Upregulation of selective cholesteryl ester uptake pathway in mice with deletion of low-density lipoprotein receptor function

This study examines the effect of mutation of the low-density lipoprotein receptor (LDLR) on cholesterol metabolism, and especially lipoprotein-derived cholesteryl ester uptake, in murine ovarian granulosa cells. Although the tests were conducted on cells prepared by two different procedures, the results are similar. Deletion of LDLR function did not noticeably affect key enzymes of the steroidogenic pathway or affect progestin production and secretion in granulosa cells. No change was found in expression of LDL-related protein (LRP). These data suggested that cholesterol turnover in cells from the knockout animals is within normal limits and that the cells are not stressed to acquire more cholesterol. Both biochemical and morphological data indicate that unstimulated granulosa cells from LDLR^−/− mice are nonetheless programmed to take in double the amount of lipoprotein-derived cholesteryl ester (via the selective cholesteryl ester uptake pathway) and to process (hydrolyze, re-esterify, or utilize) more than twofold the cholesteryl ester processed by cells from wildtype (LDLR^+/+) animals. Bt₂cAMP stimulation of the murine granulosa cells increases the mass of cholesteryl ester taken up by the selective pathway by an additional 38%. To determine to what extent this increase is related to high-density lipoprotein (HDL) scavenger receptor protein (SR-BI) or caveolin function, Western blots and immunohistochemical studies were performed under a variety of conditions. SR-BI levels are found to be low in unstimulated cells of both LDLR^+/+ and LDLR^−/− animals, but highly expressed (∼20-fold increase over basal levels) in stimulated (Bt₂cAMP) cells of both animal models. Thus, the functional relationship between selective cholesteryl ester uptake and SR-BI receptor protein is not as tight as in previously reported studies, suggesting a requirement for other tissue factors. Caveolin expression did not change under any of the conditions tested and appears not to be functionally involved in this process. J. Cell. Physiol. 180:190–202, 1999. Published 1999 Wiley-Liss, Inc.     

Elucidation of insulin assembly at acidic and neutral pH: Characterization of low molecular weight oligomers

Deficiency in insulin secretion and function that characterize type 2 diabetes often requires administration of extraneous insulin, leading to injection‐site amyloidosis. Insulin aggregation at neutral pH is not well understood. Although oligomer formation is believed to play an important role, insulin oligomers have not been fully characterized yet. Here, we elucidate similarities and differences between in vitro insulin aggregation at acidic and neutral pH for a range of insulin concentrations (2.5–100 μM) by using kinetic thioflavin T fluorescence, circular dichroism, atomic force and electron microscopy imaging. Importantly, we characterize the size distribution of insulin oligomers at different assembly stages by the application of covalent cross‐linking and gel electrophoresis. Our results show that at the earliest assembly stage, oligomers comprise up to 40% and 70% of soluble insulin at acidic and neutral pH, respectively. While the highest oligomer order increases with insulin concentration at acidic pH, the opposite tendency is observed at neutral pH, where oligomers up to heptamers are formed in 10 μM insulin. These findings suggest that oligomers may be on‐ and off‐pathway assemblies for insulin at acidic and neutral pH, respectively. Agitation, which is required to induce insulin aggregation at neutral pH, is shown to increase fibril formation rate and fibrillar mass both by an order of magnitude. Insulin incubated under agitated conditions at neutral pH rapidly aggregates into large micrometer‐sized aggregates, which may be of physiological relevance and provides insight into injection‐site amyloidosis and toxic pulmonary aggregates induced by administration of extraneous insulin.     

Enzymatic pre‐treatment of microalgae cells for enhanced extraction of proteins

Crude proteins and pigments were extracted from different microalgae strains, both marine and freshwater. The effectiveness of enzymatic pre‐treatment prior to protein extraction was evaluated and compared to conventional techniques, including ultrasonication and high‐pressure water extraction. Enzymatic pre‐treatment was chosen as it could be carried out at mild shear conditions and does not subject the proteins to high temperatures, as with the ultrasonication approach. Using enzymatic pre‐treatment, the extracted proteins yields of all tested microalgae strains were approximately 0.7 mg per mg of dry cell weight. These values were comparable to those achieved using a commercial lytic kit. Ultrasonication was not very effective for proteins extraction from Chlorella sp., and the extracted proteins yields did not exceed 0.4 mg per mg of dry cell weight. For other strains, similar yields were achieved by both treatment methods. The time‐course effect of enzymatic incubation on the proteins extraction efficiency was more evident using laccase compared to lysozyme, which suggested that the former enzyme has a slower rate of cell disruption. The crude extracted proteins were fractionated using an ion exchange resin and were analyzed by the electrophoresis technique. They were further tested for their antioxidant activity, the highest of which was about 60% from Nannochloropsis sp. The total phenolic contents in the selected strains were also determined, with Chlorella sp. showing the highest content reaching 17 mg/g. Lysozyme was also found to enhance the extraction of pigments, with Chlorella sp. showing the highest pigments contents of 16.02, 4.59 and 5.22 mg/g of chlorophyll a, chlorophyll b and total carotenoids, respectively.     

Determination of food sources and trophic position in Malaysian tropical highland streams using carbon and nitrogen stable isotopes

Stable isotope analysis has been extensively used as an effective tool in determination of trophic relationship in ecosystems. In freshwater ecosystem, aquatic invertebrates represent main component of a river food web. This study was carried out to determine potential food sources of freshwater organism together with pattern of trophic position along the river food web. In this study, rivers of Belum-Temengor Forest Complex (BTFC) has been selected as sampling site as it is a pristine area that contains high diversity and abundance of organisms and can be a benchmark for other rivers in Malaysia. Stable isotope ratios of carbon (δ¹³C) and nitrogen (δ¹⁵N) were applied to estimate trophic position and food web paradigm. Analysis of stable isotopes based on organic material collected from the study area revealed that the highest δ¹³C value was reported from filamentous algae (? 22.68 ± 0.126⁰/₀₀) and the lowest δ¹³C was in allocthonous leaf packs (? 31.58 ± 0.187⁰/₀₀). Meanwhile the highest δ¹⁵N value was in fish (8.45 ± 0.177⁰/₀₀) and the lowest value of δ¹⁵N was in autochthonous aquatic macrophyte (2.00 ± 1.234⁰/₀₀). Based on the δ¹⁵N results, there are three trophic levels in the study river and it is suggested that the trophic chain begins with organic matter followed by group of insects and ends with fish (organic matter < insects < fish).     

Large scale of identification of differentially expressed genes in the regenerating rat liver after SISPH

After partial hepatectomy (PH), the remnant paren-chyma can completely recover lost liver mass and function in about one week[1,2]. Although adult hepa-tocytes are normally quiescent, they are readily primed to pass from G0 to G1 phase within 2―6 h after PH. The first peak of DNA synthesis appears 24 h after PH, while cell division peaks at 36 h. The liver cells then enter a second cell cycle, and redifferentiation and reconstruction of structure and function[3―6] take place. A great nu…     

Tricyclic HIV integrase inhibitors: potent and orally bioavailable C5-aza analogs

A series of C5-aza tricyclic HIV integrase inhibitors was prepared. A highly potent and orally bioavailable compound (compound 9) was identified and selected for development.     

Reproductive strategies in female polar and deep-sea bobtail squid genera <Emphasis Type="Italic">Rossia</Emphasis> and <Emphasis Type="Italic">Neorossia</Emphasis> (Cephalopoda: Sepiolidae)

Female reproductive features have been investigated in five polar and deep-sea bobtail squid genera Rossia and Neorossia (R. macrosoma, R. moelleri, R. pacifica, N.c. caroli and N.c. jeannae). These species are characterized by asynchronous ovary maturation, very large eggs (>10% ML), fecundity of several hundred oocytes, very high reproductive output, and continuous spawning with low batch fecundity. This adaptive complex of reproductive traits evolved in these small animals as an optimum strategy for polar and deep-water habitats.