Introduction Blood-based Alzheimer's disease (AD) biomarkers show promise, but pre-analytical protocol differences may pose problems. We examined seven AD blood biomarkers (amyloid beta [A beta]42${\rm{A\beta }}]{_{42}}$, A beta 40${\rm{A}}{{{\beta}}_{40}}$, phosphorylatedtau[p-tau181${\rm{phosphorylated\;tau\;[p - ta}}{{\rm{u}}_{181}}$, total tau [t-tau], neurofilament light chain [NfL], A beta 4240,${\rm{A}}{{{\beta}}_{\frac{{42}}{{40}}}},$ and p-tau181A beta 42$\frac{{{\rm{p - ta}}{{\rm{u}}_{181}}}}{{{\rm{A}}{{{\beta}}_{42}}}}$) in three collection tube types (ethylenediaminetetraacetic acid [EDTA] plasma, heparin plasma, serum). Methods Plasma and serum were obtained from cerebrospinal fluid or amyloid positron emission tomography-positive and -negative participants (N = 38) in the Wisconsin Registry for Alzheimer's Prevention. We modeled AD biomarker values observed in EDTA plasma versus heparin plasma and serum, and assessed correspondence with brain amyloidosis. Results Results suggested bias due to tube type, but crosswalks are possible for some analytes, with excellent model fit for NfL (R2${{\rm{R}}<^>2}\;$= 0.94), adequate for amyloid (R2${{\rm{R}}<^>2}\;$= 0.40-0.69), and weaker for t-tau (R2${{\rm{R}}<^>2}\;$= 0.04-0.42) and p-tau181${\rm{p - ta}}{{\rm{u}}_{181}}$ ( R2${{\rm{R}}<^>2}\;$= 0.22-0.29). Brain amyloidosis differentiated several measures, especially EDTA plasma pTau181A beta 42$\frac{{{\rm{pTa}}{{\rm{u}}_{181}}}}{{{\rm{A}}{\beta _{42}}}}$ (d$d\;$= 1.29). Discussion AD biomarker concentrations vary by tube type. However, correlations for some biomarkers support harmonization across types, suggesting cautious optimism for use in banked blood.

Mutations in cardiac myosin binding protein C (cMyBP-C) are prevalent causes of hypertrophic cardiomyopathy (HCM). Although HCM-causing truncation mutations in cMyBP-C are well studied, the growing number of disease-related cMyBP-C missense mutations remain poorly understood. Our objective was to define the primary contractile effect and molecular disease mechanisms of the prevalent cMyBP-C E258K HCM-causing mutation in nonremodeled murine engineered cardiac tissue (mECT). Wild-type and human E258K cMyBP-C were expressed in mECT lacking endogenous mouse cMyBP-C through adenoviral-mediated gene transfer. Expression of E258K cMyBP-C did not affect cardiac cell survival and was appropriately incorporated into the cardiac sarcomere. Functionally, expression of E258K cMyBP-C caused accelerated contractile kinetics and severely compromised twitch force amplitude in mECT. Yeast two-hybrid analysis revealed that E258K cMyBP-C abolished interaction between the N terminal of cMyBP-C and myosin heavy chain sub-fragment 2 (S2). Furthermore, this mutation increased the affinity between the N terminal of cMyBP-C and actin. Assessment of phosphorylation of three serine residues in cMyBP-C showed that aberrant phosphorylation of cMyBP-C is unlikely to be responsible for altering these interactions. We show that the E258K mutation in cMyBP-C abolishes interaction between N-terminal cMyBP-C and myosin S2 by directly disrupting the cMyBP-C-S2 interface, independent of cMyBP-C phosphorylation. Similar to cMyBP-C ablation or phosphorylation, abolition of this inhibitory interaction accelerates contractile kinetics. Additionally, the E258K mutation impaired force production of mECT, which suggests that in addition to the loss of physiological function, this mutation disrupts contractility possibly by tethering the thick and thin filament or acting as an internal load.

Hypertrophic cardiomyopathy (HCM) is a primary cardiac disease inherited in an autosomal dominant fashion (Spirito et al., 1997). HCM is the most prevalent cause of sudden cardiac death in apparently healthy young individuals (Fananapazir and Epstein, 1991), and is estimated to affect about one in five hundred people (Maron et al., 1995; Seidman and Seidman, 2001). Globally, mutations in cardiac myosin binding protein C (cMyBP-C), encoded by the human cardiac myosin binding protein C gene (MYBPC3) gene, are among the most prevalent causes of HCM, accounting for 34% of all mutations identified (Richard et al., 2003; Olivotto et al., 2008). To date, most recognized HCM-causing mutations in

Congestive heart failure is a debilitating disease in which the principal pathology is impaired ventricular contractility leading to diminished cardiac output, and previous work indicates that reduced contractility is based in part on the ratio of myosin heavy chain (MyHC) isoforms, α- and β-MyHC, expressed in the ventricles. Normal human ventricles express ∼10% of the fast α-MyHC on a background of the slower β-MyHC, while in failing hearts α-MyHC is reduced to virtually undetectable levels with complete replacement by β-MyHC. Data from permeabilized myocardial preparations suggests that this isoform switch may be partly responsible for reduced myocardial twitch force and pressure development by failing ventricles since β-MyHC is a slower motor protein, yet most experiments have used non-human myosins and experimental conditions in which preparations were steadily activated, thus little is known about the response of human myosins to a time-varying Ca2+ transient. To address these limitations, we recently developed a human 3D engineered cardiac tissue (hECT) system in which we can express recombinant human muscle myosin motors. Using commercially available cloning and adenoviral expression systems, α- or β-MyHC isoform expressing adenoviral particles were used to transduce human cardiomyocytes produced from human iPS cells and construct hECTs. Preparations displayed well-defined cellular structure with elongated morphology aligned in the direction of preparation shortening during electrical pacing, while histological analysis of hECT revealed appropriate protein expression and localization within the sarcomere. In response to a Ca2+ transient, the time-course of twitch force development was accelerated in hECT expressing α-MyHC compared to β-MyHC, while peak twitch force was greater in hECT expressing α-MyHC. These results demonstrate the relative contribution of myosin isoforms to myocardial twitch kinetics in human engineered cardiac tissue constructs expressing a stable background of myofibrillar proteins.