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Introduction: Von Willebrand factor propeptide (VWF:Ag II) is proposed to be a more sensitive marker of acute endothelial activation than von Willebrand factor antigen (VWF:Ag). Simultaneous data on VWF:Ag and VWF:Ag II profiles are very limited following TIA and ischaemic stroke., Methods: In this prospective, observational, case-control study, plasma VWF:Ag and VWF:Ag II levels were quantified in 164 patients≤4weeks of TIA or ischaemic stroke (baseline), and then ≥14days (14d) and ≥90days (90d) later, and compared with those from 27 healthy controls. TIA and stroke subtyping was performed according to the TOAST classification. The relationship between VWF:Ag and VWF:Ag II levels and platelet activation status was assessed., Results: 'Unadjusted' VWF:Ag and VWF:Ag II levels were higher in patients at baseline, 14d and 90d than in controls (p≤0.03). VWF:Ag levels remained higher in patients than controls at baseline (p≤0.03), but not at 14d or 90d after controlling for differences in age or hypertension, and were higher in patients at baseline and 90d after controlling for smoking status (p≤0.04). 'Adjusted' VWF:Ag II levels were not higher in patients than controls after controlling for age, hypertension or smoking (p≥0.1). Patients with symptomatic carotid stenosis (N=46) had higher VWF:Ag and VWF:Ag II levels than controls at all time-points (p≤0.002). There was no significant correlation between platelet activation status and VWF:Ag or VWF:Ag II levels., Conclusions: VWF:Ag and VWF:Ag II levels are increased in an overall TIA and ischaemic stroke population, especially in patients with recently symptomatic carotid stenosis. VWF:Ag II was not superior to VWF:Ag at detecting acute endothelial activation in this cohort and might reflect timing of blood sampling in our study., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objective: This study aimed to investigate the age-related changes in the position of zonular insertion onto the anterior lens capsule., Design: The study design was an experimental cadaveric eye study., Participants: Two hundred twenty-three cadaveric eyes (range, 16-100 years old) from eye banks in the United States were studied., Intervention: The zonular insertion onto the anterior capsule was detected using a modified posterior photographic technique., Main Outcome Measures: The distances from the anterior zonular insertion (AZI) to the lens equator and to the ciliary sulcus, size of zonular-free zone, lens diameter, ciliary sulcus diameter, and axial length of the globe were measured., Results: Multiple regression analysis showed that age, lens diameter, and axial length of the globe had significant correlations with AZI (r2 = 0.748), among which age showed the highest relevance with AZI. Multiple regression equation was expressed as AZI = (0.0079 x age) + (0.202 x lens diameter) - (0.041 x axial length of the globe) + 0.0114. The increase in AZI was accompanied by a decrease in the size of zonular-free zone with age (r = -0.206, P < 0.001)., Conclusion: Aging correlates with an increase in the distance from the anterior zonular insertion to the lens equator and a decrease in zonular-free zone.

A new homogeneous liposome immunoassay system was developed in which analyte-phospholipase C conjugates are used instead of complement or melittin. This system was applied for the determination of insulin. Liposomes incorporated with calcein as a marker were prepared by the reverse-phase evaporation method. The lysis of liposomes was determined by measuring the fluorescence intensity of calcein released from liposomes and it was increased with increasing concentration of phospholipase C. Insulin was conjugated to phospholipase C by a three-step procedure with hetero-bifunctional crosslinking reagents, succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate and 3-(2-pyridyldithio)propionic acid N-hydroxysuccinimide ester. The lytic activity of phospholipase C was not affected by the reaction for conjugation. Both p-nitrophenylphosphatidylcholine hydrolytic activity and liposome lytic activity of insulin-phospholipase C conjugate were inhibited in the presence of insulin antiserum. However, lower concentration of conjugate and shorter incubation time were required when liposomes were used in the inhibition study. The antibody inhibition of conjugate-induced lysis could be reversed by addition of a competing amount of free insulin. The standard calibration curve was obtained in the range between 4 and 130 microIU/ml (r = 0.994). The detection limit (8 microIU/ml) was comparable with those of conventional heterogeneous enzyme immunoassays. This new assay approach offers a simple, sensitive, and inexpensive testing procedure for determining ultratrace amounts of bioactive substances.

Protein kinase C (PKC) plays an important role in activation of human immunodeficiency virus type 1 (HIV-1). Because of a molecular and biochemical heterogeneity of PKC, we have studied the effects of PKC isozymes in HIV-1 activation in a latently infected promonocytic cell line, U1, using various PKC isozyme agonists. 12-Deoxyphorbol 13-phenylacetate (dPP), an agonist of both Ca(++)-dependent and Ca(++)-independent isozymes, and thymeleatoxin (TT), an agonist of Ca(++)-dependent PKC isozymes, induced HIV-1 production at 10 nM with increase in a concentration dependent manner, whereas 12-deoxyphorbol 13-phenylacetate 20-acetate (dPPA), an PKC beta I isozyme agonist, did not induce viral production at 100 nM. We verified that dPPA induced translocation of PKC beta isozyme with the isozyme-specific monoclonal antibody using flow cytometry. This study demonstrates that activation of PKC isozymes leads to an induction of latent HIV-1 in U1 cells whereas PKC beta I isozyme may not be important.

Immunoliposomes were prepared and their immunoassay applications investigated. Liposomes were prepared from cholesterol and phospholipids including maleimidobenzoylphosphatidylethanolamine (MBPE) for conjunction with thiol-containing antigens. Alkaline phosphatase (APase) was entrapped in the liposome and BSA, the antigen, was modified by reaction with 3-(2-pyridyl-dithio)propionyl N-hydroxysuccinimide ester (SPDP) to introduce thiol groups for efficient coupling. BSA-coupled liposomes (immunoliposomes) were incubated with anti-BSA serum, complement, and then with APase substrate. The amount of coupled BSA was affected by the reaction time, the composition of the liposome and the BSA concentration in the reactant. The amount of enzyme released from immunoliposomes as a final result of the immunoreaction increased with increasing concentrations of complement and antibody. The liposome immunoassay offers a relatively rapid and simple testing procedure to quantitatively or qualitatively determine the presence or absence of antibodies, or antigenic materials for diagnostic purposes.