Determination of glucose exchange rates and permeability of erythrocyte membrane in preeclampsia and subsequent oxidative stress-related protein damage using dynamic-<Superscript>19</Superscript>F-NMR

The cause of the pregnancy condition preeclampsia (PE) is thought to be endothelial dysfunction caused by oxidative stress. As abnormal glucose tolerance has also been associated with PE, we use a fluorinated-mimic of this metabolite to establish whether any oxidative damage to lipids and proteins in the erythrocyte membrane has increased cell membrane permeability. Data were acquired using ¹⁹F Dynamic-NMR (DNMR) to measure exchange of 3-fluoro-3-deoxyglucose (3-FDG) across the membrane of erythrocytes from 10 pregnant women (5 healthy control women, and 5 from women suffering from PE). Magnetisation transfer was measured using the 1D selective inversion and 2D EXSY pulse sequences, over a range of time delays. Integrated intensities from these experiments were used in matrix diagonalisation to estimate the values of the rate constants of exchange and membrane permeability. No significant differences were observed for the rate of exchange of 3-FDG and membrane permeability between healthy pregnant women and those suffering from PE, leading us to conclude that no oxidative damage had occurred at this carrier-protein site in the membrane.     

Cell source,differentiation, functional stimulation,and potential application of human thermogenic adipocytes in vitro

Recent investigations have showed that the functional thermogenic adipocytes are present in both infants and adult humans. Accumulating evidence suggests that the coexistence of classical and inducible brown (brite) adipocytes in humans at adulthood and these adipocytes function to generate heat from energy resulting in reducing body fat and improving glucose metabolism. Human thermogenic adipocytes can be differentiated in vitro from stem cells, cell lines, or adipose stromal vascular fraction. Pre-activated human brite adipocytes in vitro can maintain their thermogenic function in normal or obese immunodeficient mice; therefore, they improve glucose homeostasis and reduce fat mass in obese animals. These key findings have opened a new door to use in vitro thermogenic adipocytes as a cell therapy to prevent obesity and related disorders. Thus, this paper intends to highlight our knowledge in aspects of in vitro human brite/brown adipocytes for the further studies.     

Making metabolism accessible and meaningful: is the definition of a central metabolic dogma within reach?

Intermediary metabolism, a dominant research area before the emergence of molecular biology, is attracting renewed interest for fundamental and applied reasons as documented here. Nonetheless, the field may appear to be a thicket precluding entry to all but the most determined. Here we present a metabolic overview that makes this important and fascinating area accessible to a broad range of the molecular biological and biotechnological communities that are being attracted to biological problems crying out for metabolic solutions. This is accomplished by identifying seven key concepts, a so-called metabolic central dogma, that provide a core understanding analogous to the “Central Dogma of Molecular Biology” which focused upon maintenance and flow of genetic information.     

Optimization of cell culture conditions for production of biologically active proteins

We investigated the basic technology of cell culture conditions for production of useful substances such as cytokines, and related proteins produced by Namalwa cells. Namalwa cells (Klein, 1972), human B lymphoblastoid cells, were used for large scale production of alpha-interferon (Klein, 1979). Namalwa KJM-1, a subline of Namalwa cells, adapted to serum- and albumin-free medium, can grow at a high density above 1 × 10⁷ cells/ml in suspension mode by the use of a perfusion culture system, Biofermenter^?, containing a cone-type cell-sedimentation column as cell separator (Sato, 1983).Several kinds of cytokine cDNA can be introduced and expressed in Namalwa KJM-1 cells (Miyaji, 1990a,b,c). Some of these were produced in large quantities by use of a gene amplification method with dhfr (Miyaji, 1990c), even though the Namalwa KJM-1 cells contained endogenous dhfr genes. For stable production of the target protein, Namalwa KJM-1 cells are very useful host cells, because they have no effective endogenous protease activity in the conditioned medium.Using Biofermenter with micro-silicone fibers and a dialysis system, the specific productivity of the target proteins was not depressed at a high cell density.     

Behavior of acyl carrier proteins on western blots

Acyl carrier proteins (ACPs) from Escherichia coli and Euglena were analyzed on Western blots using rabbit antibodies raised against E. coli ACP. Euglena ACP, unlike that from E. coli, behaves upon electrophoresis under denaturing conditions as its size would predict. Oligomeric forms of both ACPs were evident on Western blots, but the bacterial ACP had more tendency to aggregate. That the oligomeric forms were not due to impurities was shown by their regeneration from low-Mr protein, reaction with antibodies isolated from low-Mr protein, and by molecular weight determination of the ACP by low-angle laser light scattering.