Action Of Lysolecithin Research Articles

Role of various phospholipases A2 and inhibitors in the pathogenesis and prevention of pancreatic acinar cell necrosis: studies with isolated rat pancreatic acini.

Phospholipase A2 (PLA2) may play a central role in the pathogenesis of pancreatic acinar cell necrosis. Several questions, however, are unsolved: Is acinar cell necrosis caused by PLA2 derived from infiltrating leukocytes or from pancreatic PLA2 itself? Does PLA2 cause cellular lysis by the release of lysolecithin from lecithin or by generation of free radicals? The aims of this study were to determine which form of PLA2 is responsible for cellular damage and how to inhibit its action. Isolated rat pancreatic acini were prepared by collagenase digestion. Newly synthesized proteins were labeled by 35S-methionine. Acini were incubated in buffer to which various factors, such as porcine pancreatic PLA2 or bee venom PLA2, homogenates of either leukocytes or pancreatic homogenates, all with or without lecithin and with or without potential inhibitors (aprotinin, 4-bromophenacylbromide, BM 16.2115, quinacrine, various analogs of arachidonic acid), or free radicals (hydrogen peroxide, xanthine/ xanthine oxidase) with or without allo-purinol or dismutase/catalase were added. Cellular destruction was measured by the release of radiolabeled proteins. PLA2 alone, free radicals, and granulocytes were not harmful to acini within 30 min of incubation. Free radicals caused significant release of radiolabeled proteins only after 3 h of incubation; this release could be inhibited by scavengers. Incubation of pancreatic acini with PLA2 in combination with lecithin caused rapid release of radiolabeled proteins. Addition of high concentrations of enterokinase activated pancreatic homogenates both alone and with lecithin caused release of cellular proteins, suggesting that pancreatic PLA2 uses lecithin from pancreatic membranes as substrate. Almost all tested potential inhibitors of PLA2 were unable to prevent the destruction caused by either pancreatic or bee venom PLA2 and lecithin. However, HK 42, a polyunsaturated fatty acid analog, was able to reduce dose dependently the release of acinar proteins caused by pancreatic PLA2 and lecithin. Pancreatic PLA2 and not PLA2 from infiltrating leukocytes may play a role in pancreatic acinar cell necrosis. Cellular lysis is caused upon the action of lysolecithin and probably not via the action of free radicals.

Properties of potassium and sodium channels in frog internode.

1. Voltage-clamp experiments were performed on single frog internodes after acute demyelination with lysolecithin. The action of lysolecithin was stopped by washing out the lysolecithin with normal Ringer solution containing bovine albumin when the first delayed current was observed. After washing, the temperature was lowered from 25 to 15 degrees C. These procedures greatly prolonged the survival of the demyelinated internode up to 1 h. 2. External tetraethylammonium chloride (TEA+, 110 mM) reduced the K+ current in the internode only to 11% of the control value. 110 mM-TEA+ increased the time constant tau n of K+ activation by a factor of two in the node and by a factor of four in the internode. 120 mM-CsCl at the cut ends of the fibre also reduced the delayed outward current recorded at 60 mV in the internode to 11% of the control value, hardly changing the time constant tau n. 3. After a depolarization, the K+ tail current decayed in two phases, suggesting that the K+ conductance of the internodal membrane may be composed of at least two components, a slow one (gKs) and a fast one (gKf). As in the node, the fast K+ conductance of the internode can be further decomposed into two components (gKf1 and gKf2) with different activation potential ranges. The fast phase of the tail current was blocked by external application of 1 mM-4-aminopyridine (4-AP). The slow phase was almost unaltered by 1 mM-4-AP. The extrapolated slow tail current was 33% of the total tail current in the internode and 15% at the node, i.e. the proportion of slow K+ channels is larger in the internode than in the node. 4. Tetrodotoxin (TTX)-sensitive transient inward currents could be measured in the demyelinated internode, provided the large K+ currents were blocked by internal Cs+. The time course, TTX sensitivity, reversal potential and steady-state inactivation of the transient early inward current indicate that this current is caused mainly by Na+ passing through Na+ channels. 5. The density of K+ and Na+ channels in the demyelinated internode is estimated from the size of the K+ and Na+ current, respectively, and the capacity of the demyelinated segment. The K+ channel density of the internode seems to be about 20 times smaller than in the node, whereas the Na+ channel density in the internode appears to be about 500 times smaller than in the node.

Experimental pancreatitis in the rat. Light and electron microscopical observations on early pancreatic lesions induced by intraductal injection of trypsin, phospholipase A2, lysolecithin and non-ionic detergent

Trypsin, phospholipase A2, lysolecithin or non-ionic detergent polyoxyethylene p-t-octyl phenol solutions were injected into the rat biliopancreatic duct. Histological and ultrastructural changes in the gland were studied 15 min and 3 h after the injections. The rough surfaced endoplasmic reticulum disintegrated in two ways: (1) the endoplasmic reticulum in the cell periphery was vesiculated but ribosomes were well preserved at 15 min, and (2) large, round membranous structures appeared in apical cytoplasm at 3 h. Zymogen granules disintegrated in the second type, which possibly represents autodigestion. Both types of injury lead ultimately to structureless necrosis. Lesions induced by phospholipase A2 and lysolecithin were identical. Trypsin-induced damage developed slowly and the two phases of endoplasmic reticulum disintegration were not sharply separable. Lesions caused by polyoxyethylene p-t-octyl phenol were variable at 15 min, but at 3 h the type 2 injury described above was observed. It was concluded that although the initial damage in pancreatic acinar cells may vary, necrotic changes are similar despite the injected material at the later time interval. During acute pancreatitis, the acinar cell necrosis is most probably due to the action of lysolecithin produced by the activation of phospholipase A2.

Mechanisms for albumin-mediated membrane damage.

1. Bee venom phospholipase A2 causes a small increase in the sublytic leakiness of rabbit erythrocytes which is strongly inhibited by lysolecithin and potentiated by albumin. 2. Albumin extracts long-chain fatty acids from the erythrocyte membrane by a process which is faster than diffusion-determined efflux of fatty acid and during the process it greatly increases the sublytic leakiness of the membrane. In contrast, albumin rapidly terminates sublytic responses to lysolecithin. 3. These actions of albumin are abolished only when albumin is preloaded with four molecules of oleic acid per protein molecule. 4. A minor proportion of the response of phospholipase-A2-treated cells to albumin is due to accumulated fatty acid products; the combination of active enzyme and accumulated fatty acids is much more effective than either alone. Membrane partially depleted of phospholipids by enzyme attack is not highly leaky. 5. Albumin may, in part, act by removing lysolecithin, the inhibitory reaction product. A model is proposed in which susceptibility to phospholipase A2 attack is not uniform on the cell surface and is used to explain the inhibitory action of lysolecithin and the high sensitivity of the membrane in the presence of albumin to damage by newly released fatty acids.

Action of lysolecithin on normal and leukemic human leukocytes.

Action of lysolecithin on normal and leukemic human leukocytes.

Research on brain proteins during the action of lysolecithin on brain homogenates

Bei Zusatz von Lysolecithin in einer 7–8 mM Endkonzentration zu Hirnhomogenaten erfolgt eine Vermehrung des wasserloslichen, extrahierbaren Proteins um den Faktor 3–5.