Structural Bases of the Cytolytic Mechanisms of Entamoeba histolytica1

The cellular bases of the powerful cytolytic activity of the human protozoan parasite Entamoeba histolytica were explored by studying the effect of the virulent strain HM1:IMSS on epithelial monolayers of MDCK cells using a combination of time-lapse microcinematography and transmission and scanning electron microscopy. Early alterations of the epithelial cell membranes were detected by measuring changes in the transepithelial electrical resistance of MOCK monolayers mounted in Ussing chambers. The aggressive mechanism of E. histolytica trophozoites was found to be a complex, multifactorial phenomenon that included hit-and-run damage to the plasma membrane of effector cells mediated through contact, phagocytosis of lysed or apparently intact, but detached, MDCK cells, and inlracellular degradation of ingested cells. Following contact with amebas, the epithelial monolayers showed a pronounced lowering of transepithelial resistance, opening of tight junctions, distortion of microvilli, surface blebbing, and the presence of minute focal discontinuities in the plasma membrane. There was no evidence of amebic exocytosis, membrane fusion, or junction formation between the parasite and host plasma membranes. Although modifications in the epithelial cell membranes usually preceded lysis, the cytolytic activity of the parasite did not exclusively involve damage to the plasma membrane of the cultured host cells but also was mediated by avid phagocytosis, the displacement and separation of neighboring cells by means of pseudopodial activity, and the “pinching-off” of the peripheral cytoplasm of epithelial cells.     

A Strain of Eggplant Mosaic Virus Causing a Severe Disease of Tomato in Argentina

A virus causing a severe disease of tomatoes in Argentina was identified as a strain of eggplant mosaic virus (EMV). It resembles the type, Abelia latent and various Andean potato latent strains of EMV in its host range and its transmissibility at allow rate by Epitrix sp. It differs from these strains, however, serologically and in some of its cytopathic effects. In serological agar gel double diffusion tests it proved to be closely related to the tomato white necrosis isolate of EMV studied by Barradas (1983) in Brazil.     

Annexin V membrane interaction: an electrostatic potential study

The possible role of electrostatic interactions for membrane binding and pore formation of annexin V has been analysed on the basis of a simple dielectric model. It is suggested that the binding of phospholipids to annexin V is regulated, at least initially, by the protein's electrostatic potential. The calculations show that a strong local gradient of the electrostatic potential exists at the membrane-protein interface and a membrane pore may be generated by electroporation. The observed specificity and regulation of ion conduction is suggested to reside in the protein part of the pore. On the basis of the three-dimensional structures of the protein and its hypothetical membrane complex, and electrophysiological measurements, a mechanical model of the transmembrane voltage regulation of the annexin's ion conduction properties is proposed.     

Role of phosphatidylinositol turnover in alpha1 and of adenylate cyclase inhibition in alpha2 effects of catecholamines

Ligand binding and pharmacological studies have indicated that alpha-adrenergic receptors can be divided into alpha₁ and alpha₂. We suggest that alpha₁ receptors mediate those metabolic effects of alpha catecholamines which involve phosphatidylinositol turnover and the release of bound intracellular Ca²⁺ as well as the entry of extracellular Ca²⁺. In contrast, alpha effects of catecholamines are due to non-specific inhibition of adenylate cyclase through a mechanism independent of Ca²⁺. A similar classification for the effects of both histamine and serotonin suggests that they have separate type 1 or alpha receptors for Ca²⁺ dynamics which are different from type 2 or beta receptors which regulate adenylate cyclase.There is a significant correlation between hormone effects on phosphatidylinositol turnover and elevation of intracellular Ca²⁺. The available data suggest that the turnover of membrane-bound phosphatidylinositol is involved in Ca²⁺ gating in rat hepatocytes, rat and hamster adipocytes and blowfly salivary glands. In hamster adipocytes adenylate cyclase activity is also inhibited by alpha₂ catecholamines through a Ca²⁺ independent mechanism.     

Up-Regulation of the Neuronal Form of Nitric Oxide Synthase in Response to Prolonged Muscarinic M1 Receptor Stimulation

Abstract: It is generally believed that the neuronal form of nitric oxide synthase (nNOS) is constitutively expressed and that regulation of this enzyme's activity is mediated solely by changes in cytosolic calcium concentration. Serendipitously, however, we observed that pretreatment of Chinese hamster ovary (CHO) cells, which coexpress muscarinic M₁ receptors and nNOS, with 3.3 µ M or 1 m M carbachol (CCh) for 48 h resulted in marked enhancement of maximal muscarinic receptor-stimulated nNOS activity as determined by l -[³H]citrulline and cyclic [³H]GMP production. This was accompanied by a decrease in the potency of CCh. Muscarinic receptor density was reduced in the agonist-pretreated cells, as determined by specific [ N-methyl -³H]scopolamine methyl chloride binding, whereas competition binding studies revealed no changes in agonist affinity. Both receptor-stimulated inositol phosphate formation and elevation of intracellular calcium concentrations were found to be desensitized in agonist-pretreated cells in a manner dependent on CCh pretreatment concentration. It is interesting that ionomycin-stimulated nNOS activity was greater in CCh-pretreated cells. Also, western analysis revealed increased nNOS immunoreactivity in pretreated cells. A similar increase in nNOS immunoreactivity following agonist treatment was demonstrated in N1E-115 neuroblastoma cells, which endogenously express nNOS and muscarinic M₁ receptors. Thus, the enhancement of maximal receptor-stimulated nNOS activity following agonist pretreatment can be attributed to up-regulation of nNOS. It is interesting that this augmentation of the response takes place in spite of receptor down-regulation and desensitization of multiple steps involved in nNOS activation.     

The Effects of Snake Predation on Metamorphosis of Western Toads,Bufo boreas (Amphibia,Bufonidae)

For some anuran species, synchronous metamorphosis may function as an antipredator adaptation by swamping predators during the period of transformation. We examined the levels of synchrony of emergence from the water of metamorphosing western toads (Bufo boreas) in the presence and absence of a live snake predator, the common garter snake (Thamnophis sirtalis) in a laboratory experiment. To compare between the treatments, we measured the time to emergence from the water, the number of metamorphs emerging together, and the level of aggregation (before and during emergence) of the toads in each treatment. There was a difference between the treatments when all three factors were considered. We attributed these differences to a behavioral response in which B. boreas emerged sooner in the presence of the predator, regardless of whether individual toads had reached the point at which they were physically better suited to the terrestrial environment than the larval environment.     

The Atapuerca sites and the ibeas hominids

The Atapuerca railway Trench and Ibeas sites near Burgos, Spain, are cave fillings that include a series of deposits ranging from below the Matuyama/Bruhnes reversal up to the end of Middle Pleistocene. The lowest fossil-bearing bed in the Trench contains an assemblage of large and small Mammals includingMimomys savini, Pitymys gregaloides, Pliomys episcopalis, Crocuta crocuta, Dama sp. and Megacerini; the uppermost assemblage includesCanis lupus, Lynx spelaea, Panthera (Leo) fossilis, Felis sylvestris, Equus caballus steinbeimensis, E.c. germanicus, Pitymys subterraneus, Microtus arvalis agrestis, Pliomys lenki, and alsoPanthera toscana, Dicerorbinus hemitoechus, Bison schoetensacki, which are equally present in the lowest level. The biostratigraphic correlation and dates of the sites are briefly discussed, as are the paleoclimatic interpretation of the Trench sequences. Stone artifacts are found in several layers; the earliest occurrences correspond to the upper beds containingMimomys savini. A set of preserved human occupation floors has been excavated in the top fossil-bearing beds. The stone-tool assemblages of the upper levels are of upper-medial Acheulean to Charentian tradition. The rich bone breccia SH, in the Cueva Mayor-Cueva del Silo, Ibeas de Juarros, is a derived deposit, due to a mud flow that dispersed and carried the skeletons of many carnivores and humans. The taxa represented are:Ursus deningeri (largely dominant),Panthera (Leo) fossilis, Vulpes vulpes, Homo sapiens var. Several traits of both mandibular and cranial remains are summarized. Preliminary attempts at dating suggest that the Ibeas fossil man is older than the Last Interglacial, or oxygen-isotope stage 5.     

Preparative synthesis of C-(alpha-D-glucopyranosyl)-alkenes and -alkadienes: Diels-Alder reaction

The reaction of 2,3,4,6-tetra-O-benzyl-1-O-(p-nitrobenzoyl)-alpha-D-glucopyranose with (E)-penta-2,4-dienyltrimethylsilane and boron trifluoride etherate in acetonitrile afforded stereoselectively (E)-5-(tetra-O-benzyl-alpha-D-glucopyranosyl)-1,3-pentadiene in good yield. The readily available penta-O-benzoyl-alpha-D-glucopyranose reacted with allyltrimethylsilane in the presence of boron trifluoride etherate in acetonitrile to give 3-(tetra-O-benzoyl-alpha-D-glucopyranosyl)-1-propene and its beta anomer in yields of 60% and 2.3%, respectively. Diels-Alder cycloaddition of maleic anhydride to diene 1 afforded the adduct cis,cis-3-(tetra-O-benzyl-alpha-D-glucopyranosylmethyl)cyclohex -4-ene- 1,2-dicarboxylic anhydride in high yield.