Improvement Science Increases Routine Lipid Screening in General Pediatric Cardiology.

OBJECTIVE: To evaluate the effectiveness of patient education, physician counseling, and point-of-care (POC) testing on improving adherence to lipid screening national guidelines in a general pediatric cardiology practice (2017-2023). STUDY DESIGN: Regional primary care providers were surveyed regarding lipid screening practices. Key drivers were categorized (physician, patient, and system) with corresponding interventions. Pediatric cardiologists started offering lipid screening during regular visits by providing families with preventive cardiovascular education materials and lab phlebotomy testing. System redesign included educational posters, clinical intake protocol, physician counseling, electronic health record integration, and POC testing. Run charts and statistical process control charts measured screening rates and key processes. RESULTS: The primary care survey response rate was 32% (95/294); 97% supported pediatric cardiologists conducting routine lipid screening. Pediatric cardiology mean baseline lipid screening rate was 0%, increased to 7% with patient education, and to 61% after system redesign including POC testing. Screening rates among 1467 patients were similar across age groups (P = .98). More patients received lipid screening by POC (91.7%) compared with phlebotomy (8.3%). Lipid abnormalities detected did not differ by screening methodology (P = .49). CONCLUSION: Patient education, counseling, and POC testing improved adherence to national lipid screening guidelines, providing a possible model for primary care implementation.

Lipoprotein(a): a Case for Universal Screening in Youth.

PURPOSE OF REVIEW: Lipoprotein(a) has emerged as a strong independent risk factor for cardiovascular disease. Targeted screening recommendations for Lp(a) measurement exist for adults and youth known to be at high-risk. However, Lp(a) measurements are not included in universal screening guidelines in the US; hence, most families in the US with high Lp(a) levels who are at risk of future atherosclerotic heart disease, stroke, or aortic stenosis are not recognized. Lp(a) measurement included as part of routine universal lipid screening in youth would identify those children at risk of ASCVD and enable family cascade screening with identification and early intervention for affected family members. RECENT FINDINGS: Lp(a) levels can be reliably measured in children as young as two years of age. Lp(a) levels are genetically determined. The Lp(a) gene is inherited in a co-dominant fashion. Serum Lp(a) attains adult levels by two years of age and is stable for the lifetime of the individual. Novel therapies that aim to specifically target Lp(a) are in the pipeline, including nucleic acid-based molecules such as antisense oligonucleotides and siRNAs. Inclusion of a single Lp(a) measurement performed as part of routine universal lipid screening in youth (ages 9-11; or at ages 17-21) is feasible and cost effective. Lp(a) screening would identify youth at-risk of ASCVD and enable family cascade screening with identification and early intervention for affected family members.

Increasing lipid screenings in children 9-11 years old at a federally qualified health center - A quality improvement project.

BACKGROUND: Coronary artery disease (CAD) remains the leading cause of death in the U.S. Atherosclerotic changes leading to CAD begin in early childhood. Universal lipid screenings remain low nationwide despite the 2011 National Heart, Lung, and Blood Institute's (NHLBI) Expert Panel Guidelines. LOCAL PROBLEM: The aim of this quality improvement project was to examine the benefit of an educational intervention on the implementation of universal lipid screening guidelines within a federally qualified health center tasked with providing care to a high-risk population. INTERVENTION: An educational intervention was offered detailing the 2011 NHLBI guidelines. A total of seven medical providers participated in the intervention. METHOD: Following the intervention, a pre- and post- knowledge survey was given to assess improvement in knowledge. A retrospective chart review was performed to evaluate application to practice. RESULTS: The number of lipid screenings improved from 7.8% (n = 384) pre-intervention to 39.2% (n = 74) post intervention. There was a statistically significant increase in screenings post-intervention t (456) = 7.842, p = .000, two-tailed). CONCLUSION: More studies are needed to adequately identify the impact of universal screening guidelines on the health of both children and adults alike. PRACTICE IMPLICATIONS: Universal lipid screenings remain promising in early identification of CAD in the pediatric population. Interventions related to expanding the knowledge of healthcare providers, patients, and families are key to decreasing CAD morbidity and mortality.

Inhibition of dengue virus infection by 1-stearoyl-2-arachidonoyl-phosphatidylinositol in vitro.

Dengue fever is an acute febrile infectious disease caused by dengue virus (DENV). Despite the significant public health concerns posed by DENV, there are currently no effective anti-DENV therapeutic agents. To develop such drugs, a better understanding of the detailed mechanisms of DENV infection is needed. Both lipid metabolism and lipid synthesis are activated in DENV-infected cells, so we used lipid screening to identify potential antiviral lipid molecules. We identified 1-stearoyl-2-arachidonoyl-phosphatidylinositol (SAPI), which is the most abundant endogenous phosphatidylinositol (PI) molecular species, as an anti-DENV lipid molecule. SAPI suppressed the cytopathic effects induced by DENV2 infection as well as the replication of all DENV serotypes without inhibiting the entry of DENV2 into host cells. However, no other PI molecular species or PI metabolites, including lysophosphatidylinositols and phosphoinositides, displayed anti-DENV2 activity. Furthermore, SAPI suppressed the production of DENV2 infection-induced cytokines and chemokines, including C-C motif chemokine ligand (CCL)5, CCL20, C-X-C chemokine ligand 8, IL-6, and IFN-ß. SAPI also suppressed the TNF-α production induced by LPS stimulation in macrophage cells differentiated from THP-1 cells. Our results demonstrated that SAPI is an endogenous inhibitor of DENV and modulated inflammatory responses in DENV2-infected cells, at least in part via TLR 4.-Sanaki, T., Wakabayashi, M., Yoshioka, T., Yoshida, R., Shishido, T., Hall, W. W., Sawa, H., Sato, A. Inhibition of dengue virus infection by 1-stearoyl-2-arachidonoyl-phosphatidylinositol in vitro.

Co-translational formation and pharmacological characterization of beta1-adrenergic receptor/nanodisc complexes with different lipid environments.

G protein-coupled receptors are of key significance for biomedical research. Streamlined approaches for their efficient recombinant production are of pivotal interest in order to explore their intrinsic conformational dynamics and complex ligand binding behavior. We have systematically optimized the co-translational association and folding of G protein-coupled receptors with defined membranes of nanodiscs by cell-free expression approaches. Each optimization step was quantified and the ligand binding active fraction of the receptor samples could drastically be improved. The strategy was exemplified with a stabilized and a non-stabilized derivative of the turkey beta1-adrenergic receptor. Systematic lipid screens with preformed nanodiscs revealed that generation of ligand binding active conformations of the analyzed beta1-adrenergic receptors strongly depends on lipid charge, flexibility and chain length. The lipid composition of the nanodisc membranes modulates the affinities to a variety of ligands of both receptor derivatives. In addition, the thermostabilization procedure had a significant impact on specific ligand affinities of the receptor and abolished or reduced the binding of certain antagonists. Both receptors were highly stable after purification with optimized nanodisc membranes. The procedure avoids any detergent contact of the receptors and sample production takes less than two days. Moreover, even non-stabilized receptors can be analyzed and their prior purification is not necessary for the formation of nanodisc complexes. The established process appears therefore to be suitable as a new platform for the functional or even structural characterization of recombinant G protein-coupled receptors associated with defined lipid environments.

Referral Patterns and Cascade Screening for Familial Hypercholesterolemia in a Pediatric Lipid Clinic.

Charts of 42 children with familial hypercholesterolemia from a dyslipidemia clinic were reviewed for initial cholesterol screen indication and cascade screening results. Indications were universal screening (8/28 after guideline release, none before), family history (26/42), risk factor (5/42), and other (3/42). Cascade screening identified 63 relatives with unknown familial hypercholesterolemia.

Functional properties of cell-free expressed human endothelin A and endothelin B receptors in artificial membrane environments.

The human endothelin receptors are members of the rhodopsin class A of G-protein coupled receptors and key modulators of blood pressure regulation. Their functional in vitro characterization has widely been limited by the availability of high quality samples. We have optimized cell-free expression protocols for the human endothelin A and endothelin B receptors by implementing co-translational association approaches of the synthesized proteins with supplied liposomes or nanodiscs. Efficiency of membrane association and ligand binding properties of the receptors have systematically been studied in correlation to different membrane environments and lipid types. Ligand binding was analyzed by a number of complementary assays including radioassays, surface plasmon resonance and fluorescence measurements. High affinity binding of the peptide ligand ET-1 to both endothelin receptors could be obtained with several conditions and the highest Bmax values were measured in association with nanodiscs. We could further obtain the characteristic differential binding pattern of the two endothelin receptors with a panel of selected agonists and antagonists. Two intrinsic properties of the functionally folded endothelin B receptor, the proteolytic processing based on conformational recognition as well as the formation of SDS-resistant complexes with the peptide ligand ET-1, were observed with samples obtained from several cell-free expression conditions. High affinity and specific binding of ligands could furthermore be obtained with non-purified receptor samples in crude cell-free reaction mixtures, thus providing new perspectives for fast in vitro screening applications.

Overcoming the real and imagined barriers to cholesterol screening in pediatrics.

Recent guidance by the United States Preventive Services Task Force has renewed the debate surrounding the benefits of pediatric lipid screening. This commentary reviews the evolution of the pediatric lipid screening recommendations in the United States, followed by an exploration of real and imagined challenges that prevent optimal cholesterol screening rates in children. Real challenges substantively prevent the uptake of these guidelines into practice; imagined challenges, such as identifying the best age to screen, are often context-dependent and can also be surmounted. Experiences from other countries identify potential facilitators to improving screening and additional barriers. Implementation science provides guidance on overcoming the real barriers, translating evidence-based recommendations into clinical practice, and informing the next wave of solutions to overcome these challenges.

"I don't think people should die young": perspectives of parents with children diagnosed with familial hypercholesterolemia.

Familial Hypercholesterolemia (FH) is an inherited disorder that significantly increases an individual's risk of developing premature cardiovascular disease (CVD). Early intervention involving lifestyle modification and medication is crucial in preventing CVD. Prior studies have shown that lipid-lowering therapy in children is safe and effective. Despite FH being a treatable and manageable condition, the condition is still underdiagnosed and undertreated. Universal lipid screening (ULS) in children has been recommended by some medical experts in the United States as a strategy to identify cases of FH and maximize the benefits of early invention. However, lipid screening is not routinely offered in pediatric clinics. This study aimed to explore parental experience with FH diagnosis in their children, identify key facilitators and barriers in children's diagnosis and care, and examine parental perspectives on ULS in children in the United States. A total of fourteen semi-structured interviews were conducted with participants recruited through the Family Heart Foundation. Thematic analysis identified three key themes: role of family history in facilitating child's FH diagnosis, barriers and challenges in post-diagnosis care, and attitudes towards ULS in children. All participants supported ULS in children and emphasized the value of early diagnosis and treatment for FH. However, a lack of guidance or referral after the child's diagnosis was a concern raised by many participants. This underscores the need for accessible and comprehensive care amid ongoing efforts to increase pediatric diagnosis of FH.

Fluorescence-Detection Size-Exclusion Chromatography-Based Thermostability Assay for Membrane Proteins.

Green fluorescent proteins (GFPs) have lightened up almost every aspect of biological research including protein sciences. In the field of membrane protein structural biology, GFPs have been used widely to monitor membrane protein localization, expression level, the purification process and yield, and the stability inside the cells and in the test tube. Of particular interest is the fluorescence-detector size-exclusion chromatography-based thermostability assay (FSEC-TS). By simple heating and FSEC, the generally applicable method allows rapid assessment of the thermostability of GFP-fused membrane proteins without purification. Here we describe the experimental details and some typical results for the FSEC-TS method.

Adherence With Lipid Screening Guidelines in Children With Acquired and Congenital Heart Disease: An Observational Study Using Data From The MarketScan Commercial and Medicaid Databases.

Background Universal lipid screening in children provides an opportunity to mitigate the lifetime risk of atherosclerosis, particularly in children with chronic conditions that are predisposed to early atherosclerosis. In response, national guidelines recommend additional early screening in a subset of cardiac conditions. The penetration of such guidelines has not been evaluated. Methods and Results We performed a retrospective study of a geographically representative sample of US children using the MarketScan Commercial and Medicaid claims databases. The study population was children with cardiac disease between ages 2 and 18 years and ≥3 years of continuous coverage from January 1, 2013, to June 30, 2018, divided into 4 major strata of heart disease. We assessed the likelihood of screening between these classifications and compared with healthy children and calculated multivariate models to identify patient factors associated with screening likelihood. Of the eligible 8.4 million children, 155 000 children had heart disease, of which 1.8% (31 216) had high-risk conditions. Only 17.5% of healthy children underwent lipid screening. High-risk children were more likely to be screened (odds ratio [OR], 2.1; 95% CI, 2.09-2.19; P<0.001) than standard-risk children, but that likelihood varied depending on strata of cardiac disease (22%-77%). Timing of screening also varied, with most occurring between ages 9 and 11 years. Among cardiac conditions, heart transplantation (OR, 16.8; 95% CI, 14.4-19.7) and cardiomyopathy (OR, 2.9; 95% CI, 2.8-3.1) were associated with the highest likelihood of screening. Conclusions Children with cardiac disease are more likely to undergo recommended lipid screening than healthy children, but at lower rates and later ages than recommended, highlighting the importance of quality improvement and advocacy for this vulnerable population.

Racial/Ethnic Disparities in Screening for and Awareness of High Cholesterol Among Pregnant Women Receiving Prenatal Care.

Background Atherosclerotic cardiovascular disease remains a leading cause of morbidity and mortality among women, with younger women being disproportionately affected by traditional cardiovascular risk factors such as dyslipidemia. Despite recommendations for lipid screening in early adulthood and the risks associated with maternal dyslipidemia during pregnancy, many younger women lack access to and utilization of early screening. Accordingly, our objective was to assess the prevalence of and disparities in lipid screening and awareness of high cholesterol as an atherosclerotic cardiovascular disease risk factor among pregnant women receiving prenatal care. Methods and Results We invited 234 pregnant women receiving prenatal care at 1 of 3 clinics affiliated with the University of Pennsylvania Health System to complete our survey. A total of 200 pregnant women (86% response rate) completed the survey. Overall, 59% of pregnant women (mean age 32.2 [±5.7] years) self-reported a previous lipid screening and 79% of women were aware of high cholesterol as an atherosclerotic cardiovascular disease risk factor. Stratified by racial/ethnic subgroups, non-Hispanic Black women were less likely to report a prior screening (43% versus 67%, P=0.022) and had lower levels of awareness (66% versus 92%, P<0.001) compared with non-Hispanic White women. Non-Hispanic Black women were more likely to see an obstetrician/gynecologist for their usual source of non-pregnancy care compared with non-Hispanic White women (18% versus 5%, P=0.043). Those seeing an obstetrician/gynecologist for usual care were less likely to report a prior lipid screening compared with those seeing a primary care physician (29% versus 63%, P=0.007). Conclusions Significant racial/ethnic disparities persist in lipid screening and risk factor awareness among pregnant women. Prenatal care may represent an opportunity to enhance access to and uptake of screening among younger women and reduce variations in accessing preventive care services.

In pediatric familial hypercholesterolemia, lipoprotein(a) is more predictive than LDL-C for early onset of cardiovascular disease in family members.

BACKGROUND: Low-density lipoprotein cholesterol (LDL-C) level and lipoprotein(a) [Lp(a)] ≥ 50 mg/dL predict atherosclerotic cardiovascular disease (ASCVD) risk in adults with familial hypercholesterolemia (FH), but their role for children with FH is less clear. OBJECTIVE: This study examined the relationship between elevated Lp(a) and LDL-C levels in a pediatric population with FH and onset of ASCVD in family members. METHODS: Retrospective review of pediatric patients with FH identified LDL-C, Lp(a), and family history of ASCVD. Logistic regression modeling evaluated the association between the child's Lp(a) and peak LDL-C level with earliest age of ASCVD onset in their family. RESULTS: One hundred twenty-nine children from 109 families were identified. Children from families with early-onset ASCVD were 3 times more likely to have high Lp(a) than those with a family history of late-onset ASCVD (OR: 3.77, 95% CI: 1.16-12.25, P = .027) but were not more likely to have highly elevated peak LDL-C (≥190 mg/dL) (OR: 0.45, 95% CI: 0.11-1.80, P = .26). CONCLUSION: Children with FH and family history of early-onset ASCVD were more likely to have Lp(a) ≥50 mg/dL than children with FH and family history of late-onset ASCVD. Family history of early-onset ASCVD was more predictive of a child's Lp(a) level than of a child's peak LDL-C. Measurement of Lp(a) in children with FH may better characterize their cardiovascular risk, particularly when knowledge of family history is limited. Lp(a) testing may also identify children with FH that could benefit from more aggressive management to reduce ASCVD risk.

Membrane Protein Production in E. coli Lysates in Presence of Preassembled Nanodiscs.

Cell-free expression allows to synthesize membrane proteins in completely new formats that can relatively easily be customized for particular applications. Amphiphilic superstructures such as micelles, lipomicelles, or nanodiscs can be provided as nano-devices for the solubilization of membrane proteins. Defined empty bilayers in the form of nanodiscs offer native like environments for membrane proteins, supporting functional folding, proper oligomeric assembly as well as stability. Even very difficult and detergent-sensitive membrane proteins can be addressed by the combination of nanodisc technology with efficient cell-free expression systems as the direct co-translational insertion of nascent membrane proteins into supplied preassembled nanodiscs is possible. This chapter provides updated protocols for the synthesis of membrane proteins in presence of preassembled nanodiscs suitable for emerging applications such as screening of lipid effects on membrane protein function and the modulation of oligomeric complex formation.

Lipoprotein(a): A Lipoprotein Whose Time Has Come.

OPINION STATEMENT: Cardiovascular disease is a leading cause of morbidity and mortality in the USA and around the world. While we are now able to achieve significant low-density lipoprotein cholesterol (LDL-C) lowering with current therapies, many patients remain at risk for cardiovascular disease (CVD). Elevated lipoprotein(a) [Lp(a)] has been shown to be an independent risk factor for CVD and accounts for some of the residual CVD risk after LDL-C lowering in several large clinical trials. Moreover, there is now strong evidence supporting the causal relationship between Lp(a) and aortic stenosis as well as peripheral arterial disease. Despite the growing interest in this lipoprotein, the current therapeutic options for Lp(a) reduction are limited. Our general approach in patients with elevated Lp(a) levels is to aggressively manage other modifiable cardiovascular risk factors including lifestyle modification, consideration of aspirin therapy, and LDL-C lowering. Unfortunately, there are conflicting reports on how effective this strategy is at reducing the risk for cardiovascular events attributed to elevated Lp(a). As a result, targeted Lp(a)-lowering strategies are needed. Lp(a) therapeutics is an active area of research with several promising classes of pharmacotherapies under investigation to address this causal biomarker.

Cell-Free Production of Membrane Proteins in Escherichia coli Lysates for Functional and Structural Studies.

The complexity of membrane protein synthesis is largely reduced in cell-free systems and it results into high success rates of target expression. Protocols for the preparation of bacterial lysates have been optimized in order to ensure reliable efficiencies in membrane protein production that are even sufficient for structural applications. The open accessibility of the semisynthetic cell-free expression reactions allows to adjust membrane protein solubilization conditions according to the optimal folding requirements of individual targets. Two basic strategies will be exemplified. The post-translational solubilization of membrane proteins in detergent micelles is most straightforward for crystallization approaches. The co-translational integration of membrane proteins into preformed nanodiscs will enable their functional characterization in a variety of natural lipid environments.

Counterpoint: The evidence does not support universal screening and treatment in children.

Few pediatric guidelines have generated the amount or intensity of controversy that the pediatric lipid guidelines have. In the following article, I will synthesize the arguments against universal lipid screening and treatment in childhood. Direct evidence that relates the presence of cardiovascular risk factors in childhood to cardiovascular disease outcomes in adulthood is unavailable, and as a consequence, the guidelines were formulated based on a chain of indirect evidence. The debate centers on the strength of the indirect evidence that links risk factors present in childhood to adult disease outcomes. The arguments against universal lipid screening and treatment of children include (1) a history of unanticipated harms caused by screening tests or treatments that were enacted based on indirect evidence, (2) the poor test performance characteristics of lipid profiles in childhood when used as a screening test, (3) problems with the effectiveness of lipid testing done in the office setting, and (4) concerns regarding the safety of statins when used in children.

Dyslipidemia in Overweight and Obese School-Aged Children.

Dyslipidemia often affects overweight and obese adolescents and can be present along with hypertension, insulin resistance, type 2 diabetes, and polycystic ovarian syndrome. This article is the third of six discussing the comorbidities of childhood obesity and will focus on the individual parts of the lipid profile and the impact of dyslipidemia on the heart and other body systems. Since few pharmacologic therapies are approved to treat dyslipidemia in children and adolescents younger than 18, treatment consists of lifestyle changes that can be supported and modeled by the school nurse. The school nurse can also be an advocate for a healthy lifestyle in the school district and community. More success in the treatment of dyslipidemia will be realized with less attention to changing the individual and more attention to changing the wider populations, including schools and the community.

Comparison of two point of care devices for capillary lipid screening in fasting and postprandial adults.

OBJECTIVES: The aim of this study was to assess the performance of two point of care (POC) devices for capillary lipid screening in fasting and post-prandial adults. DESIGN AND METHODS: Fasting and post-prandial capillary whole blood samples collected from 57 adult donors were analyzed simultaneously on Cholestech LDX Lipid Profile (Alere San Diego, Inc., San Diego, CA) cassettes and CardioChek Lipid Panel (Polymer Technology Systems, Indianapolis, IN) strips. Paired serum samples were collected from the same donors and analyzed with CDC-certified methods for total cholesterol, high density lipoprotein cholesterol (HDL-C) and non-blanked triglycerides. Non-HDL-C (total cholesterol minus HDL-C) and low density lipoprotein cholesterol (LDL-C) were calculated. Mean bias between capillary whole blood and serum laboratory lipids was calculated. RESULTS: HDL-C measurements were not affected by triglyceride content on either device. However, both devices exhibited significant variability in triglyceride measurement relative to the reference method. Compared to reference methods, Cholestech was more accurate than CardioChek for non-HDL-C while CardioChek was more accurate for HDL-C. Among the calculated cardiovascular risk parameters (LDL-C and non-HDL-C), Cholestech-calculated non-HDL-C exhibited the least average bias in both fasting and postprandial samples. CONCLUSIONS: The optimal approach to capillary lipid screening may be to use Cholestech non-HDL cholesterol; as it exhibited little bias relative to CDC reference methods in both fasting and postprandial samples, facilitating lipid screening in non-fasting adults.

Screening for lipid requirements of membrane proteins by combining cell-free expression with nanodiscs.

Cell-free (CF) protein expression has emerged as one of the most efficient production platforms for membrane proteins. Central bottlenecks prevalent in conventional cell-based expression systems such as mistargeting, inclusion body formation, degradation as well as product toxicity can be addressed by taking advantage of the reduced complexity of CF expression systems. However, the open accessibility of CF reactions offers the possibility to design customized artificial expression environments by supplying synthetic hydrophobic compounds such as micelles or membranes of defined composition. The open nature of CF systems therefore generally allows systematic screening approaches for the identification of efficient cotranslational solubilization environments of membrane proteins. Synergies exist in particular with the recently developed nanodisc (ND) technology enabling the synthesis of stable and highly soluble particles containing membrane discs of defined composition. Specific types of lipids frequently modulate folding, stability, and activity of integrated membrane proteins. One recently reported example are phospho-MurNAc-pentapeptide (MraY) translocases that catalyze a crucial step in bacterial peptidoglycan biosynthesis making them interesting as future drug targets. Production of functionally active MraY homologues from most human pathogens in conventional cellular production systems was so far not successful due to their obviously strict lipid dependency for functionally folding. We demonstrate that the combination of CF expression with ND technologies is an efficient strategy for the production of folded MraY translocases, and we present a general protocol for the rapid screening of lipid specificities of membrane proteins.