Aceruloplasminemia: A Severe Neurodegenerative Disorder Deserving an Early Diagnosis

Aceruloplasminemia (ACP) is a rare, adult-onset, autosomal recessive disorder, characterized by systemic iron overload due to mutations in the Ceruloplasmin gene (CP), which in turn lead to absence or strong reduction of CP activity. CP is a ferroxidase that plays a key role in iron export from various cells, especially in the brain, where it maintains the appropriate iron homeostasis with neuroprotective effects. Brain iron accumulation makes ACP unique among systemic iron overload syndromes, e.g., various types of genetic hemochromatosis. The main clinical features of fully expressed ACP include diabetes, retinopathy, liver disease, and progressive neurological symptoms reflecting iron deposition in target organs. However, biochemical signs of the disease, namely a mild anemia mimicking iron deficiency anemia because of microcytosis and low transferrin saturation, but with "paradoxical" hyperferritinemia, usually precedes the onset of clinical symptoms of many years and sometimes decades. Prompt diagnosis and therapy are crucial to prevent neurological complications of the disease, as they are usually irreversible once established. In this mini-review we discuss some major issues about this rare disorder, pointing out the early clues to the right diagnosis, instrumental to reduce significant disability burden of affected patients

Urinary extracellular vesicles carry valuable hints through mRNA for the understanding of endocrine hypertension

Urinary extracellular vesicles (uEVs), released from cells of the urogenital tract organs, carry precious information about originating tissues. The study of molecules transported through uEVs such as proteins, lipids and nucleic acids provides a deeper understanding of the function of the kidney, an organ involved in the pathogenesis of hypertension and a target of hypertension-mediated organ damage. Molecules derived from uEVs are often proposed for the study of disease pathophysiology or as possible disease diagnostic and prognostic biomarkers. Analysis of mRNA loading within uEVs may be a unique and readily obtainable way to assess gene expression patterns of renal cells, otherwise achievable only by an invasive biopsy procedure. Interestingly, the only few studies investigating transcriptomics of hypertension-related genes through the analysis of mRNA from uEVs are inherent to mineralocorticoid hypertension. More specifically, it has been observed that perturbation in human endocrine signalling through mineralcorticoid receptors (MR) activation parallels changes of mRNA transcripts in urine supernatant. Furthermore, an increased copy number of uEVs-extracted mRNA transcripts of the 11β-hydroxysteroid dehydrogenase type 2 (HSD11B2) gene were detected among subjects affected by apparent mineralocorticoid excess (AME), a hypertension-inducing autosomal recessive disorder due to a defective enzyme function. Moreover, by studying uEVs mRNA, it was observed that the renal sodium chloride cotransporter (NCC) gene expression is modulated under different conditions related to hypertension. Following this perspective, we illustrate here the state of the art and the possible future of uEVs transcriptomics towards a deeper knowledge of hypertension pathophysiology and ultimately more tailored investigational, diagnostic-prognostic approaches

Not Just Arterial Damage: Increased Incidence of Venous Thromboembolic Events in Cardiovascular Patients With Elevated Plasma Levels of Apolipoprotein CIII

Background Apolipoprotein CIII (apo CIII ) is a crucial player in triglyceride-rich lipoprotein metabolism, but may also act pleiotropically, provoking inflammatory responses and stimulating coagulation. Elevated apo CIII plasma levels have been associated with increased activity of coagulation factors. Since these features of prothrombotic diathesis are linked with venous thromboembolism ( VTE ), we hypothesized that apo CIII plays a role in VTE . Methods and Results We recorded nonfatal VTE events in 1020 patients (age 63.3\ub111.4 years; 29.1% women) with or without coronary artery disease (79.1% with coronary artery disease and 20.9% without coronary artery disease) during a long follow-up. Complete plasma lipid and apolipoproteins were available for all patients. Forty-five patients (4.4%) experienced nonfatal VTE events during a median follow-up period of 144 months. Apo CIII plasma concentration at enrollment was higher in patients with VTE compared with patients without VTE (12.2 [95% CI, 11.10-13.5] mg/dL vs 10.6 [95% CI, 10.4-10.9] mg/dL, respectively; P=0.011). Patients with apo CIII levels above the median value (10.6 mg/dL) exhibited an increased risk of VTE (incidence rate, 6.0 [95% CI , 4.0-8.0] vs 1.8 [95% CI, 0.7-2.9] VTE events/1000 person-years; unadjusted hazard ratio [ HR ], 3.42 [95% CI , 1.73-6.75]; P<0.001). This association was confirmed after adjustment for sex, age, coronary artery disease diagnosis, body mass index, hypertension, and anticoagulant treatment at enrollment ( HR , 2.66; 95% CI , 1.31-5.37 [ P=0.007]), with inclusion of lipid parameters in the Cox model (HR, 3.74; 95% CI , 1.24-11.33 [ P=0.019]), and even with exclusion of patients who died at follow-up ( HR, 3.92; 95% CI , 1.68-9.14 [ P=0.002]) or patients taking anticoagulants ( HR , 3.39; 95% CI , 1.72-6.69 [ P<0.001]). Conclusions Our results suggest that high plasma apo CIII concentrations may predict an increased risk of VTE in patients with cardiovascular disease

Surface plasmon resonance based on molecularly imprinted nanoparticles for the picomolar detection of the iron regulating hormone Hepcidin-25

Molecular modelling

High resolution preparation of monocyte-derived macrophages (MDM) protein fractions for clinical proteomics

<p>Abstract</p> <p>Background</p> <p>Macrophages are involved in a number of key physiological processes and complex responses such as inflammatory, immunological, infectious diseases and iron homeostasis. These cells are specialised for iron storage and recycling from senescent erythrocytes so they play a central role in the fine tuning of iron balancing and distribution. The comprehension of the many physiological responses of macrophages implies the study of the related molecular events. To this regard, proteomic analysis, is one of the most powerful tools for the elucidation of the molecular mechanisms, in terms of changes in protein expression levels.</p> <p>Results</p> <p>Our aim was to optimize a protocol for protein fractionation and high resolution mapping using human macrophages for clinical studies. We exploited a fractionation protocol based on the neutral detergent Triton X-114. The 2D maps of the fractions obtained showed high resolution and a good level of purity. Western immunoblotting and mass spectrometry (MS/MS analysis) indicated no fraction cross contamination. On 2D-PAGE mini gels (7 × 8 cm) we could count more than five hundred protein spots, substantially increasing the resolution and the number of detectable proteins for the macrophage proteome. The fractions were also evaluated, with preliminary experiments, using Surface Enhanced Laser Desorption Ionization Time of Flight Mass Spectrometry (SELDI-TOF-MS).</p> <p>Conclusion</p> <p>This relatively simple method allows deep investigation into macrophages proteomics producing discrete and accurate protein fractions, especially membrane-associated and integral proteins. The adapted protocol seems highly suitable for further studies of clinical proteomics, especially for the elucidation of the molecular mechanisms controlling iron homeostasis in normal and disease conditions.</p

Detection of urinary exosomal HSD11B2 mRNA expression: a useful novel tool for the diagnostic approach of dysfunctional 11β-HSD2-related hypertension

Apparent mineralocorticoid excess (AME) is an autosomal recessive disorder caused by the 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) enzyme deficiency, traditionally assessed by measuring either the urinary cortisol metabolites ratio (tetrahydrocortisol+allotetrahydrocortisol/tetrahydrocortisone, THF+5αTHF/THE) or the urinary cortisol/cortisone (F/E) ratio. Exosomal mRNA is an emerging diagnostic tool due to its stability in body fluids and its biological regulatory function. It is unknown whether urinary exosomal HSD11B2 mRNA is related to steroid ratio or the HSD11B2 662 C&gt;G genotype (corresponding to a 221 A&gt;G substitution) in patients with AME and essential hypertension (EH)

Novel protein-truncating variant in the APOB gene may protect from coronary artery disease and adverse cardiovascular events

Background and aims: Genetic testing is still rarely used for the diagnosis of dyslipidemia, even though gene variants determining plasma lipids levels are not uncommon.Methods: Starting from a a pilot-analysis of targeted Next Generation Sequencing (NGS) of 5 genes related to familial hypercholesterolemia (LDLR, APOB, PCSK9, HMGCR, APOE) within a cardiovascular cohort in subjects with extreme plasma concentrations of low-density lipoprotein (LDL) cholesterol, we discovered and characterized a novel point mutation in the APOB gene, which was associated with very low levels of apolipoprotein B (ApoB) and LDL cholesterol.Results: APOB c.6943 G &gt; T induces a premature stop codon at the level of exon 26 in the APOB gene and generates a protein which has the 51% of the mass of the wild type ApoB-10 0 (ApoB-51), with a trun-cation at the level of residue 2315. The premature stop codon occurs after the one needed for the synthesis of ApoB-4 8, allowing chylomicron production at intestinal level and thus avoiding potential nutritional impairments. The heterozygous carrier of APOB c.6943G &gt; T, despite a very high-risk profile encompassing all the traditional risk factors except for dyslipidemia, had normal coronary arteries by angiography and did not report any major adverse cardiovascular event during a 20-years follow-up, thereby obtaining advantage from the gene variant as regards protection against atherosclerosis, apparently without any metabolic retaliation.Conclusions: Our data support the use of targeted NGS in well-characterized clinical settings, as well as they indicate that.a partial block of ApoB production may be well tolerated and improve cardiovascular outcomes. (C) 2022 The Authors. Published by Elsevier B.V

Red Blood Cell Morphologic Abnormalities in Patients Hospitalized for COVID-19

Peripheral blood smear is a simple laboratory tool, which remains of invaluable help for diagnosing primary and secondary abnormalities of blood cells despite advances in automated and molecular techniques. Red blood cells (RBCs) abnormalities are known to occur in many viral infections, typically in the form of mild normo-microcytic anemia. While several hematological alterations at automated complete blood count (including neutrophilia, lymphopenia, and increased red cell distribution width—RDW) have been consistently associated with severity of COVID-19, there is scarce information on RBCs morphological abnormalities, mainly as case-reports or small series of patients, which are hardly comparable due to heterogeneity in sampling times and definition of illness severity. We report here a systematic evaluation of RBCs morphology at peripheral blood smear in COVID-19 patients within the first 72 h from hospital admission. One hundred and fifteen patients were included, with detailed collection of other clinical variables and follow-up. A certain degree of abnormalities in RBCs morphology was observed in 75 (65%) patients. Heterogenous alterations were noted, with spiculated cells being the more frequent morphology. The group with &gt;10% RBCs abnormalities had more consistent lymphopenia and thrombocytopenia compared to those without abnormalities or &lt;10% RBCs abnormalities (p &lt; 0.018, and p &lt; 0.021, respectively), thus underpinning a possible association with an overall more sustained immune-inflammatory “stress” hematopoiesis. Follow-up analysis showed a different mortality rate across groups, with the highest rate in those with more frequent RBCs morphological alterations compared to those with &lt;10% or no abnormalities (41.9%, vs. 20.5%, vs. 12.5%, respectively, p = 0.012). Despite the inherent limitations of such simple association, our results point out towards further studies on erythropoiesis alterations in the pathophysiology of COVID-19

ApoCIII glycoforms determination and proteomic analysis in plasma of coronary patients with different ApoCIII levels .

BACKGROUND:The aims of this study were:i) to analyze and quantify the apolipoproteinCIIIglycoforms characterized by none, one or two sialic acids,(having different lipoprotein lipase-LPLinhibitoryactivity), ii) to assess their relationship with LPL activity and ApoA-V in CAD patients,iii) to analyze some previously identified plasma proteins in relation to lipids status.METHODS: ApoCIII glycoforms in four groups of patients (from \u201cVerona Heart Study\u201d biobank,)classified according to the total plasma concentration of ApoCIII and different triglyceride (TG)levels, were analyzed by a classical (isoelectric focusing/western blotting) and by a shotgun MSapproach. LPL activity (Fluorescent assay) and ApoA-V concentration (ELISA assay) weredetermined, and their correlations with lipid metabolism parameters were analyzed. We alsovalidated by 2D-WB some plasma proteins previously identified by 2-DE analysis.RESULTS The distribution of the ApoCIII glycoforms in the selected groups of patients are relatedto the TG levels, particularly the mono-sialylated isoform (ApoCIII-1) prevails in patients with thehighest TG levels. Moreover, investigating the relation among ApoA-V level, LPL activity and theother parameters of lipid metabolism, we obtained useful information on their interplay as antiatherogenicfactors. ApoA-V concentration was found within the normal range, however higherthan previously reported. LPL activity measured as Vmax did not show significant correlations withApoCIII . Finally 2D-WB showed peculiar proteomic patterns of several proteins associated withelevated apoCIII plasma concentrations .CONCLUSIONS: As compared with the other ones, mono \u2013 sialylated isoform of apo CIII ispreferentially associated with TG levels. Samples with elevated levels of apoCIII are characterizedby specific proteomic patterns

Plasma proteomic analysis of patients with Coronary Artery Disease with different levels of apolipoprotein CIII

BACKGROUND: Coronary artery disease (CAD) is a multifactorial condition, involving alsogenetic factors. Among candidate genes associated with CAD risk there are lipoprotein lipase (LPL)gene and APOC3 gene. APOC3 encodes for the apolipoprotein CIII (ApoCIII) which acts as aninhibitor of LPL. Three different ApoCIII glycoforms (with different LPL inhibitory activity) -characterized by none, one or two sialic acids - have been described. Changes in the relativeabundance of these glycoforms have been observed in a variety of pathologies.The aim of this study was to analyze and quantify the apoCIII glycoforms and to assess theirrelationship with LPL activity and with the levels of the LPL activator apoA-V.METHODS: ApoCIII glycoforms in four groups of patients (from \u201cVerona Heart Study\u201d biobank,)classified according to the total plasma concentration of ApoCIII and different TG levels, wereanalyzed by a classical (isoelectric focusing/western blotting) and by a shotgun MS approach. LPLactivity (Fluorescent assay) and ApoA-V concentration (ELISA assay) were determined, and theircorrelations with lipid metabolism parameters were analyzed. We also validated by 2D-WB someproteins previously identified by 2-DE analysis.RESULTS Our results indicate that the distribution of the three ApoCIII glycoforms in the selectedgroups of patients are related to the TG levels, particularly the mono-sialylated isoform (ApoCIII-1)prevails in patients with the highest TG levels. A positive correlation between apoCIII and TGlevels as well a positive correlation between TG and more glycosylated ApoCIII were observed.Moreover, investigating the relation among ApoA-V level, LPL activity and the other parameters oflipid metabolism, we obtained useful information on their interplay as anti-atherogenic factor. Finally2D-WB patterns of serum proteins (i.e. Fibrinogen beta/gamma chain, Complement C3) confirmed thetrend in previously proteomics experiments.CONCLUSIONS: These data could provide important insights regarding the interaction amongapoCIII glycoforms, ApoA-V level and LPL activity in predicting cardiovascular risk inpredisposed individuals