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Background Evidence suggests that LPA risk genotypes are a possible contributor to the clinical diagnosis of familial hypercholesterolemia (FH). This study aimed at determining the prevalence of LPA risk variants in adult individuals with FH enrolled in the Italian LIPIGEN (Lipid Transport Disorders Italian Genetic Network) study, with (FH/M+) or without (FH/M-) a causative genetic variant. Methods and Results An lp(a) [lipoprotein(a)] genetic score was calculated by summing the number risk-increasing alleles inherited at rs3798220 and rs10455872 variants. Overall, in the 4.6% of 1695 patients with clinically diagnosed FH, the phenotype was not explained by a monogenic or polygenic cause but by genotype associated with high lp(a) levels. Among 765 subjects with FH/M- and 930 subjects with FH/M+, 133 (17.4%) and 95 (10.2%) were characterized by 1 copy of either rs10455872 or rs3798220 or 2 copies of either rs10455872 or rs3798220 (lp(a) score ≥1). Subjects with FH/M- also had lower mean levels of pretreatment low-density lipoprotein cholesterol than individuals with FH/M+ (t test for difference in means between FH/M- and FH/M+ groups

Background and Aims Familial hypercholesterolemia (FH) is an autosomal dominant disease that leads to cardiovascular (CV) disease. Proprotein convertase subtilisin/kexin type 9 inhibitors (PCSK9-I) demonstrated efficacy in low-density lipoprotein cholesterol (LDL-C) reduction and in prevention of CV events. The aim of our study is to evaluate the relationship between LDL receptor (LDLR) mutations and response to PCSK9-I therapy. Methods and Results We evaluated total cholesterol (TC), LDL-C, high-density lipoprotein cholesterol (HDL-C) and triglycerides (TG) in consecutive patients with FH before PCSK9-I treatment and after 12 (T12w) and 36 (T36w) weeks of treatment. We evaluated LDL-C target achievement according to different mutations in LDLR. Eighty FH subjects (mean age:54±13.3 years), 39 heterozygous (He) with defective LDLR gene mutations, 30 He with null mutations and 11 compound-He or homozygotes (Ho) were recruited. At baseline, 69 subjects were under maximal lipid lowering therapy (MLLT) and 11 subjects had statin-intolerance. From baseline to T36w we observed an overall 51% reduction in LDL-C. We found no difference in LDL-C changes between subjects with He-defective mutation and He-null mutations both at T12w (p=1.00) and T36w (p=0.538). At T36w, LDL-C target was achieved in 59% of He-defective mutations subjects and in 36% of He-null mutations subgroup (p=0.069), whereas none of compound-He/Ho-FH achieved LDL-C target. Conclusions After 36 weeks there were no differences in response to PCSK9-I therapy between different groups of He-FH subjects. Response to PCSK9-I was significantly lower in carriers of compound-He/Ho mutations. Registration number for clinical trials: NCT04313270 extension.

Background Periprocedural myocardial infarction (MI) according to the Society for Cardiovascular Angiography and Interventions (SCAI) criteria has prognostic relevance among patients undergoing percutaneous coronary intervention (PCI). However, it is unclear whether the type of cardiac biomarker used for the diagnosis of periprocedural MI plays a role in terms of event frequency and outcomes. Objectives To compare the characteristics of SCAI periprocedural MI based on creatine kinase-myocardial band fraction (CK-MB) vs. high-sensitivity cardiac troponin (hs-cTn) in patients undergoing elective PCI. Methods and results Between 2017 and 2021, periprocedural MI was assessed in a prospective study. The primary clinical outcome of interest was all-cause death at 1-year follow-up. A total of 1010 patients undergoing elective PCI were included. SCAI periprocedural MI based on CK-MB vs. hs-cTnI occurred in 1.8 and 13.5% of patients, respectively. hs-cTnI periprocedural MI in the absence of concomitant CK-MB criteria was associated with lower rates of ancillary criteria, including angiographic, ECG, and cardiac imaging criteria. At 1-year follow-up, periprocedural MI defined by CK-MB (adjusted hazard ratio, HR, 4.27, 95% confidence intervals, CI, 1.23–14.8; P = 0.022) but not hs-cTnI (adjusted HR 2.04, 95% CI 0.94–4.45; P = 0.072) was associated with a higher risk of all-cause death. Hs-cTnI periprocedural MI was not predictive of death unless accompanied by CK-MB criteria (adjusted HR 4.64, 95% CI 1.32–16.31; P = 0.017). Conclusion In the setting of elective PCI, using hs-cTn instead of CK-MB resulted in a substantial increase in SCAI periprocedural MI events, which were not prognostically relevant in the absence of concurrent CK-MB elevations.

Familial Hypercholesterolemia (FH) is the most common genetic disease caused by variants in LDLR, APOB, PCSK9 genes; it is characterized by high levels of LDL-cholesterol and premature cardiovascular disease. We aim to perform a retrospective analysis of a genetically screened population (528 unrelated patients - 342 adults and 186 children) to evaluate the biochemical and clinical correlations with the different genetic statuses. Genetic screening was performed by traditional sequencing and some patients were re-analyzed by next-generation-sequencing. Pathogenic variants, mainly missense in the LDLR gene, were identified in 402/528 patients (76.1%), including 4 homozygotes, 17 compound heterozygotes and 1 double heterozygotes. A gradual increase of LDL-cholesterol was observed from patients without pathogenic variants to patients with a defective variant, to patients with a null variant and to patients with two variants. Six variants accounted for 51% of patients; a large variability of LDL-cholesterol was observed among patients carrying the same variant. The frequency of pathogenic variants gradually increased from unlikely FH to definite FH, according to the Dutch Lipid Clinic Network criteria. Genetic diagnosis can help prognostic evaluation of FH patients, discriminating between the different genetic statuses or variant types. Clinical suspicion of FH should be considered even if few symptoms are present or if LDL-cholesterol is only mildly increased. This article is protected by copyright. All rights reserved.

Aims Lomitapide is a lipid-lowering agent indicated as an adjunct therapy for adult homozygous familial hypercholesterolaemia (HoFH). This study evaluated the medium-term effectiveness and safety of lomitapide in a large cohort of HoFH patients in Europe. Methods and results In a multicentre retrospective, observational study including 75 HoFH patients treated with lomitapide in a real-world clinical setting from 9 European countries, low-density lipoprotein cholesterol (LDL-C) changes, adverse events (AEs), and major adverse cardiovascular events (MACE) were assessed. After a median 19 months (interquartile range 11–41 months) of treatment with a mean dosage of 20 mg of lomitapide. Low-density lipoprotein cholesterol decreased by 60%, from baseline 280.5 mg/dL (191.8–405.0 mg/dL) to 121.6 mg/dL (61.0–190.5 mg/dL). At the last visit, 32.0% of patients achieved LDL-C Conclusion In this medium-term real-world experience, lomitapide proved to be very effective in reducing LDL-C in HoFH. Gastrointestinal AEs were common, but liver safety was reassuring with no sign of increased risk of liver fibrosis. A signal of cardiovascular protection was also observed.

Backgroundand aim: Autosomal recessive hypercholesterolemia (ARH) is a rare autosomal recessive disorder of low-density lipoprotein (LDL) metabolism caused by pathogenic variants in the LDLRAP1 gene. Like homozygous familial hypercholesterolemia, ARH is resistant to conventional LDL-lowering medications and causes a high risk of atherosclerotic cardiovascular diseases (ASCVDs) and aortic valve stenosis. Lomitapide is emerging as an efficacious therapy in classical HoFH, but few data are available for ARH.Results: This is a subanalysis carried out on nine ARH patients included in the Pan-European Lomitapide Study. The age at starting lomitapide was 46 (interquartile range (IQR), 39.0–65.5) years, with a median treatment duration of 31.0 (IQR 14.0–40.5) months. At baseline, four (44.4%) patients had hypertension, one (11.1%) had diabetes mellitus, two (22.2%) were active smokers, and five (55.5%) reported ASCVD. The baseline LDL-C was 257.0 (IQR, 165.3–309.2) mg/dL. All patients were on statins plus ezetimibe, three were receiving Lipoprotein apheresis (LA), and one was also receiving proprotein convertase subtilisin/kexin type 9 inhibitors (PCSK9i). The addition of lomitapide (mean dose, 10 mg) resulted in the achievement of a median on-treatment LDL-C of 101.7 mg/dL (IQR, 71.3–138.3; 60.4% reduction from baseline), with a best LDL-C value of 68.0 mg/dL (IQR, 43.7–86.7; 73.5% reduction from baseline). During follow-up, one patient stopped both PCSK9i and LA. Recurrence of ASCVD events was reported in one patient. The median on-treatment aspartate transaminase and alanine transaminase values were 31.1 (IQR, 22.6–48.3) U/L and 31.1 (IQR, 27.2–53.8) U/L, respectively. Among six ARH patients with available fibroscan examination, liver stiffness values recorded at the last visit were within the normal range (median, 4.7 KPa; IQR, 3.6–5.3 KPa).Conclusion: Lomitapide is effective and safe in ARH therapy as well as in classical HoFH.

Pathology registers can be a useful tool to overcome obstacles in the identification and management of familial hypercholesterolemia since childhood. In 2018, the LIPIGEN pediatric group was constituted within the Italian LIPIGEN study to focus on FH subjects under 18 years. This work aimed at discussing its recent progress and early outcomes. Demographic, biochemical, and genetic baseline characteristics were collected, with an in-depth analysis of the genetic defects. The analysis was carried out on 1,602 children and adolescents (mean age at baseline 9.9 ± 4.0 years), and almost the whole cohort underwent the genetic test (93.3%). Overall, the untreated mean value of LDL-C was 220.0 ± 97.2 mg/dl, with an increasing gradient from subjects with a negative (N = 317; mean untreated LDL-C = 159.9 ± 47.7 mg/dl), inconclusive (N = 125; mean untreated LDL-C = 166.4 ± 56.5 mg/dl), or positive (N = 1,053; mean untreated LDL-C = 246.5 ± 102.1 mg/dl) genetic diagnosis of FH. In the latter group, the LDL-C values presented a great variability based on the number and the biological impact of involved causative variants. The LIPIGEN pediatric group represents one of the largest cohorts of children with FH, allowing the deepening of the characterization of their baseline and genetic features, providing the basis for further longitudinal investigations for complete details.

Summary Systematic insight into cellular dysfunction can improve understanding of disease etiology, risk assessment, and patient stratification. We present a multiparametric high-content imaging platform enabling quantification of low-density lipoprotein (LDL) uptake and lipid storage in cytoplasmic droplets of primary leukocyte subpopulations. We validate this platform with samples from 65 individuals with variable blood LDL-cholesterol (LDL-c) levels, including familial hypercholesterolemia (FH) and non-FH subjects. We integrate lipid storage data into another readout parameter, lipid mobilization, measuring the efficiency with which cells deplete lipid reservoirs. Lipid mobilization correlates positively with LDL uptake and negatively with hypercholesterolemia and age, improving differentiation of individuals with normal and elevated LDL-c. Moreover, combination of cell-based readouts with a polygenic risk score for LDL-c explains hypercholesterolemia better than the genetic risk score alone. This platform provides functional insights into cellular lipid trafficking and has broad possible applications in dissecting the cellular basis of metabolic disorders.

Little is known about the role of Lp(a) in the assessment of cardiovascular risk in the paediatric population. Trying to clarify the clinical relevance of Lp(a) in risk stratification, the aim of the study is to evaluate the association between Lp(a) plasma levels in children with familial hypercholesterolaemia (FH) and positive family history for premature cardiovascular disease (pCVD) in first- and second-degree relatives.653 Caucasian children and adolescents (334 females and 319 males), aged 2-17 years, with diagnosis of FH from a paediatric cohort included in the LIPIGEN Network, were selected. We compared family history of pCVD, lipid and genetic profile in two groups based on Lp(a) levels below or above 30 mg/dL. To determine the independent predictors of pCVD, a multivariate logistic regression was used, with all clinical characteristics and blood measurements as predictors.Subjects with Lp(a) 30 mg/dl more frequently reported positive family history of pCVD compared to subjects with Lp(a)≤30 mg/dl (69.90% vs 36.66%, p 0.0001), while did not show differences in terms of median [interquartile range] LDL-cholesterol level (153.00 [88.00 vs 164.50 [90.25] mg/dL, p = 0.3105). In the regression analysis, Lp(a) 30 mg/dl was an independent predictor of family history of pCVD. Comparing subjects with or without family history of pCVD, we reported significant differences for Lp(a) 30 mg/dl (46.25% vs 17.65%, p 0.0001), FH genetic mutation (50.48% vs 40.75%, p = 0,0157), as well as for LDL-cholesterol (p = 0.0013) and total cholesterol (p = 0.0101).Children/adolescents with FH and Lp(a) 30 mg/dl where more likely to have a positive family history of pCVD. Lp(a) screening in children and adolescents with FH may enhance risk assessment and help identify those subjects, children and relatives, at increased pCVD risk.

Abstract Background Homozygous familial hypercholesterolemia (HoFH) is a rare life-threatening condition that represents a therapeutic challenge. The vast majority of HoFH patients fail to achieve LDL-C targets when treated with the standard protocol, which associates maximally tolerated dose of lipid-lowering medications with lipoprotein apheresis (LA). Lomitapide is an emerging therapy in HoFH, but its place in the treatment algorithm is disputed because a comparison of its long-term efficacy versus LA in reducing LDL-C burden is not available. We assessed changes in long-term LDL-C burden and goals achievement in two independent HoFH patients’ cohorts, one treated with lomitapide in Italy (n = 30) and the other with LA in France (n = 29). Results The two cohorts differed significantly for genotype (p = 0.004), baseline lipid profile (p padj = 0.004). Yearly on-treatment LDL-C padj = 0.002). A trend towards fewer total cardiovascular events was observed in the Lomitapide than in the LA cohort. Conclusions In comparison with LA, lomitapide appears to provide a better control of LDL-C in HoFH. Further studies are needed to confirm this data and establish whether this translates into a reduction of cardiovascular risk.

Increased concentrations of B-type natriuretic peptide (BNP), N-terminal pro-B-type natriuretic peptide (NT-proBNP) and high-sensitivity troponin I (HsTnI) in COVID-19 patients have already been reported. The aim of this study is to evaluate which of these common markers of cardiac disease is the most useful predictor of fatal outcome in COVID-19 patients. One hundred and seventy-four patients affected with COVID-19 were recruited, and markers of cardiac disease and the clinical history of the patients were collected at admission in the infectious disease unit or intensive care unit. NT-proBNP, BNP and HsTnI values were higher in in-hospital non-surviving patients. Receiver operating characteristic (ROC) curve analysis of NT-proBNP, BNP and HsTnI was performed, with NT-proBNP (AUC = 0.951) and HsTnI (AUC = 0.947) being better performers (p = 0.01) than BNP (AUC = 0.777). Logistic regression was performed assessing the relation of HsTnI and NT-proBNP to fatal outcome adjusting for age and gender, with only NT-proBNP being significant. The population was then divided into two groups, one with higher NT-proBNP values at admission than the cut-off resulted from the ROC curve (511 ng/L) and a second one with lower values. The Kaplan–Meier analysis showed an absence of fatal outcome in the group of patients with NT-proBNP values lower than the cut-off (p &lt, 0.001). NT-proBNP proved to be the best prognostic tool for fatal outcome among markers of cardiac disease in COVID-19 patients.

Background and aims Familial hypercholesterolemia (FH) is a monogenic disease characterized by high levels of low-density lipoprotein cholesterol and premature atherosclerotic cardiovascular disease. FH is caused by loss of function mutations in genes encoding LDL receptor (LDLR), and Apolipoprotein B (APOB) or gain of function (GOF) mutations in proprotein convertase subtilisin/kexin type 9 (PCSK9). In this study, we identified a novel variant in PCSK9, p.(Arg499His), located in the C-terminal domain, in two unrelated FH patients from Spain and Italy. Methods We studied familial segregation and determined variant activity in vitro. Results We determined PCSK9 expression, secretion and activity of the variant in transfected HEK293 cells; extracellular activity of the recombinant p.(Arg499His) PCSK9 variant in HEK 293 and HepG2 cells; PCSK9 affinity to the LDL receptor at neutral and acidic pH; the mechanism of action of the p.(Arg499His) PCSK9 variant by co-transfection with a soluble construct of the LDL receptor and by determining total PCSK9 intracellular accumulation when endosomal acidification is impaired and when an excess of soluble LDLr is present in the culture medium. Our results show high LDL-C concentrations and FH phenotype in p.(Arg499His) carriers. In vitro functional characterization shows that p.(Arg499His) PCSK9 variant causes a reduction in LDLr expression and LDL uptake. An intracellular activity for this variant is also shown when blocking the activity of secreted PCSK9 and by inhibiting endosomal acidification. Conclusions We demonstrated that p.(Arg499His) PCSK9 variant causes a direct intracellular degradation of LDLr therefore causing FH by reducing LDLr availability.

SummarySystematic insight into cellular dysfunctions can improve understanding of disease etiology, risk assessment and patient stratification. We present a multiparametric high-content imaging platform enabling quantification of low-density lipoprotein (LDL) uptake and lipid storage in cytoplasmic droplets of primary leukocyte subpopulations. We validated this platform with samples from 65 individuals with variable blood LDL-cholesterol (LDL-c) levels, including familial hypercholesterolemia (FH) and non-FH subjects. We integrated lipid storage data into a novel readout, lipid mobilization, measuring the efficiency with which cells deplete lipid reservoirs. Lipid mobilization correlated positively with LDL uptake and negatively with hypercholesterolemia and age, improving differentiation of individuals with normal and elevated LDL-c. Moreover, combination of cell-based readouts with a polygenic risk score for LDL-c explained hypercholesterolemia better than the genetic risk score alone. This platform provides functional insights into cellular lipid trafficking from a few ml’s of blood and is applicable to dissect metabolic disorders, such as hypercholesterolemia.MotivationWe have limited information on how cellular lipid uptake and processing differ between individuals and influence the development of metabolic diseases, such as hypercholesterolemia. Available assays are labor intensive, require skilled personnel and are difficult to scale to higher throughput, making it challenging to obtain systematic functional cell-based data from individuals. To overcome this problem, we established a scalable automated analysis pipeline enabling reliable quantification of multiple cellular readouts, including lipid uptake, storage and mobilization, from different white blood cell populations. This approach provides new personalized insights into the cellular basis of hypercholesterolemia and obesity.Graphical Abstract

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Despite the progress in cardiovascular research, atherosclerosis still represents the main cause of death worldwide. Clinically, the diagnosis of Atherosclerotic Cardiovascular Disease (ASCVD) relies on imaging methodologies including X-ray angiography and computed tomography (CT), which however still fails in the identification of patients at high risk of plaque rupture, the main cause of severe clinical events as stroke and heart attack. Magnetic resonance imaging, which is characterized by very high spatial resolution, could provide a better characterization of atherosclerotic plaque (AP) anatomy and composition, aiding in the identification of “vulnerable” plaques. In this context, hydrogel matrices, which have been demonstrated able to boost relaxometric properties of Gd-based contrast agents (CAs) by the effect of Hydrodenticity, represent a valuable tool towards the precision imaging of ASCVD improving the performance of this class of CAs while reducing systemic toxicity. In particular, hydrogel nanoparticles encapsulating Gd-DTPA can further contribute to providing CA-specific accumulation in the AP by nanoparticle surface decoration triggering an active targeting of the AP with the overall effect of allowing an earlier and more accurate diagnosis. In this work, we tested crosslinked Hyaluronic Acid Nanoparticles (cHANPs) in the complex environment of human atherosclerotic plaque. In addition, the surface of cHANPs was decorated with the antibody anti-CD36 (Ab36-cHANPs) for the active targeting of AP-associated macrophages. Results demonstrate that the Hydrodenticity of cHANPs and Ab36-cHANPs is preserved in this complex system and, preliminarily, that interaction of these probes with the AP is present.

Introduction Familial hypercholesterolemia (FH) is an autosomal dominant disease that leads to premature cardiovascular disease (CAD). The availability of ab-PCSK9 has changed the approach to therapy. Purpose To evaluate the relationship between different types of mutations in LDLR gene and response to ab-PCSK9. Methods 73 FH patients, 33 women and 40 men (53.9±13. yrs), in primary prevention (N=46) and secondary prevention (N=27), were recruited. This sample included patients with mutations in LDLR gene: heterozygotes for missense mutations (N=31), for null mutations (N=31), compound heterozygotes or homozygotes (N=11). At baseline, the whole sample had a maximally tolerated lipid lowering therapy (MT-LLT) without ab-PCSK9; 16 patients had MT-LLTs intolerance. After 160 days with ab-PCSK9 therapy we evaluated the achievement of a goal (LDL-C Results After 160 days of therapy with ab-PCSK9 (45 patients on Alirocumab, 28 patients on Evolocumab) and MT-LLT, 29/73 patients (39.7%) of the whole sample achieve the goal of LDL-C. Of them 14/29 (48.2%) were in primary prevention, 15/29 (51.7%) in secondary prevention, no difference in achievement of the goal. We then evaluated the percent of patients achieving the goal of LDL-C: 15/31 (48.3%) patients with missense mutation and 14/31 (45.1%) patients with null mutation, no significant difference among groups; 0/11 compound heterozygotes or homozygotes; 3/16 (18.7%) MT-LLTs intolerance. The other main cardiovascular risk factors did not influence of the achievement the goal of LDL cholesterol. Conclusions Lack of correlation between type of mutation in heterozygous FH patients and ab-PCSK9 therapy response; response was significantly poorest in patients with compound heterozygosis or homozygosis mutation as compared to heterozygotes; the intolerance to MT-LLT was significant in the achievement of the goal of LDL-C. Different between guideline 2016 vs 2019 Funding Acknowledgement Type of funding source: None

Background and aim: Protein convertase subtilisin kexin type 9 (PCSK-9) inhibitors demonstrated efficacy in cholesterol reduction and in the prevention of cardiovascular events. We evaluated changes in lipid profile and carotid stiffness in patients with familial hypercholesterolemia during 12 weeks of treatment with a PCSK-9 inhibitor, Evolocumab®. Methods and results: Patients with familial hypercholesterolemia starting a treatment with Evolocumab® were included. Total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), small dense LDL (assessed by LDL score) and carotid stiffness were evaluated before starting treatment with Evolocumab® and during 12 weeks of treatment. Twenty-five subjects were enrolled (52% males, mean age 51.5 years). TC and LDL-C were reduced of 38% and 52%, respectively during treatment, with LDL score reduced of 46.1%. In parallel, carotid stiffness changed from 8.8 (IQR: 7.0–10.4) m/sec to 6.6 (IQR: 5.4–7.5) m/sec, corresponding to a median change of 21.4% (p < 0.001), with a significant increase in carotid distensibility (from 12.1, IQR: 8.73–19.3 kPA−1 × 10−3 at T0 to 21.8, IQR: 16.6–31.8 kPA−1 × 10−3 at T12w) corresponding to a median change of 62.8% (p < 0.001). A multivariate analysis showed that changes in LDL score were independently associated with changes in carotid stiffness (β = 0.429, p = 0.041). Conclusion: Small dense LDL reduction, as assessed by LDL score, is associated with changes in carotid stiffness in patients with familial hypercholesterolemia treated with Evolocumab®.

Familial hypercholesterolemia (FH) is an autosomal dominant disease caused by pathogenic variants in genes encoding for LDL receptor (LDLR), Apolipoprotein B and Proprotein convertase subtilisin/kexin type 9 (PCSK9). Among PCSK9 variants, only Gain-of- Function (GOF) variants lead to FH. Greater attention should be paid to the classification of variants as pathogenic. Two hundred sixty nine patients with a clinical suspect of FH were screened for variants in LDLR and the patients without pathogenic variants were screened for variants in PCSK9 and APOB. Functional characterization of PCSK9 variants was performed by assessment of protein secretion, of LDLR activity in presence of PCSK9 variant proteins as well as of the LDLR affinity of the PCSK9 variants. Among 81 patients without pathogenic variants in LDLR, 7 PCSK9 heterozygotes were found, 4 of whom were carriers of variants whose role in FH pathogenesis is still unknown. Functional characterization revealed that two variants (p.(Ser636Arg) and p.(Arg357Cys)) were GOF variants. In Conclusions, we demonstrated a GOF effect of 2 PCSK9 variants that can be considered as FH-causative variants. The study highlights the important role played by functional characterization in integrating diagnostic procedures when the pathogenicity of new variants has not been previously demonstrated.

Background Numerous reports have shown that psoriasis patients are more exposed to lipoprotein peroxidation and to a decrease in the activity of paraoxonase (PON)1, an antioxidant and anti-inflammatory enzyme. Thus, it has been suggested that malfunction of the antioxidant system and an increased production of reactive oxygen species drive immune inflammatory events, that result in progressive skin cell damage in patients with psoriasis. The PON protein family, including PON1, PON2 and PON3, is one of the most important endogenous defense mechanisms against oxidative stress. In the present study, we investigated PON gene expression in psoriasis and in cutaneous oxidative stress. Methods The study population included 10 patients affected by moderate-to-severe plaque psoriasis and 15 healthy donors who have undergone to plastic surgery, were used as control. Skin punch biopsies of lesional and non lesional psoriatic skin were performed for analysis of PON2 and PON3 gene expression. In addition, oxidation assays in ex vivo full-thickness healthy skin organ cultures were performed. Results No significant differences were observed between PON2 and PON3 gene expression in psoriatic lesional and non lesional skin compared with healthy controls. H2O2 treatment induced a significant decrease of PON2 and PON3 expression in ex vivo full-thickness healthy skin organ cultures; conversely the pretreatment of samples with the antioxidant reagent N-acetyl-L-cysteine (NAC) induced a significant increase. Interestingly, no significant alterations were reported for PON2 and PON3 expression in ex vivo full-thickness healthy skin organ cultures stimulated with IL-17. Conclusions Taken together our findings have revealed that a strong pro-oxidative activity is not effectively countered by antioxidant endogenous mechanisms both in psoriatic skin and in ex vivo experimental model.

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.

Introduction: atherosclerosis is one of the leading causes of death and morbidity worldwide. It consists in thedevelopment of plaques in the intima media layers of arteries due to lipid accumulation and oxidation, causingmassive inflammation. We aim to better understand the role of Galectin-3 (Gal-3) and Lipoprotein(a) [Lp(a)] aspossible peripheral markers of plaque presence. Methods: Gal-3 and Lp(a) were measured in plasma samples from 99 patients undergoing carotid endarterectomyand 78 healthy controls, by immunometric assays. Plaques were classified histologically, according to the AmericanHeart Association (AHA) guidelines as type Va (fibroatheroma), Vb (mainly calcific) and Vl (complicated lesion). Results: Gal-3 and Lp(a) plasma levels are higher in patients compared to controls [19.8 ng/mL (SD 5.8) vs 14.0ng/mL (3.6)], p

Galectin-3 (Gal-3) is a β-galactoside-binding protein belonging to the lectin family with pleiotropic regulatory activities and several physiological cellular functions, such as cellular growth, proliferation, apoptosis, differentiation, cellular adhesion, and tissue repair. Inflammation, tissue fibrosis and angiogenesis are the main processes in which Gal-3 is involved. It is implicated in the pathogenesis of several diseases, including organ fibrosis, chronic inflammation, cancer, atherosclerosis and other cardiovascular diseases (CVDs). This review aims to explore the connections of Gal-3 with cardiovascular diseases since they represent a major cause of morbidity and mortality. We herein discuss the evidence on the pro-inflammatory role of Gal-3 in the atherogenic process as well as the association with plaque features linked to lesion stability. We report the biological role and molecular mechanisms of Gal-3 in other CVDs, highlighting its involvement in the development of cardiac fibrosis and impaired myocardium remodelling, resulting in heart failure and atrial fibrillation. The role of Gal-3 as a prognostic marker of heart failure is described together with possible diagnostic applications to other CVDs. Finally, we report the tentative use of Gal-3 inhibition as a therapeutic approach to prevent cardiac inflammation and fibrosis.

Aims Familial Hypercholesterolemia (FH) is one of the most frequent dyslipidemias, the autosomal dominant form of which is primarily caused by mutations in the LDL receptor ( LDLR ), apolipoprotein B ( APOB ), and proprotein convertase subtilisin/kexin type 9 ( PCSK9 ) genes, although in around 20% of patients the genetic cause remains unidentified. Genetic testing has notably improved the identification of patients suffering from FH, the most frequent cause of which is the presence of mutations in the LDLR gene. Although more than 1200 different mutations have been identified in this gene, about 80% are recognized to be pathogenic. We aim to overview the current methods used to perform the functional characterization of mutations causing FH and to highlight the conditions requiring a functional characterization of the variant in order to obtain a diagnostic report. Data synthesis In the current review, we summarize the different types of functional assays – including their advantages and disadvantages – performed to characterize mutations in the LDLR , APOB and PCSK9 genes helping to better define their pathogenic role. We describe the evaluation of splicing alterations and two major procedures for functional characterization: 1. ex vivo methods, using cells from FH patients; 2. in vitro methods using cell lines. Conclusions Functional characterization of the LDLR , APOB and PCSK9 mutant genes associated with FH can be considered a necessary integration of its genetic diagnosis.

Homozygous familial hypercholesterolemia (HoFH), the severest form of familial hypercholesterolemia (FH), is characterized by very high LDL-cholesterol levels and a high frequency of coronary heart disease. The disease is caused by the presence of either a pathogenic variant at homozygous status or of two pathogenic variants at compound heterozygous status in the LDLR, APOB, PCSK9 genes. We retrospectively analyzed data of 23 HoFH patients (four children and 19 adults) identified during the genetic screening of 724 FH patients. Genetic screening was performed by sequencing FH causative genes and identifying large rearrangements of LDLR. Among the HoFH patients, four out of 23 (17.4%) were true homozygotes, whereas 19 out of 23 (82.6%) were compound heterozygotes for variants in the LDLR gene. Basal LDL-cholesterol was 12.9 &plusmn, 2.9 mmol/L. LDL-cholesterol levels decreased to 7.2 &plusmn, 1.8 mmol/L when treated with statin/ezetimibe and to 5.1 &plusmn, 3.1 mmol/L with anti-PCSK9 antibodies. Homozygous patients showed higher basal LDL-cholesterol and a poorer response to therapy compared with compound heterozygotes. Since 19 unrelated patients were identified in the Campania region (6,000,000 inhabitants) in southern Italy, the regional prevalence of HoFH was estimated to be at least 1:320,000. In conclusion, our results revealed a worse phenotype for homozygotes compared with compound heterozygotes, thereby highlighting the role of genetic screening in differentiating one genetic status from the other.

Background Familial hypercholesterolemia (FH) is a genetic disorder caused by mutations in genes involved in low-density lipoprotein (LDL) uptake (LDLR, APOB and PCSK9). Genetic diagnosis is particularly useful in asymptomatic children allowing for the detection of definite FH patients. Furthermore, defining their genetic status may be of considerable importance as the compound heterozygous status is much more severe than the heterozygous one. Our study aims at depicting the genetic background of an Italian pediatric population with FH focusing on the correlation between lipid profile and genetic status. Methods Out of 196 patients with clinically suspected FH (LDL-cholesterol [LDL-C] levels above 3.37 mmol/L, cholesterol level above 6.46 mmol/L in a first-degree relative or the presence of premature cardiovascular acute disease in a first/second-degree relative), we screened 164 index cases for mutations in the LDLR, APOB and PCSK9 genes. Results Patients with mutations (129/164) showed increased levels of LDL-C, 95th percentile-adjusted LDL-C and LDL/high-density lipoprotein (HDL) ratio and decreased levels of HDL-C, adjusted HDL-C. The association of the LDL/HDL ratio with the presence of mutations was assessed independently of age, (body mass index) BMI, parental hypercholesterolemia, premature coronary artery disease (CAD), triglycerides by multivariate logistic regression (odds ratio [OR]=1.701 [1.103–2.621], p=0.016). The LDL/HDL ratio gradually increased from patients without mutations to patients with missense mutations, null mutations and compound heterozygotes. Conclusions In conclusion, the LDL/HDL ratio proved to be a better parameter than LDL-C for discriminating patients with from patients without mutations across different genetic statuses.

Aim: Atherosclerosis is one of the leading causes of death and morbidity worldwide. Plaque instability is the most relevant predisposing factor for acute events. We aim to better understand the role of Galectin-3 (Gal-3) and Lipoprotein(a) (Lp(a)) in atherosclerotic process and to define their potential role as peripheral marker of plaque instability. Methods: Advanced human carotid plaques from 99 patients undergoing carotid endarterectomy were classified histologically according to American Heart Association (AHA) guidelines. Plaque complication (type VI) is defined by the presence of hemorrhage, ulceration or hematoma. Gal-3 and Lp(a) was measured in serum samples from patients and 17 healthy controls by immunometric assays. The quantification of Gal-3 in plaque was performed by immunohistochemistry. Results: Gal-3 and Lp(a)/LDL serum levels showed increased levels in patients compared with the control group (p¼0.0001 and p¼0.001 respectively). No correlations have been observed between Gal-3 serum levels and Gal-3 levels in atherosclerotic plaques. No differences were found between Gal-3 serum levels among the different plaques types, nor between complicated and uncomplicated plaques. At multivariate logistic regression the presence of complicated lesion or patient symptoms (surrogate marker of plaque instability) are not related to both Gal-3 or Lp(a)/LDL. Conclusions: Our data showed that Gal-3 and Lp(a) are good markers of advanced atherosclerotic plaque. The absence of differences among the different lesion types suggest that the increase of Gal-3 and Lp(a) is independent by the specific plaque features. Both Gal-3 and Lp(a)/LDL levels cannot be considered markers of plaque instability.

BACKGROUND: Severe hypertriglyceridemia is a rare disease characterized by triglyceride levels higher than 1000 mg/dl (11.3 mmol/L) and acute pancreatitis. The disease is caused by pathogenic variants in genes encoding lipoprotein lipase (LPL), apolipoprotein A5 (APOA5), apolipoprotein C2 (APOC2), glycosylphosphatidylinositol-anchored high density lipoproteinbinding protein 1 (GPIHBP1) and lipase maturation factor 1 (LMF1). OBJECTIVE: We aim to identify the genetic cause of severe hypertriglyceridemia and characterize the new variants in a patient with severe hypertriglyceridemia. METHODS: The proband was a male showing severe hypertriglyceridemia (triglycerides 1416 mg/dL, 16.0 mmol/L); proband’s relatives were also screened. Genetic screening included direct sequencing of the above genes and identification of large rearrangements in the LPL gene. Functional characterization of mutant LMF1 variants was performed by complementing LPL maturation in transfected Lmf1-deficient mouse fibroblasts. RESULTS: The proband and his affected brother were compound heterozygotes for variants in the LMF1 gene never identified as causative of severe hypertriglyceridemia c.[157delC;1351C>T];[410C>T], p.[(Arg53Glyfs*5)];[(Ser137Leu)]. Functional analysis demonstrated that the p.(Arg53Glyfs*5) truncation completely abolished and the p.(Ser137Leu) missense variant dramatically diminished the lipase maturation activity of LMF1. CONCLUSIONS: In addition to a novel truncating variant, we describe for the first time a missense variant functionally demonstrated affecting the lipase maturation function of LMF1. This is the first case in which compound heterozygous variants in LMF1 was functionally demonstrated as causative of severe hypertriglyceridemia.