decades, biotechnology produced a growth of knowledge on the causes and mechanisms of metabolic diseases that have formed the basis for their study, diagnosis and treatment. Unfortunately, it is well known that the clinical features of metabolic diseases can manifest themselves with very different characteristics and escape early detection. Also, it is well known that the prognosis of many metabolic diseases is excellent if diagnosed and treated early. In this editorial we briefly summarized two groups of inherited metabolic diseases, the defects of cholesterol biosynthesis and those of bile acids. Both groups show variable clinical manifestations but some clinical signs and symptoms are common in both the defects of cholesterol and bile acids. The differential diagnosis can be made analyzing sterol profiles in blood and/or bile acids in blood and urine by chromatographic techniques (GC-MS and LC-MS/MS). Several defects of both biosynthetic pathways are treatable so early diagnosis is crucial. Unfortunately their diagnosis is made too late, due either to the clinical heterogeneity of the syndromes (severe, mild and very mild) that to the scarcity of scientific dissemination of these rare diseases. Therefore, the delay in diagnosis leads the patient to the medical observation when the disease has produced irreversible damages to the body. Here, we highlighted simple clinical and laboratory descriptions that can potentially make you to suspect a defect in cholesterol biosynthesis and/or bile acids, as well, we suggest appropriate request of the laboratory tests that along with common clinical features can help to diagnose these defects.

Hepatocellular carcinoma (HCC) is a very aggressive neoplasia requiring early and accurate diagnosis to improve patient outcomes with timely treatment. The liver is also very frequently colonized by metastases, and the most frequent differential diagnosis is HCC against intrahepatic cholangiocarcinoma or metastatic adenocarcinoma. Metabolomics is a powerful tool for identification of altered biomarkers in cancer, and to evaluate the efficacy of drug treatments. Here we analyzed by HILIC-MS/MS methylated arginines, basic amino acids (Arg, Cit, Orn), and their ratios in the extracts of primary HCC tissues, liver metastases from colorectal carcinoma (MET), cirrhotic related hepatitis-C-virus (CIR), and non-cirrhotic normal liver (NT) adjacent tissues. We found high levels of Arg (p

The use of Fourier transform infrared spectromicroscopy and mass spectrometry (MS) allowed us to characterize the composition of polar and non-polar binders present in sporadic wall paint fragments taken from Pompeii's archaeological excavation. The analyses of the polar and non-polar binder components extracted from paint powder layer showed the presence of amino acids, sugars, and fatty acids but the absence of proteinaceous material. These results are consistent with a water tempera painting mixture composed of pigments, flours, gums, and oils and are in agreement with those obtained from a simulated wall paint sample made for mimicking an ancient "a secco" technique. Notably, for the first time, we report the capability to discriminate by tandem MS the presence of free amino acids in the paint layer. © 2012 Springer-Verlag.

Background Although the pigment composition of Pompeian wall paintings has been the object of several studies, a comprehensive characterization of paint binder components is still lacking. This work aimed investigated at a molecular level the binder composition differences among wall paintings belonging to different periods of Pompeii's history. Analytical investigations were performed on representative samples of the first, second, third, and fourth painting styles excavated from the house of Marcus Fabius Rufus (Insula Occidentalis). The application of sensitive experimental methodologies was complemented by historical knowledge to gain insight in painting techniques and materials used by Pompeian artists. Results Fourier transform infrared spectroscopy and Raman spectroscopy were used to investigate the organic components and pigments present in powders obtained from samples of the four painting styles. No proteinaceous components were detected in the samples with liquid chromatography-electrospray ionization-hybrid quadrupole/time-of-flight mass spectrometry. Liquid chromatography, gas chromatography with flame-ionization detection, and gas chromatography–mass spectrometry of polar and non-polar components extracted from powders were used to evaluate and compare the free amino acids, sugars, and fatty acids profiles. Conclusions Pigments and natural products (lipids, gums and wheat flours) were the main components of all samples. This supports the hypothesis that artists likely used water tempera for Pompeian wall paintings.

Since the early 1960s, dried blood spots (DBS) on filter paper have been used in clinical applications. The first key milestone in the use of DBS was the screening of phenylketonuria and other inborn errors of metabolism using microbiological and enzymatic analytical methods. 20 years after its introduction, advanced mass spectrometers and new soft ionization techniques have permitted the coupling of liquid chromatography with MS and tandem MS (MS/MS) and since the 1990s, DBS analysis by LC-MS/MS expanded screening to many inborn errors of metabolism simultaneously. Recently, DBS-LC-MS/MS analysis has been used in other fields such as pharmacology, toxicology and forensic sciences. Today, new ambient ionization techniques, coupled to MS, directly desorb/ionize molecules from solid samples. This presents new opportunities for the in situ analysis of DBS. Most likely, ambient MS methods will be used to analyze DBS, increasing the clinical applications of MS within the next 10 years. © 2010 Future Science Ltd

Here is proposed a rapid and sensitive method involving atmospheric pressure thermal desorption chemical ionization mass spectrometry (APTDCI-MS) for specific laboratory screening of the Smith-Lemli-Opitz syndrome (SLOS), an inherited defect of cholesterol biosynthesis. Biochemical findings in the blood of SLOS patients are low cholesterol (Chol), high 7- and 8-dehydrocholesterol (DHCs) levels and high DHCs/Chol ratios. The APTDCI proposed method is able to ionize sterols for qualitative and quantitative analysis directly from dried plasma/blood spots. Critical APTDCI parameters - desolvation gas flow and temperature - were optimized analyzing Chol, 7-DHC and cholesteryl stearate standards spotted onto a glass slide acquiring the full scan spectra in positive ion mode. Chol levels in dried plasma spots of unaffected controls (n = 23) obtained by the proposed method were compared with those of the enzymatic method (y = 0.9166x + 0.3811; r = 0.8831) while Chol and DHCs of SLOS patients (n = 9) were compared with the gas chromatography flame ionization detection (GC-FID) method (y = 0.8214x + 0.7388; r = 0.8288). The APTDCI-MS method is also able to differentiate normal from SLOS samples directly analyzing whole blood and washed red cells spotted on paper. In conclusion, the intrinsic analytical high-throughput of APTDCI-MS method for sterol analysis could be useful to screen SLO syndrome. © 2010 The Royal Society of Chemistry