Total cholesterol and anthropometric measurements distributions in a cross-sectional study of school children

Cross-sectional age and sex specific distributions of serum total cholesterol were described for 1091 children age 6-18 years, in The Woodlands, Texas. Associations of serum total cholesterol with five anthropometric measurements (weight, height, body mass index, arm circumference, and triceps skinfold thickness) were examined by correlation and regression analyses. Examination of serum total cholesterol distributions showed lower levels in boys than in girls for most of the age groups studied. Mean levels of total cholesterol peaked at age 9 for boys and 8 for girls. Serum total cholesterol leveled off until age 14 for boys and 11 for girls, and then dropped through age 18 for both boys and girls. These results support the hypothesis that serum total cholesterol concentration drops at pre-adolescence.^ Age adjusted correlations were observed between serum total cholesterol and triceps skinfold thickness for both boys and girls. This association was stronger in boys. Triceps skinfold thickness and arm circumference were consistently the strongest correlates for serum total cholesterol in boys. Weight and arm circumference were consistently the strongest correlates for serum total cholesterol in girls. ^

Host-pathogen interactions of secreted and surface Staphylococcus aureus factors

Staphylococcus aureus is an opportunistic bacterial pathogen that can infect humans and other species. It utilizes an arsenal of virulence factors to cause disease, including secreted and cell wall anchored factors. Secreted toxins attack host cells, and pore-forming toxins destroy target cells by causing cell lysis. S. aureus uses cell-surface adhesins to attach to host molecules thereby facilitating host colonization. The Microbial Surface Components Recognizing Adhesive Matrix Molecules (MSCRAMMs) are a family of cell-wall anchored proteins that target molecules like fibronectin and fibrinogen. The Serine-aspartate repeat (Sdr) proteins are a subset of staphylococcal MSCRAMMs that share similar domain organization. Interestingly, the amino-terminus, is composed of three immunoglobulin-folded subdomains (N1, N2, and N3) that contain ligand-binding activity. Clumping factors A and B (ClfA and ClfB) and SdrG are Sdr proteins that bind to fibrinogen (Fg), a large, plasma glycoprotein that is activated during the clotting cascade to form fibrin. In addition to recognizing fibrinogen, ClfA and ClfB can bind to other host ligands. Analysis of S. aureus strains that cause osteomyelitis led to the discovery of the bone-sialoprotein-binding protein (Bbp), an Sdr protein. Because several MSCRAMMs target more than one molecule, I hypothesized that Bbp may recognize other host proteins. A ligand screen revealed that the recombinant construct BbpN2N3 specifically recognizes human Fg. Surface plasmon resonance was used to determine the affinity of BbpN2N3 for Fg, and a dissociation constant of 540 nM was determined. Binding experiments performed with recombinant Fg chains were used to map the binding of BbpN2N3 to the Fg Aalpha chain. Additionally, Bbp expressed on the surface of Lactococcus lactis and S. aureus Newman bald mediated attachment of these bacteria to Fg Aalpha. To further characterize the interaction between the two proteins, isothermal titration calorimetry and inhibition assays were conducted with synthetic Fg Aalpha peptides. To determine the physiological implications of Bbp binding to Fg, the effect of Bbp on fibrinogen clotting was studied. Results show that Bbp binding to Fg inhibits the formation of fibrin. The consequences of this interaction are currently under investigation. Together, these data demonstrate that human Fg is a novel ligand for Bbp. This study indicates that the MSCRAMM Bbp may aid in staphylococcal attachment by targeting both an extracellular matrix and a blood plasma protein. The implications of these novel findings are discussed.

Commissioning an Anthropomorphic Spine and Lung Phantom for Remote Dose Verification of Institutions Participating in RTOG 0631

The RPC developed a new phantom to ensure comparable and consistent radiation administration in spinal radiosurgery clinical trials. This study assessed the phantom’s dosimetric and anatomic utility. The ‘spine phantom’ is a water filled thorax with anatomy encountered in spinal radiosurgery: target volume, vertebral column, spinal canal, esophagus, heart, and lungs. The dose to the target volume was measured with axial and sagittal planes of radiochromic film and thermoluminescent dosimeters (TLD). The dose distributions were measured with the radiochromic film calibrated to the absolute dose measured by the TLD. Four irradiations were administered: a four angle box plan, a seven angle conformal plan, a seven angle IMRT plan, and a nine angle IMRT plan (denoted as IMRT plan #1 and plan #2, respectively). In each plan, at least 95% of the defined tumor volume received 8 Gy. For each irradiation the planned and administered dose distributions were registered via pinpricks, and compared using point dose measurements, dose profiles, isodose distributions, and gamma analyses. Based on previous experience at the RPC, a gamma analysis was considering passing if greater than 95% of pixels passed the criteria of 5% dose difference and 3 mm distance-to-agreement. Each irradiation showed acceptable agreement in the qualitative assessments and exceeded the 95% passing rate at the 5% / 3 mm criteria, except IMRT plan #1, which was determined to have been poorly localized during treatment administration. The measured and planned dose distributions demonstrated acceptable agreement at the 5% / 3 mm criteria, and the spine phantom was determined to be a useful tool for the remote assessment of an institution’s treatment planning and dose delivery regimen.

THE NEW ROLE OF PROPROTEIN CONVERTASE SUBTILISIN/KEXIN TYPE 9: A CONNECTION OF PROPROTEIN CONVERTASE SUBTILISIN/KEXIN TYPE 9, APOLIPOPROTEIN B, and AUTOPHAGY

Plasma low-density lipoprotein (LDL) levels are positively correlated with the incidence of coronary artery disease. In the circulation, the plasma LDL clearance is mainly achieved by the uptake via LDL receptor (LDLR). Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a newly discovered gene, playing an important role in LDL metabolism. Gain-of-function mutations of PCSK9 lead to hypercholesterolemia and loss-of-function mutations of PCSK9 are associated with decrease of LDL cholesterol. The effects of PCSK9 on cholesterol levels are the consequence of a strong interaction between the catalytic domain of PCSK9 and epidermal growth factor-like repeat A (EGF-A) domain of LDLR on the cell surface of hepatocytes. This PCSK9/LDLR complex enters the cell via endocytosis, where both PCSK9 and LDLR are removed via the lysosome pathway, resulting in decreased levels of LDLR and accumulation of LDL in the plasma. However, whether this is the exclusive function of PCSK9 on LDL metabolism was challenged by us; we observed PCSK9 interacted with apolipoprotein B (apoB) and increased apoB production, irrespective of the LDLR. ApoB is the primary structure protein of LDL particle and it also serves as the ligand for the LDL receptor. There is ample evidence showing that the levels of apoB are a better indicator for heart disease than either total cholesterol or LDL cholesterol levels. We used a second-generation adenoviral vector to overexpress PCSK9 (Ad-PCSK9) in wild-type C57BL/6 and LDLR deficient mice (Ldlr-/- and Ldlr-/-Apobec1-/-). Our study revealed that overexpression of PCSK9 promoted the production and secretion of apoB in the form of very-low density lipoprotein (VLDL), which is the precursor of LDL, in the 3 mouse models studied (C57BL/6J, Ldlr-/-, and Ldlr-/-Apobec1-/-). The increased apoB production in mice was regulated at post-transcriptional levels, since there was no difference in apoB mRNA levels between mice treated with Ad-PCSK9 and control vector Ad-Null. By using pulse-chase experiment on primary hepatocytes, we showed that overexpression of PCSK9 increased the secretion of apoB, independent of LDLR. In the circulation, we showed that PCSK9 was associated with LDL particles. By using 3 different protein–protein interaction assays of co-immunoprecipitation, mammalian two-hybrid system, and in situ proximity ligation assay, we demonstrated a direct protein–protein interaction between PCSK9 and apoB. The impact of this interaction inhibited the physiological removal process of apoB via autophagosome/lysosome pathway in an LDLR-independent fashion, resulting in increased production and secretion of apoB-containing lipoproteins. The significance of this process was shown in the Pcsk9 knockout mice in the background of Ldlr-/-Apobec1-/- mice (triple knockout mice); in the absence of Pcsk9 (triple knockout mice) the levels of cholesterol, triacylglycerol, and apoB decreased significantly in comparison to that of Ldlr-/-Apobec1-/- mice. Taken together, our study demonstrated a direct intracellular interaction of PCSK9 with apoB, resulting in the inhibition of apoB degradation via the autophagosome/lysosome pathway independent of LDLR. This discovery provides a new concept of the importance of PCSK9 and suggests new approaches for the therapeutic intervention of hyperlipidemia.

A simulation study using the hybrid statistic and the meta-analysis t-test for data with matched and unmatched subjects in the interim analysis of clinical trials

An interim analysis is usually applied in later phase II or phase III trials to find convincing evidence of a significant treatment difference that may lead to trial termination at an earlier point than planned at the beginning. This can result in the saving of patient resources and shortening of drug development and approval time. In addition, ethics and economics are also the reasons to stop a trial earlier. In clinical trials of eyes, ears, knees, arms, kidneys, lungs, and other clustered treatments, data may include distribution-free random variables with matched and unmatched subjects in one study. It is important to properly include both subjects in the interim and the final analyses so that the maximum efficiency of statistical and clinical inferences can be obtained at different stages of the trials. So far, no publication has applied a statistical method for distribution-free data with matched and unmatched subjects in the interim analysis of clinical trials. In this simulation study, the hybrid statistic was used to estimate the empirical powers and the empirical type I errors among the simulated datasets with different sample sizes, different effect sizes, different correlation coefficients for matched pairs, and different data distributions, respectively, in the interim and final analysis with 4 different group sequential methods. Empirical powers and empirical type I errors were also compared to those estimated by using the meta-analysis t-test among the same simulated datasets. Results from this simulation study show that, compared to the meta-analysis t-test commonly used for data with normally distributed observations, the hybrid statistic has a greater power for data observed from normally, log-normally, and multinomially distributed random variables with matched and unmatched subjects and with outliers. Powers rose with the increase in sample size, effect size, and correlation coefficient for the matched pairs. In addition, lower type I errors were observed estimated by using the hybrid statistic, which indicates that this test is also conservative for data with outliers in the interim analysis of clinical trials.^

Volumetric Modulated Arc Therapy Evaluation with the Radiological Physics Center Head and Neck Phantom

Validation of treatment plan quality and dose calculation accuracy is essential for new radiotherapy techniques, including volumetric modulated arc therapy (VMAT). VMAT delivers intensity modulated radiotherapy treatments while simultaneously rotating the gantry, adding an additional level of complexity to both the dose calculation and delivery of VMAT treatments compared to static gantry IMRT. The purpose of this project was to compare two VMAT systems, Elekta VMAT and Varian RapidArc, to the current standard of care, IMRT, in terms of both treatment plan quality and dosimetric delivery accuracy using the Radiological Physics Center (RPC) head and neck (H&N) phantom. Clinically relevant treatment plans were created for the phantom using typical prescription and dose constraints for Elekta VMAT (planned with Pinnacle3 Smart Arc) and RapidArc and IMRT (both planned with Eclipse). The treatment plans were evaluated to determine if they were clinically comparable using several dosimetric criteria, including ability to meet dose objectives, hot spots, conformity index, and homogeneity index. The planned treatments were delivered to the phantom and absolute doses and relative dose distributions were measured with thermoluminescent dosimeters (TLDs) and radiochromic film, respectively. The measured and calculated doses of each treatment were compared to determine if they were clinically acceptable based upon RPC criteria of ±7% dose difference and 4 mm distance-to-agreement. Gamma analysis was used to assess dosimetric accuracy, as well. All treatment plans were able to meet the dosimetric objectives set by the RPC and had similar hot spots in the normal tissue. The Elekta VMAT plan was more homogenous but less conformal than the RapidArc and IMRT plans. When comparing the measured and calculated doses, all plans met the RPC ±7%/4 mm criteria. The percent of points passing the gamma analysis for each treatment delivery was acceptable. Treatment plan quality of the Elekta VMAT, RapidArc and IMRT treatments were comparable for consistent dose prescriptions and constraints. Additionally, the dosimetric accuracy of the Elekta VMAT and RapidArc treatments was verified to be within acceptable tolerances.