The axonal synthesis of phospholipids has been demonstrated in compartmented cultures of rat sympathetic neurons. In this model of neuron culture, metabolic events occurring in distal axons were studied independently of those occurring in cell bodies. Using radiolabeled tracers the axonal biosynthesis of the major membrane phospholipids and fatty acids but not cholesterol was detected. The capacity of axons for synthesis of phosphatidylcholine (PC), the major membrane lipid, was confirmed by the demonstration that key enzymes of PC biosynthesis were present in distal axons. A double-labeling experiment showed that at least 50% of axonal PC was synthesized locally in axons, with the remainder being made in cell bodies and transported into axons. The requirement of axonal PC synthesis for axonal elongation was investigated. When PC biosynthesis in distal axons alone was inhibited by two independent approaches (deprivation of choline or addition of the inhibitor hexadecylphosphocholine) axonal growth was markedly retarded. Our experiments demonstrated that PC synthesis in cell bodies was neither necessary nor sufficient for growth of distal axons, whereas local synthesis of PC in distal axons was required for normal axonal elongation.

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PubMedDate not supplied

Use of a low ionization potential dopant in a kinestatic charge detector: experimental findings.

The broadening of the line spread function (LSF) in the drift direction with increasing drift distance in the kinestatic charge detector is substantially reduced when small amounts (less than 1%) of trimethylamine [(CH3)3 N] are added to the x-ray detection medium (krypton or xenon). The LSF of a mixture of Kr and 0.01% trimethylamine (TMA) was measured as a function of distance at 15, 25, and 35 atm absolute pressure. The full width at half-maximum (FWHM) of the LSF was reduced from about 1.0 mm to less than 0.5 mm at a drift distance of 4.0 mm for the three pressures. The LSF's of mixtures of xenon and TMA at concentrations ranging from 0.0004% to 0.4% in one run and 0.06% to 4.0% in a second run were measured at a constant pressure of 20 atm. The FWHM of the LSF was reduced from 0.6 to 0.4 mm at 4.0 mm for the xenon measurements. The optimum concentration of TMA in Xe was found to be in the neighborhood of 0.1%. The use of TMA reduced the drift distance-dependent LSF broadening to the level expected from ionic diffusion, space charge repulsion, and electric field nonuniformity, and it may be possible to reduce the 0.4-mm FWHM plateau through the use of an improved Frisch grid design. Observation of negative charge carriers showed that electron attachment increases with increasing TMA concentration, although this could be caused by impurities in the TMA. The implications of these results are discussed in terms of extending the maximum drift distance attainable in a kinestatic charge detector.

Humans↗

PubMedDate not supplied

Lipoprotein lipase in cultured pig aortic smooth muscle cells.

Acetone powder extracts prepared from cultured pig aortic smooth muscle cells and the culture medium from these cells (particularly when incubated with heparin) were shown to contain a lipolytic enzyme which was identified as lipoprotein lipase by the following criteria: 1) stimulation by apolipoprotein C-II; 2) an optimal activity at approximately pH 8.0; 3) inhibition by NaCl, and 4) binding to a heparin-Sepharose affinity column. In addition, we found that cultured arterial smooth muscle cells from guinea pig and rabbit secreted a similar lipase into the culture medium. In contrast, studies using cultured bovine aortic endothelial cells yielded no evidence for either the synthesis or secretion of lipoprotein lipase by these cells. The production of lipoprotein lipase by the smooth muscle cells of the artery may play a role in the process of atherogenesis.

Acetone↗

PubMedDate not supplied

Neutral triglyceride lipase in macrophages.

High levels of neutral triglyceride lipase activity have been demonstrated in several types of macrophages (J774 cells, human monocyte/macrophages, rabbit alveolar macrophages, and resident mouse peritoneal macrophages). The pH optima ranged from 6.5 to 7.4 depending upon the buffer and the conditions of incubation. The addition of bovine serum albumin stimulated activity at low concentrations, as expected for a fatty acid-releasing reaction, but strongly inhibited at higher concentrations; maximal activity was observed in the presence of 0.625 mg/ml of bovine serum albumin. The enzyme was remarkably thermostable, showing no apparent loss of activity at 50 degrees C for as long as 6 hours. The lipase was inhibited 80% by 0.1 M NaCl. Assayed under optimal conditions, the specific activity of the neutral triglyceride lipase from J774 cells was more than 100-fold greater than the activity of lipoprotein lipase or neutral cholesterol esterase from those cells; this activity was 10-fold greater than the levels of hormone-sensitive lipase from 3T3-L1 adipocytes. This neutral triglyceride lipase may play an important role in the degradation and mobilization of cytosolic triglyceride in macrophage-derived foam cells.

Animals↗

PubMedDate not supplied

Intracellular trafficking of phospholipids: import of phosphatidylserine into mitochondria.

Three mechanisms are commonly suggested for the movemerlt of lipids between intracellular organelles: transfer mediated by cytosolic lipid transfer proteins, vesicle-mediated transfer, and transfer via regions of membrane continuity between organelles. The mechanism of translocation of phosphatidylserine from its site of synthesis on the endoplasmic reticulum (and related membranes) to the site of phosphatidylserine decarboxylation in the mitochondria has been investigated. Several experiments indicate that a transfer mediated by soluble cytosolic phospholipid transfer proteins, or by vesicles, is unlikely. Rather, the most likely mode of import of newly-synthesized phosphatidylserine into mitochondria is contact between the endoplasmic reticulum and mitochondrial membranes. In support of this mechanism we have isolated an endoplasmic reticulum-like."mitochondria-associated membrane" fraction and shown that it has some, but not all, properties of the endoplasmic reticulum. The mitochondria-associated membranes are enriched in lipid biosynthetic enzymes, especially phosphatidylserine synthase. When either phosphatidylserine translocation to mitochondria is blocked (by ATP depletion), or phosphatidylserine decarboxylation is blocked (with hydroxylamine), newly-synthesized phosphatidylserine accumulates in the mitochondria-associated membrane but not in microsomes, suggesting that phosphatidylserine traverses the mitochondria-associated membrane on its route from the endoplasmic reticulum to mitochondria.

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