HDL3-mediated cholesterol efflux from cultured enterocytes: The role of apoproteins A-I and A-II

High density lipoproteins (HDL) were recently demonstrated in an enterocyte model (CaCo-2 cells) to mediate reverse cholesterol transport by retroendocytosis. The present study was carried out to define the role of the major HDL apoproteins (apo) A-I and apo A-II in this pathway. HDL₃ was fractionated by heparin affinity chromatography into the two main fractions containing either apo A-I only (fraction A) or both apo A-I and apo A-II (fraction B). In addition, liposomes were reconstituted from purified apo A-I or apo A-II and dimyristoyl phosphatidylcholine. The cell binding properties and cholesterol efflux potential were studied in the lipoprotein fractions and the liposomes. Both fractions exhibited similar maximal binding capacities of 4427 (A) and 5041 (B) ng/mg cell protein, but their dissociation constants differed (40.5 and 167.7 μg/mL, respectively). Fraction A induced cholesterol efflux and stimulated cholesterol synthesis more than did fraction B. Fraction A mobilized both cellular free and esterified cholesterol, whereas fraction B preferentially mobilized cholesteryl esters. Liposomes, containing either apo A-I or apo A-II, showed specific binding, endocytosis and endosomal transport, and were released as intact particles. Apo A-I liposomes also mediated cholesterol efflux. In conclusion, there is evidence that the HDL₃ subfractions A and B, as well as reconstituted liposomes containing either apo A-I or apo A-II, were specifically bound and entered a retroendocytosis pathway which was directly linked to cholesterol efflux. Quantitatively, the apo A-I subfraction appeared to play the dominant role in normal enterocytes. The apo A-II content of fraction B was related to the mobilization of cholesteryl esters.     

Assessing landscape change and biodiversity values of the Middle Vistula river valley, Poland, using BIO-SAFE

This paper deals with the relation between legally protected biodiversity and riverine ecotopes and with the assessment of biodiversity values of the Middle Vistula river valley (Poland). Furthermore, it describes the effects of landscape change on spatial distribution of ecotope patches and biodiversity values in Kazimierski Landscape Park. Biodiversity values were calculated using BIO-SAFE, a model meant to quantify biodiversity and to value ecotopes based on legally protected species. Dissimilarity indices depict high uniqueness of ecotope types regarding their species assemblages (e.g., river dunes, banks and bars). The actual biodiversity values of the river valley in Kazimierski Landscape Park are high in comparison with floodplains of lowland rivers in Western Europe. GIS analyses of remotely sensed ecotope maps show remarkable differences in number, acreage and patchiness of ecotopes for the years 1953 and 2003. The total number of patches increased by almost 44%. Side channels and floodplain lakes became fragmented. The average and total surface area of bush, forest and arable land increased, but decreased for bare soil, pioneer vegetation and grassland. These landscape changes indicate natural vegetation succession, intensification of agriculture and progressive impacts of river regulation. The Vistula river valley still represents high biodiversity values for higher plants, birds, herpetofauna and fish. However, current landscape changes negatively affect potential values for protected and endangered species. Assessments with BIO-SAFE can help to balance biodiversity conservation, river management and landscape planning.     

Exchange of domains of glutamate dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus and the mesophilic bacterium Clostridium difficile: effects on catalysis, thermoactivity and stability

The glutamate dehydrogenase gene from the hyperthermophilicarchaeon Pyrococcus furiosus has been functionally expressedin Escherichia coli under the control of the X, PL promoter.The P.furiosus glutamate dehydrogenase amounted to 20% of thetotal E.coli cell protein, and the vast majority consisted ofhexamers. Following activation by heat treatment, an enzymecould be purified from E.coli that was indistinguishable fromthe glutamate dehydrogenase purified from P.furiosus. Hybridgenes, that consisted of the coding regions for the homologousglutamate dehydrogenases from P.furiosus and the mesophilicbacterium Clostridium difficile, were constructed and successfullyexpressed in E.coli. One of the resulting hybrid proteins, containingthe glutamate binding domain of the C.difficile enzyme and thecofactor binding domain of the P.furiosus enzyme, did not showa detectable activity. In contrast, the complementary hybridcontaining the P.furiosus glutamate and the C.difficile cofactorbinding domain was a catalytically active hexamer that showeda reduced substrate affinity but maintained efficient cofactorbinding with the specificity found in the Clostridium symbiosumenzyme. Compared with the C.difficile glutamate dehydrogenase,the archaeal-bacterial hybrid is slightly more thermoactive,less thermostable but much more stable towards guanidinium chloride-inducedinactivation and denaturation     

In vitro characterization and ex vivo surgical evaluation of human hair keratin films in ocular surface reconstruction after sterilization processing

The disadvantages of human amniotic membrane (hAM), used for ocular surface reconstruction, necessitate the development of standardized alternatives. Keratin-derived-films (KF) have been indicated as transferable substrate for cell cultivation and tissue engineering. The impact of different sterilization procedures on KF and surgical feasibility were investigated. Human hair KF were prepared and sterilized; optical, biomechanical properties, in vitro cell seeding efficiency and proliferation of human corneal epithelial cells were studied and compared with hAM. Surgical feasibility was tested on enucleated porcine eye. Sterilized KF showed higher light transmission and significantly higher E-modulus than hAM; cell-seeding-efficiency and proliferation rate were not affected. Although KF could be surgically handled, suture placement was more difficult compared to hAM. Plasma treatment seems the best sterilization method for KF; it does not affect cell biology or optical and biomechanical properties. However material modifications are requested before KF may represent a feasible alternative for ocular surface reconstruction.     

Bio-geomorphic alteration through shifting flow regime in a modified monsoonal river system in Korea

River processes and patterns are affected by changes in the watershed driven by natural and human-induced causes. A sudden pattern alteration from a “white river” (bare soils) to a “green river” (vegetated) influences riverine biodiversity and can increase flood risk. Despite these significant impacts, knowledge on the triggers that kickstart feedback exacerbating changes in bio-geomorphic patterns is insufficient. In this study, we collected and analyzed detailed monitoring data on a sandy, hydro-morphologically active, and monsoon-driven river in Korea. The surface area covered by vegetation has been increasing; this increase intensified after the 2014–2015 drought, which provided a window of opportunity for vegetation establishment. During the drought, pioneer vegetation densely colonized bare sandbars and temporarily exposed riverbed. Despite partial rejuvenation by several subsequent floods, succession to woody vegetation occurred, resulting in a stable vegetation cover. Narrowing and incision of a low-water channel occurred, and secondary channels formed inside the floodplain. The results of this study show a rapid bio-geomorphic alteration triggered by the shifts in flow regime in a river primed by human-induced changes. Furthermore, modified monsoon-driven rivers might be on the brink of similar bio-physical alterations triggered through shifting flow regime following climate change, leading to increased flood risk and impacts on endemic biodiversity.     

The effect of physical accessibility and service level of water supply on economic accessibility: a case study of Bandung City,Indonesia

Achieving equitable access to water, in the sense of both physical and economic accessibility, remains a challenge. The article evaluates these two types of accessibility across households of different income groups in Central Cikapundung Basin, Indonesia. Higher-income households are more likely to use piped water, bottled water, or combinations thereof and have higher water expenditures than their lower-income counterparts. We estimate the hidden mitigation costs of groundwater extraction and water boiling and highlight the importance of incorporating mitigation costs when assessing the impacts of poor service quality of water supply on household water expenditure and affordability.     

HDL3-mediated cholesterol efflux from cultured enterocytes: the role of apoproteins A-I and A-II

High density lipoproteins (HDL) were recently demonstrated in an enterocyte model (CaCo-2 cells) to mediate reverse cholesterol transport by retroendocytosis. The present study was carried out to define the role of the major HDL apoproteins (apo) A-I and apo A-II in this pathway. HDL3 was fractionated by heparin affinity chromatography into the two main fractions containing either apo A-I only (fraction A) or both apo A-I and apo A-II (fraction B). In addition, liposomes were reconstituted from purified apo A-I or apo A-II and dimyristoyl phosphatidylcholine. The cell binding properties and cholesterol efflux potential were studied in the lipoprotein fractions and the liposomes. Both fractions exhibited similar maximal binding capacities of 4427 (A) and 5041 (B) ng/mg cell protein, but their dissociation constants differed (40.5 and 167.7 micrograms/mL, respectively). Fraction A induced cholesterol efflux and stimulated cholesterol synthesis more than did fraction B. Fraction A mobilized both cellular free and esterified cholesterol, whereas fraction B preferentially mobilized cholesteryl esters. Liposomes, containing either apo A-I or apo A-II, showed specific binding, endocytosis and endosomal transport, and were released as intact particles. Apo A-I liposomes also mediated cholesterol efflux. In conclusion, there is evidence that the HDL3 subfractions A and B, as well as reconstituted liposomes containing either apo A-I or apo A-II, were specifically bound and entered a retroendocytosis pathway which was directly linked to cholesterol efflux. Quantitatively, the apo A-I subfraction appeared to play the dominant role in normal enterocytes. The apo A-II content of fraction B was related to the mobilization of cholesteryl esters.     

Diode laser thermokeratoplasty: application strategy and dosimetry

PURPOSE: To investigate suitable application parameters for efficient hyperopic correction by laser thermokeratoplasty (LTK) using mid-infrared laser diodes. SETTING: Medical Laser Center Lübeck, Lübeck, Germany. METHOD: A tunable continuous-wave laser diode in the spectral range between 1.845 and 1.871 microns was used. Transmitted by waveguides, the laser energy was used to induce coagulations on freshly enucleated porcine eyes to increase corneal curvature. The coagulations were equidistantly applied by a fiber-cornea contact and a noncontact focusing device that were adjusted on a ring concentric to the corneal apex. Different laser parameters and application geometries were evaluated. Refractive changes were measured by computer-assisted corneal topography before and after treatment. Polarization light microscopy and temperature calculations were used to analyze the coagulations. RESULTS: Because of the tunability of the laser diode, the influence of the corneal absorption coefficient (between 0.9 and 1.6 mm-1) on the refractive change could be measured. A laser power between 125 and 200 mW was adequate to achieve refractive changes up to 10.0 diopters. In the preferable focusing device, the refractive change increased almost logarithmically with the irradiation time up to 15 seconds. The number of coagulations on a fixed application ring showed no significant influence on refractive change; however, it showed an almost linear decrease with increasing ring diameter from 5.0 to 10.0 mm. Histological analysis revealed 3 stages of thermal damage. CONCLUSION: Diode LTK provided defined and uniform coagulations when using a well-adapted focusing device, resulting in sufficient refractive change. The results indicate that diode LTK is superior to pulsed holmium LTK.     

Designing optimally-graded depth filter media using a novel multiscale method

Even though current manufacturing methods allow to build precisely-defined graded depth filter media, it remains unclear which local filtration properties are desirable. We, therefore, introduce a multiscale approach which links pore and continuum scale (PS and CS) to address this question. Based on data from PS simulations, local filtration performance, as described by the filter coefficient, is predicted on the CS by an optimal control solution and the obtained trajectory is translated back to the PS where it is validated. Two case studies are presented: a bidisperse fibrous medium and a granular depth filter composed of bidisperse particles. Both media are optimized to achieve a homogenous deposition of separated impurities along the filter depth. It is found that the presented method allows to reach this goal reasonably well in both cases. We claim that our method forms a good basis for further developments for which promising possibilities are highlighted.