Plasma proteins associate with carotid plaques and predict incident atherosclerotic cardiovascular events.

PURPOSE: Performing non-invasive carotid imaging is challenging, owing inter-operator variability and organizational barriers, but plasma proteomics can offer an alternative. We sought plasma proteins that associate with the presence of carotid plaques, their number and predict the incidence of clinically overt atherosclerotic cardiovascular events (ASCVD) above currently recognized risk factors in "apparently healthy" subjects. METHODS: We studied the plasma levels of 368 proteins in 664 subjects from the PLIC study, who underwent an ultrasound imaging screening of the carotids to check for the presence of plaques. We clustered, by artificial intelligence (A.I.), the proteins that associate with the presence, the number of plaques and that predict incident ASCVDs over 22 years (198 events were registered). FINDINGS: 299/664 subjects had at least 1 carotid plaque (1+) (77 with only one plaque, 101 with 2 plaques, 121 with ≥3 plaques (3+)). The remaining 365 subjects with no plaques acted as controls. 106 proteins were associated with 1+ plaques, but 97 proteins significantly predicted 3+ plaques only (AUC = 0.683 (0.601-0.785), p < 0.001), when considered alone. A.I. underscored 87 proteins that improved the performance of the classical risk factors both in detecting 3+ plaques (AUC = 0.918 (0.887-0.943) versus risk factors alone, AUC = 0.760 (0.716-0.801), p < 0.001) and in predicting the incident ASCVD (AUC = 0.739 (0.704-0.773) vs risk factors alone AUC = 0.559 (0.521-0.598), p < 0.001). The chemotaxis/migration of leukocytes and interleukins/cytokines signaling were biological pathways mostly represented by these proteins. DISCUSSION AND CONCLUSIONS: Plasma proteomics marks the number of carotid plaques and improve the prediction of incidence ASCVDs in apparently healthy subjects.

High dietary inflammatory index associates with inflammatory proteins in plasma.

BACKGROUND AND AIM: Unhealthy dietary habits and highly caloric foods induce metabolic alterations and promote the development of the inflammatory consequences of obesity, insulin resistance, diabetes and cardiovascular diseases. Describing an inflammatory effect of diet is difficult to pursue, owing lacks of standardized quali-quantitative dietary assessments. The Dietary Inflammatory Index (DII) has been proposed as an estimator of the pro- or anti-inflammatory effect of nutrients and higher DII values, which indicate an increased intake of nutrients with pro-inflammatory effects, relate to an increased risk of metabolic and cardiovascular diseases and we here assessed whether they reflect biologically relevant plasmatic variations of inflammatory proteins. METHODS: In this cross-sectional study, seven days dietary records from 663 subjects in primary prevention for cardiovascular diseases were analyzed to derive the intake of nutrients, foods and to calculate DII. To associate DII with the Normalized Protein eXpression (NPX), an index of abundance, of a targeted panel of 368 inflammatory biomarkers (Olink™) measured in the plasma, we divided the population by the median value of DII (1.60 (0.83-2.30)). RESULTS: 332 subjects with estimated DII over the median value reported a higher intake of saturated fats but lower intakes of poly-unsaturated fats, including omega-3 and omega-6 fats, versus subjects with estimated dietary DII below the median value (N = 331). The NPX of 61 proteins was increased in the plasma of subjects with DII > median vs. subjects with DII < median. By contrast, in the latter group, we underscored only 3 proteins with increased NPX. Only 23, out of these 64 proteins, accurately identified subjects with DII > median (Area Under the Curve = 0.601 (0.519-0.668), p = 0.035). CONCLUSION: This large-scale proteomic study supports that higher DII reflects changes in the plasmatic abundance of inflammatory proteins. Larger studies are warranted to validate.

Real-World Effectiveness of PCSK9 Inhibitors in Reducing LDL-C in Patients With Familial Hypercholesterolemia in Italy: A Retrospective Cohort Study Based on the AIFA Monitoring Registries.

Background Information on the real-world use of proprotein convertase subtilisin kexin 9 inhibitors (PCKS9is) in familial hypercholesterolemia are limited. We evaluated the pattern of prescription and the long-term efficacy of alirocumab and evolocumab in Italian patients with familial hypercholesterolemia in clinical practice. Methods and Results The data set for analysis was extracted from the PCKS9i Italian Medicines Agency (AIFA) registry and included 2484 patients with heterozygous familial hypercholesterolemia (HeFH) and 62 patients with homozygous familial hypercholesterolemia (HoFH) who were prescribed PCKS9is from February 2017 to December 2021. As the follow-up schedules were not prespecified and could vary, persistence and adherence as well as low-density lipoprotein cholesterol (LDL-C) changes during 2 years of treatment were analyzed in a final cohort of 1299 patients with familial hypercholesterolemia. At baseline, 53.8% of patients with HeFH and 69.4% of patients with HoFH were receiving maximally tolerated lipid-lowering therapies, while 45.9% of patients with HeFH and 30.7% of patients with HoFH reported statin intolerance; mean LDL-C was 197.7±52.3 mg/dL in HeFH and 252.0±106.2 mg/dL in HoFH. The 6-month persistence and adherence to therapy were >85%, and LDL-C reduction reached 58.6% (to 79.7 mg/dL) in HeFH and 57.6% (to 95.1 mg/dL) in HoFH after 24 months of treatment. The European Atherosclerosis Society/European Society of Cardiology LDL-C goals were achieved in 43.3% of patients with HeFH and 37.5% of patients with HoFH. Conclusions PCKS9i prescribed to patients with familial hypercholesterolemia in clinical practice showed LDL-C-lowering efficacy similar to that observed in controlled trials. However, 2 of 5 HeFH cases and 2 of 6 HoFH cases achieved the recommended LDL-C goals. The full achievement of European Atherosclerosis Society/European Society of Cardiology LDL-C goals should require a lower threshold for PCKS9i initiation and a combination of multiple therapies.

Apolipoprotein E isoforms differentially affect LCAT-dependent cholesterol esterification.

BACKGROUND AND AIMS: LCAT esterifies cholesterol in both HDL (α-activity) and apoB-containing lipoproteins (ß-activity). The main activator of LCAT ß-activity is apoE, which in humans exists in 3 main different isoforms (E2, E3 and E4). Here, to gather insights into the potential role of LCAT in apoB-containing lipoprotein metabolism, we investigated the ability of apoE isoforms to promote LCAT-mediated cholesterol esterification. METHODS: We evaluated the plasma cholesterol esterification rate (CER) in 311 individuals who express functional LCAT and either apoE2, apoE3, or apoE4 and in 28 individuals who also carried LCAT mutations causing selective loss of LCAT α-activity (Fish-Eye Disease (FED)-causing mutations). The association of carrier status with CER was determined using an adjusted linear regression model. The kinetic of LCAT activity towards reconstituted HDLs (rHDLs) containing each apoE isoform was determined using the Michaelis-Menten model. RESULTS: Plasma CER was ∼20% higher in apoE2 carriers compared to apoE3 carriers, and ∼30% higher in apoE2 carriers compared to apoE4 carriers. After adjusting for age, sex, total cholesterol, HDL-C, apoA-I, apoB, chronic kidney disease diagnosis, zygosity, and LCAT concentration, CER remained significantly different among carriers of the three apoE isoforms. The same trend was observed in carriers of FED-causing mutations. rHDLs containing apoE2 were associated with a lower affinity but higher maximal esterification rate, compared to particles containing apoE3 or apoE4. CONCLUSION: The present results suggest that the apoE2 isoform is associated with a higher LCAT-mediated cholesterol esterification. This observation may contribute to the characterization of the peculiar functional properties of apoE2.

Clinical Approach in the Management of Paediatric Patients with Familial Hypercholesterolemia: A National Survey Conducted by the LIPIGEN Paediatric Group.

Detection and treatment of patients with familial hypercholesterolemia (FH) starting from childhood is fundamental to reduce morbidity and mortality. The activity of National realities such as the LIPIGEN (LIpid transPort disorders Italian GEnetic Network) Paediatric Group, founded in 2018, is a milestone in this context. The aim of this exploratory survey, conducted in October 2021 among Italian lipid clinics included in the LIPIGEN Paediatric Group, was to investigate the current clinical approach in the management and treatment of paediatric patients with suspected FH. A digital questionnaire composed of 20 questions investigating nutritional treatment and nutraceutical and pharmacological therapy for children and adolescents with FH was proposed to the principal investigators of 30 LIPIGEN centres. Twenty-four centres responded to the section referring to children aged < 10 years and 30 to that referring to adolescents. Overall, 66.7% of children and 73.3% of adolescents were given lipid-lowering nutritional treatment as the first intervention level for at least 3-4 months (29.2% and 23.3%) or 6-12 months (58.3% and 53.3%). Nutraceuticals were considered in 41.7% (regarding children) and 50.0% (regarding adolescents) of the centres as a supplementary approach to diet. Lipid-lowering drug therapy initiation was mainly recommended (91.7% and 80.0%). In 83.3% of children and 96.7% of adolescents, statins were the most frequently prescribed drug. We highlighted several differences in the treatment of paediatric patients with suspected FH among Italian centres; however, the overall approach is in line with the European Atherosclerosis Society (EAS) recommendations for FH children and adolescents. We consider this survey as a starting point to reinforce collaboration between LIPIGEN centres and to elaborate in the near future a consensus document on the management of paediatric patients with suspected FH so as to improve and uniform detection, management, and treatment of these patients in our country.

Corrigendum: The role of registers in increasing knowledge and improving management of children and adolescents affected by familial hypercholesterolemia: The LIPIGEN pediatric group.

[This corrects the article DOI: 10.3389/fgene.2022.912510.].

Neutrophil aging exacerbates high fat diet induced metabolic alterations.

BACKGROUND: High fat diet (HFD) chronically hyper-activates the myeloid cell precursors, but whether it affects the neutrophil aging is unknown. PURPOSE: We characterized how HFD impacts neutrophil aging, infiltration in metabolic tissues and if this aging, in turn, modulates the development of metabolic alterations. We immunophenotyped neutrophils and characterized the metabolic responses in physiology (wild-type mice, WT) and in mice with constitutively aged neutrophils (MRP8 driven conditional deletion of CXCR4; herein CXCR4fl/flCre+) or with constitutively fresh neutrophils (MRP8 driven conditional deletion of CXCR2; CXCR2fl/flCre+), following 20 weeks of HFD feeding (45 % kcal from fat). FINDINGS: After 20 weeks HFD, the gluco-metabolic profile of CXCR4fl/flCre+ mice was comparable to that of WT mice, while CXCR2fl/flCre+ mice were protected from metabolic alterations. CXCR4fl/flCre+ infiltrated more, but CXCR2fl/flCre+ neutrophils infiltrated less, in liver and visceral adipose tissue (VAT). As consequence, while CXCR4fl/flCre+ resulted into hepatic "suicidal" neutrophils extracellular traps (NETs) and altered immune cell architecture in VAT, CXCR2fl/flCre+ promoted proresolutive hepatic NETs and reduced accumulation of pro-inflammatory macrophages in VAT. In humans, higher plasma levels of Cxcl12 (CXCR4 ligand) correlated with visceral adiposity while higher levels of Cxcl1 (the ligand of CXCR2) correlated with indexes of hepatic steatosis, adiposity and metabolic syndrome. CONCLUSIONS: Neutrophil aging might contribute to the development of HFD induced metabolic disorders.

The effect of the 2019 ESC/EAS dyslipidaemia guidelines on low-density lipoprotein cholesterol goal achievement in patients with acute coronary syndromes: The ACS EuroPath IV project.

AIMS: To evaluate the effect of the ESC/EAS 2019 dyslipidaemia guidelines on patient management of lipid-lowering therapy in patients with acute coronary syndrome (ACS), through a survey designed to compare post-ACS patient management in 2022 with that in 2018. METHODS: Online questionnaires focused on lipid profile and medications were used to gather data from 2650 ACS patients in 6 European countries, treated between March-June 2022 (ACS EuroPath IV survey). These data were compared with data collected from 2650 patients who participated in the ACS EuroPath I survey (conducted in 2018). RESULTS: Lipid testing was performed in 90% of patients and was done sooner after admission in 2022 versus 2018 (mean 1.4 vs 1.7 days). Increased testing for non-HDL-C, lipoprotein(a), and ApoB was observed over time. At discharge, most patients (≥90%) were receiving lipid-lowering therapy. Prescribing patterns differed, with a higher proportion of patients receiving statin plus ezetimibe combination therapy in 2022 versus 2018 (34% vs 13%). LDL-C levels were lower in 2022 versus 2018 at admission and at 1st, 2nd and 3rd post-discharge follow-up points. More patients achieved low-density lipoprotein cholesterol (LDL-C) goals in 2022 versus 2018 at the first follow-up (average 14 vs 16 weeks since discharge; <70 mg/dL [1.8 mmol/L]: 34% vs 20%; <55 mg/dL [1.4 mmol/L]: 18% vs 10%) and at subsequent follow-up points. CONCLUSION: LDL-C goal achievement has improved since the release of the 2019 guidelines, but lipid management in post-ACS patients remains suboptimal.

The low-density lipoprotein receptor-mTORC1 axis coordinates CD8+ T cell activation.

Activation of T cells relies on the availability of intracellular cholesterol for an effective response after stimulation. We investigated the contribution of cholesterol derived from extracellular uptake by the low-density lipoprotein (LDL) receptor in the immunometabolic response of T cells. By combining proteomics, gene expression profiling, and immunophenotyping, we described a unique role for cholesterol provided by the LDLR pathway in CD8+ T cell activation. mRNA and protein expression of LDLR was significantly increased in activated CD8+ compared to CD4+ WT T cells, and this resulted in a significant reduction of proliferation and cytokine production (IFNγ, Granzyme B, and Perforin) of CD8+ but not CD4+ T cells from Ldlr -/- mice after in vitro and in vivo stimulation. This effect was the consequence of altered cholesterol routing to the lysosome resulting in a lower mTORC1 activation. Similarly, CD8+ T cells from humans affected by familial hypercholesterolemia (FH) carrying a mutation on the LDLR gene showed reduced activation after an immune challenge.

Oral Porphyromonas gingivalis and Fusobacterium nucleatum Abundance in Subjects in Primary and Secondary Cardiovascular Prevention, with or without Heterozygous Familial Hypercholesterolemia.

BACKGROUND: Low-grade chronic inflammation, promoted by dysbiosis of the gut and oral microbiota, has been shown to contribute to individual susceptibility to atherosclerotic cardiovascular disease (ASCVD). High oral Porphyromonas gingivalis (Pg) and lower Fusobacterium nucleatum (Fn) concentrations have been associated with clinical and experimental atherosclerosis. We assessed oral Pg and Fn abundance in very high-risk patients with previously diagnosed ASCVD, with or without heterozygous familial hypercholesterolemia (HeFH), in subjects with HeFH in primary prevention and in healthy subjects. METHODS: In this cross-sectional study, 40 patients with previously diagnosed ASCVD (10 with genetically proven HeFH, and 30 without FH), 26 subjects with HeFH in primary prevention, and 31 healthy subjects were selected to quantify oral Pg and Fn abundance by qPCR and assess oral health status. RESULTS: Compared to healthy subjects, patients with previously diagnosed ASCVD showed greater Pg abundance (1101.3 vs. 192.4, p = 0.03), but similar Fn abundance. HeFH patients with ASCVD had an even greater Pg abundance than did non-HeFH patients and healthy subjects (1770.6 vs. 758.4 vs. 192.4, respectively; p = 0.048). No differences were found in the levels of Pg and Fn abundance in HeFH subjects in primary prevention, as compared to healthy subjects. CONCLUSIONS: Greater oral Pg abundance is present in very high-risk patients with previously diagnosed ASCVD, with or without FH, suggesting a potential relationship with CV events. Future studies will assess the predictive value of Pg abundance measurement in ASCVD risk stratification.

Pancreatic PCSK9 controls the organization of the ß-cell secretory pathway via LDLR-cholesterol axis.

BACKGROUND: Cholesterol is central to pancreatic ß-cell physiology and alterations of its homeostasis contribute to ß-cell dysfunction and diabetes. Proper intracellular cholesterol levels are maintained by different mechanisms including uptake via the low-density lipoprotein receptor (LDLR). In the liver, the proprotein convertase subtilisin/kexin type 9 (PCSK9) routes the LDLR to lysosomes for degradation, thus limiting its recycling to the membrane. PCSK9 is also expressed in the pancreas and loss of function mutations of PCSK9 result in higher plasma glucose levels and increased risk of Type 2 diabetes mellitus. Aim of this study was to investigate whether PCSK9 also impacts ß-cells function. METHODS: Pancreas-specific Pcsk9 null mice (Pdx1Cre/Pcsk9 fl/fl) were generated and characterized for glucose tolerance, insulin release and islet morphology. Isolated Pcsk9-deficient islets and clonal ß-cells (INS1E) were employed to characterize the molecular mechanisms of PCSK9 action. RESULTS: Pdx1Cre/Pcsk9 fl/fl mice exhibited normal blood PCSK9 and cholesterol levels but were glucose intolerant and had defective insulin secretion in vivo. Analysis of PCSK9-deficient islets revealed comparable ß-cell mass and insulin content but impaired stimulated secretion. Increased proinsulin/insulin ratio, modifications of SNARE proteins expression and decreased stimulated­calcium dynamics were detected in PCSK9-deficient ß-cells. Mechanistically, pancreatic PCSK9 silencing impacts ß-cell LDLR expression and cholesterol content, both in vivo and in vitro. The key role of LDLR is confirmed by the demonstration that LDLR downregulation rescued the phenotype. CONCLUSIONS: These findings establish pancreatic PCSK9 as a novel critical regulator of the functional maturation of the ß-cell secretory pathway, via modulation of cholesterol homeostasis.

Targeted Plasma Proteomics to Predict the Development of Carotid Plaques.

BACKGROUND: Cardiovascular risk stratification in primary prevention is a clinical challenge. We recently identified a large set of circulating proteins improving the risk prediction for cardiovascular events. We now evaluate which of these proteins predicts the development of subclinical carotid atherosclerosis (SCA) in primary cardiovascular prevention. METHODS: Three hundred sixty-eight proteins were quantified, by proximity extension assay, from the plasma collected at basal visit from 586 subjects without previous cardiovascular events and without preclinical atherosclerosis. These subjects were reevaluated 11 years after median follow-up (10-12) in a longitudinal observational analysis, to assess the development of SCA, defined as the formation of focal lesion in any carotid tract and detected by carotid ultrasound at basal visit and after follow-up. Common carotid (intima-media thickness [IMT]) was also measured by ultrasound during the same follow-up to identify subjects with faster common carotid intima-media thickness (IMT) progression (increase IMT)>1.3 mm in the common carotid tract). RESULTS: The variation of 68 proteins predicted SCA development and, among them, higher levels of PIgR2 (polymeric immunoglobulin receptor), chemokine (C-C motif) ligand 18, CA1 (carbonic anhydrase 1), Fc gamma receptor IIa and reduced MMP10 (matrix metallopeptidase 10), GT (gastrotropin), IL7R (interleukin 7 receptor) were the most predictive for SCA development. These 7 proteins improved the sensitivity and the specificity for SCA development versus risk factors (age, sex, overweight, hypertension, low HDL-cholesterol, high triglyceride); area under the curve: 0.747 ([0.707-0.784] versus 0.620 [0.577-0.663]; P<0.001). Vice versa, 25 proteins (not in common with the previous 68) predicted faster common carotid IMT progression. Among them, increased IL7D (interleukin 7), chemokine (C-X-C motif) ligand 1, and reduced TNFS13B (TNF superfamily member 13b) significantly increased the sensitivity and the specificity to predict faster common carotid IMT progression as compared with same risk factors (area under the curve: 0.719 [0.680-0.756] versus 0.569 [0.527-0.610]; P<0.001). CONCLUSIONS: A new set of circulating proteins have been identified that may be considered as markers of preclinical atherosclerosis development. The difference of the protein identified to predict SCA versus IMT progression may reflect different etiological factors.

The Role of Registers in Increasing Knowledge and Improving Management of Children and Adolescents Affected by Familial Hypercholesterolemia: the LIPIGEN Pediatric Group.

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.

Inclisiran: How Widely and When Should We Use It?

PURPOSE OF REVIEW: Plasma levels of LDL cholesterol (LDL-C) are causally associated with cardiovascular risk. Reducing LDL-C results in a decreased incidence of cardiovascular events, proportionally to the absolute reduction in LDL-C. The inhibition of proprotein convertase subtilisin kexin 9 (PCSK) is a highly effective and safe approach to reducing LDL-C levels. In this review, we discuss the available data on the efficacy and safety of inclisiran, a siRNA targeting PCSK9 and propose a clinical profile for the patients who can benefit the most from this approach. RECENT FINDINGS: Inclisiran is a small interfering RNA targeting the mRNA of PCSK9 specifically in the liver, owing to the conjugation with triantennary N-acetylgalactosamine. Randomized clinical trials have shown that inclisiran provides robust and durable reductions of PCSK9 and LDL-C levels, with a dosing schedule of once every 6 months after the initial and 3-month doses. These effects are consistent in different categories of patients, including patients with atherosclerotic cardiovascular disease and/or risk equivalent or patients with heterozygous familial hypercholesterolaemia. Ultimately the administration schedule may improve patients' compliance given also the favourable safety profile of the drug. Completion of ongoing outcome clinical trials will provide information on both the expected clinical benefit and the safety of inclisiran administered for longer.

Lipoprotein(a) and family history for cardiovascular disease in paediatric patients: A new frontier in cardiovascular risk stratification. Data from the LIPIGEN paediatric group.

BACKGROUND AND AIMS: 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. METHODS: 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. RESULTS: 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). CONCLUSIONS: 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.

Evaluation of Factors Associated With Appropriate Drug Prescription and Effectiveness of Informative and Educational Interventions-The EDU.RE.DRUG Project.

Background: EDU.RE.DRUG study is a prospective, multicentre, open-label, parallel-arm, controlled, pragmatic trial directed to general practitioners (GPs) and their patients. Methods: The study data were retrieved from health-related administrative databases of four local health units (LHUs) of Lombardy and four LHUs in Campania. According to the LHUs, the GPs/patients were assigned to (A) intervention on both GPs (feedback reports about appropriate prescribing among their patients and online courses) and patients (flyers and posters on proper drug use), (B) intervention on GPs, (C) intervention on patients, and (D) no intervention (control arm). A set of appropriate prescribing indicators (potential drug-drug interactions [pDDIs], potential and unnecessary therapeutic duplicates [pTDs], and inappropriate prescriptions in the elderly [ERD-list]) were measured at baseline and after the intervention phase. The effectiveness of the intervention was evaluated estimating the absolute difference in percentages of selected indicators carrying out linear random-intercept mixed-effect models. Results: A cohort of 3,586 GPs (2,567 in intervention groups and 1,019 in the control group) was evaluated. In Campania, the mean pre-intervention percentage of patients with at least one pDDI was always greater than 20% and always lower than 15% in Lombardy. The pre-post difference was quite heterogeneous among the LHUs, ranging from 1.9 to -1.4 percentage points. The mean pre-intervention percentage of patients with pTDs ranged from 0.59 to 2.1%, with slightly higher values characterizing Campania LHUs. The magnitude of the pre-post difference was very low, ranging from -0.11 to 0.20. In Campania, the mean pre-intervention percentage of patients with at least one ERD criterium was considerably higher than in Lombardy (approximately 30% in Lombardy and 50% in Campania). The pre-post difference was again quite heterogeneous. The results from the models accounting for GP geographical belonging suggested that none of the interventions resulted in a statistically significant effect, for all the three indicators considered. Conclusion: The proposed strategy was shown to be not effective in influencing the voluntary changes in GP prescription performance. However, the use of a set of explicit indicators proved to be useful in quantifying the inappropriateness. Further efforts are needed to find more efficient strategies and design more tailored interventions.

Potentially Inappropriate Prescribing among Elderly Outpatients: Evaluation of Temporal Trends 2012-2018 in Piedmont, Italy.

Pharmacological intervention is one of the cornerstones in the treatment and prevention of disease in modern healthcare. However, a large number of drugs are often prescribed and used inappropriately, especially in elderly patients. We aimed at investigating the annual prevalence of potentially inappropriate prescriptions (PIPs) among older outpatients using administrative healthcare databases of the Piedmont Region (Italy) over a seven-year period (2012-2018). We included all Piedmont outpatients aged 65 years or older with at least one drug prescription per year. Polypharmacy and the prevalence of PIPs according to the ERD list explicit tool were measured on an annual basis. A range between 976,398 (in 2012) and 1,066,389 (in 2018) elderly were evaluated. Among them, the number of subjects with at least one PIP decreased from 418,537 in 2012 to 339,764 in 2018; the prevalence significantly reduced by ~25% over the study period. The stratified analyses by age groups and sex also confirmed the downward trend and identified several differences in the most prevalent inappropriately prescribed drugs. Overall, despite a reduction in PIP prevalence, one out of three older outpatients was still exposed to inappropriateness, highlighting the extensive need for intervention to improve prescribing.