Filtern
Dokumenttyp
Sprache
- Englisch (3)
Referierte Publikation
- ja (3)
Schlagworte
Organisationseinheit der BAM
Arteriosclerosis with its clinical sequelae (cardiac infarction, stroke, peripheral arterial occlusive disease) and vascular/Alzheimer dementia not only result in far more than half of all deaths but also represent dramatic economic problems. The reason is, among others, that diabetes mellitus is an independent risk factor for both disorders, and the number of diabetics strongly increases worldwide. More than one-half of infants in the first 6 months of life have already small collections of macrophages and macrophages filled with lipid droplets in susceptible segments of the coronary arteries. On the other hand, the authors of the Bogalusa Heart Study found a strong increase in the prevalence of obesity in childhood that is paralleled by an increase in blood pressure, blood lipid concentration, and type 2 Diabetes mellitus. Thus, there is a clear linkage between arteriosclerosis/Alzheimer's disease on the one hand and diabetes mellitus on the other hand. Furthermore, it has been demonstrated that distinct apoE isoforms on the blood lipids further both arteriosclerotic and Alzheimer nanoplaque formation and therefore impair flow-mediated vascular reactivity as well. Nanoplaque build-up seems to be the starting point for arteriosclerosis and Alzheimer's disease in their later full clinical manifestation.
In earlier work, we could portray the anionic biopolyelectrolytes syndecan/perlecan as blood flow sensors and lipoprotein receptors in cell membrane and vascular matrix. We described extensively molecular composition, conformation, form and function of the macromolecule heparan sulfate proteoglycan (HS-PG). In two supplementary experimental settings (ellipsometry, myography),we utilized isolated HS-PG for in vitro nanoplaque investigations and isolated human coronary artery segments for in vivo tension measurements.
With the ellipsometry-based approach, we were successful in establishing a direct connection on a molecular level between diabetes mellitus on the one side and arteriosclerosis/Alzheimer's disease on the other side. Application of glucose at a concentration representative for diabetics and leading to glycation of proteins and lipids, entailed a significant increase in arteriosclerotic and Alzheimer nanoplaque formation. IDLapoE4/E4 was by far superior to IDLapoE3/E3 in Plaque build-up, both in diabetic and non-diabetic patients. Recording vascular tension of flow-dependent reactivity in blood substitute solution and under application of different IDLapoE isoforms showed an impaired vasorelaxation for pooled IDL and IDLapoE4/E4, thus confirming the ellipsometric investigations. Incubation in IDLapoE0/E0 (apoE “knockout man”), however, resulted in a massive flowmediated contraction, also complemented by strongly aggregated nanoplaques.
In contrast, HDL was shown to present a powerful protection against nanoplaque formation on principle, both in the in vitro model and the in vivo scenario on the endothelial cell membrane. The competitive interplay with LDL is highlighted through the flow experiment, where flow-mediated, HDL-induced vasodilatation remains untouched by additional incubation with LDL. This is due to the four times higher affinity for the proteoglycan receptor of HDL as compared to LDL.
Taken together, the studies demonstrate that while simplistic, the ellipsometry approach and the endothelialmimicking proteoglycan-modified surfaces provide information on the initial steps of lipoprotein-related Plaque formation, which correlates with findings on endothelial cells and blood vessels, and afford insight into the role of lipoprotein deposition and Exchange phenomena at the onset of these pathophysiologies.
An isotopic effect of normal (H2O) vs. heavy water (D2O) is well known to fundamentally affect structure and chemical properties of proteins, for instance. Here we correlate results from small angle X-ray and neutron scattering (SAXS, SANS) with high-resolution scanning transmission electron microscopy to track the evolution of CdS nanoparticle size and crystallinity from aqeuous solution in presence of the organic ligand ethylenediaminetetraacetate (EDTA) at room temperature in both H2O and D2O. We provide evidence via SANS experiments that exchanging H2O by D2O impacts nanoparticle formation by changing the equilibria and dynamics of EDTA clusters in solution as investigated by nuclear magnetic resonance. The colloidal stability of the CdS nanoparticles, covered by a layer of [Cd(EDTA)]2- complexes, is significantly reduced in D2O despite the strong stabilizing effect of EDTA in suspensions of normal water. Hence, conclusions about nanoparticle formation mechanisms from D2O solutions can bare limited transferability to reactions in normal water due to isotopic effects, which thus need to be discussed for contrast match experiments.
Investigation of the Thermal Stability of Proteinase K for the Melt Processing of Poly(L‑lactide)
(2022)
The enzymatic degradation of aliphatic polyesters offers unique opportunities for various use cases in materials science. Although evidently desirable, the implementation of enzymes in technical applications of polyesters is generally challenging due to the thermal lability of enzymes. To prospectively overcome this intrinsic limitation, we here explored the thermal stability of proteinase K at conditions applicable for polymer melt processing, given that this hydrolytic enzyme is well established for its ability to degrade poly(L-lactide) (PLLA). Using assorted spectroscopic methods and enzymatic assays, we investigated the effects of high temperatures on the structure and specific activity of proteinase K. Whereas in solution, irreversible unfolding occurred at temperatures above 75−80 °C, in the dry, bulk state, proteinase K withstood prolonged incubation at elevated temperatures. Unexpectedly little activity loss occurred during incubation at up to 130 °C, and intermediate levels of catalytic activity were preserved at up to 150 °C. The resistance of bulk proteinase K to thermal treatment was slightly enhanced by absorption into polyacrylamide (PAM) particles. Under these conditions, after 5 min at a temperature of 200 °C, which is required for the melt processing of PLLA, proteinase K was not completely denatured but retained around 2% enzymatic activity. Our findings reveal that the thermal processing of proteinase K in the dry state is principally feasible, but equally, they also identify needs and prospects for improvement. The experimental pipeline we establish for proteinase K analysis stands to benefit efforts directed to this end. More broadly, our work sheds light on enzymatically degradable polymers and the thermal processing of enzymes, which are of increasing economical and societal relevance.