Adenine deaminase of a eukaryotic animal cell,Crithidia fasciculata

Adenine deaminase (adenine aminohydrolase, EC 3.5.4.2) has been found to occur in Crithidia fasciculata with a specific activity higher than that of the same enzymes of bacteria and yeasts. It is remarkable for its stability to heat, exhibiting no appreciable loss of activity after 60 min at 55 °C. It occurs in the soluble portion of cell extracts but can be released into the suspending medium by osmotic and/or cold shock.     

Beta 1 integrin (CD29) expression on human postnatal T cell subsets defined by selective CD45 isoform expression

The integrin beta 1 (CD29) is a marker for total very late activation Ag integrins on cells, and exhibits considerable fluctuation in cell surface density at various stages of T cell development. We have analyzed beta 1 integrin expression on subsets of human thymus, and on T cells from healthy babies and children, in comparison to healthy adults aged 26 to 75. T cells from adult peripheral blood include a CD29-, a CD29lo, and a CD29hi set. Compared with adults, PBMC T cells from children have reduced numbers of both CD29lo and CD29hi subsets but equivalent numbers of CD29- T cells. The number of CD29hi T cells increases gradually with age, achieving adult levels only at about 26 yr of age; in aged adults (69 to 75 yr), nearly all T cells have a CD29hi phenotype. Most thymocytes and cord blood T cells, in contrast, have a single peak of CD29 staining that is intermediate to the two peaks seen in adults. Multi-negative progenitor and CD45RO- thymocytes (presumptive thymic generative line-age) are 98% CD29hi. Progenitor thymocytes and adult PBMC T cells express equivalent amounts of beta 1 and alpha 4, but progenitors are alpha 5hi, whereas PBMC T cells are alpha 5lo. T cells from children have reduced beta 1hi and alpha 5lo, but nearly comparable numbers of alpha 4hi. This suggests that the major very late activation Ag integrins during childhood may be alpha 5 beta 1 and alpha 4 complexed with an alternate beta chain. In children, the majority of CD29hi cells are also CD45RAhi, in contrast to the pattern in adults, in whom the majority of CD29hi T cells are CD45RA-. This suggests that in children, the main defense against infection may reside in the CD29hi45RAhi T cells, which have not yet made the transition to CD45RO and to bona fide memory status. The proliferative response to tetanus toxoid of 4- to 6-mo-old babies correlates with the number of CD29hi45RAhi T cells, suggesting that it derives at least in part from cells that do not express a "memory" phenotype. These observations show a pattern of alternating high and low density CD29 during T cell development, which is consistent with the idea that CD29 is a marker for functionally defined T cell sets. Analysis of the CD29 expression of CD29hi thymocytes developing in vitro supports this view. We suggest that the intensity of CD29 expression on a T cell varies, dependent upon the microenvironmental interactions required by a differentiating T cell.     

Cerebral Metabolic Effects of Monoamine Oxidase Inhibition in Normal and 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Acutely Treated Monkeys

The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces dopaminergic cell death in the substantia nigra pars compacta (SNpc) and clinical parkinsonism in humans and experimental animals. Pretreatment with monoamine oxidase inhibitors prevents this cell death and associated parkinsonism by blocking the oxidation of MPTP to a toxic intermediate. The 2-deoxyglucose method was used to study the acute effects of MPTP in the monkey brain and the effects of monoamine oxidase inhibition on local cerebral glucose utilization in both normal and MPTP-treated monkeys. MPTP administration alone caused a major increase in glucose utilization in the SNpc and smaller increases in some subnuclei within the ventral tegmental area in which eventual dopaminergic cell loss also occurs. Pretreatment with pargyline abolished these metabolic increases, a finding suggesting both that the oxidized product of MPTP generates the metabolic increases and that the increased glucose consumption may contribute to cell toxicity. On the other hand, in most cortical, thalamic, striatal, brainstem, and cerebellar areas MPTP alone caused reductions in glucose utilization, and pargyline failed to prevent these effects. Pargyline alone depressed metabolism in the locus coeruleus and a few other monoaminergic structures.     

Bovine trophoblastic cell vesicle attachment to polarized endometrial epithelial cells in vitro

Summary Interactions between bovine trophoblastic cell vesicles and bovine endometrial epithelial cells were investigated by light and electron microscopy and lectin histochemistry in a cell culture model of early blastocyst attachment. Primary lines of bovine endometrial epithelial cells were polarized by subculturing on substrata and maintaining cultures at the air-medium interface. Trophoblastic cell vesicles were obtained from elongated Day 14 blastocysts. In co-cultures, trophoblastic cell vesicles adhered to endometrial epithelial cells through microvillus interdigitation and formation of primitive membrane junctional complexes. After 3 d in co-culture, a multilayered cellular plaque formed at the trophoblastic cell-endometrial epithelial cell interface. The type of cells contributing to this local proliferative response could not be identified specifically as trophoblastic or endometrial cells, and areas of membrane fusion between cells were noted. Ultrastructural features of vesicle adhesion in cultures were similar to features of conceptus attachment in vivo. Lectins bound to apical membranes of trophoblastic cells and endometrial epithelial cells in all locations except contact sites between vesicles and endometrial cells. These findings suggest that local cellular proliferation and membrane fusion between trophoblastic and endometrial epithelial cells may be early events in conceptus implantation in the cow and these events can be reproduced in culture. This work was supported by a grant from U.S. Department of Agriculture Animal Health and Disease Program, Washington, DC.     

Conformational changes of recombinant human granulocyte-colony stimulating factor induced bypH and guanidine hydrochloride

Fluorescence and circular dichroism were used to follow thepH-dependent conformational changes of granulocyte colony stimulating factor (G-CSF). Tryptophan fluorescence of the spectra monitored at 344 nm, or after deconvolution of the emission spectra, at 345 nm, showed a decrease in intensity on going frompH 7 to 4, with a midtransitionpH of 5.8. On the other hand, tyrosine fluorescence measured either by the ratio of intensity at 308 nm to that at 344 nm, or by the fluorescence intensity at 303 nm after deconvolution of the spectra, increased in intensity as thepH was changed from 6 to 2.5, with a midtransitionpH of 4.5. Near UV circular dichroic spectra also showed changes betweenpH 7.5 and 4.5, which correlated with the transition monitored by the tryptophan fluorescence. The guanidine hydrochloride-induced conformational changes of G-CSF at fivepH values from 2.5 to 7.5 were also studied. Circular dichroic and fluorescence spectra revealed minor conformational changes by the addition of 1 or 2 M guanidine HCl at allpH values examined, while the major conformational transition occurred between 2 and 4 M guanidine hydrochloride. The secondary structure of the protein was most stable betweenpH 3.3 and 4.5. The guanidine HCl-induced denaturation of G-CSF involved more than a two-state transition, with detectable intermediate(s) present, and the structure of the intermediate(s) appeared to depend on thepH used. These results are consistent with thepH dependence of the structure described above, and demonstrate the complex conformational properties of G-CSF.     

The N-terminal amino acid sequence of the beta-subunit of the legumin-like protein from seeds of Ginkgo biloba.

The sequence of the first 52 amino acids at the N-terminus of the beta-subunit of a legumin-like protein from seeds of the gymnosperm Ginkgo biloba were determined by automated sequencing and DABITC/PITC microsequence analyses of peptides derived from the protein by enzymatic digestions and chemical cleavage with CNBr. The protein from Ginkgo exhibits sequence homologies (32-49% identities) with the 11S globulins and legume-like proteins from seeds of various angiosperm monocotyledons and dicotyledons.     

An electron microscopic study of picoplanktonic organisms from a Small Lake

Picoplankton, both prokaryotic and eukaryotic, are distinguished from other aquatic organisms by their small size (0.1–2.0 m). Such organisms were recovered from waters of a small oligotrophic lake using screens, filters, and high-speed centrifugation. The majority of the picoplankton were unable to form visible colonies on common media. Cells examined in thin sections by electron microscopy showed that 60–75% of the cells had an average diameter after dehydration of 0.48–0.51 m. The maximum dimensions of the rest of the cells ranged from 0.56–1.81 m. Using details of ultrastructure, cells were classified as prokaryotic or eukaryotic. Phototrophs present included two cyanobacterial morphotypes (5–6%) and two eukaryotic algae (less than I%). The arrays of intracytoplasmic membranes in 18–20% of the cells were suggestive of methanotrophic rods and chemoautotrophs. Relatively few prosthecate bacteria were observed in the water column samples. The smallest cells (1–2%) contained magnetosomes, the presence of which were confirmed by x-ray spectroscopy. Iron was also detected in the envelopes of some rod shaped cells by the same technique. The study of in situ picoplankton populations using TEM coupled with other techniques may provide better understanding of picoplankton biomass.     

Cerebral energy metabolism in immature and mature guinea pig fetuses during acute asphyxia.

In immature fetuses circulatory centralization caused by acute asphyxia is less effective than that in mature fetuses (Jensen & Berger, 1991). This suggests that cerebral oxygenation may be poor in immature fetuses during asphyxia. On the other hand cerebral oxygen consumption is lower in immature than that in mature fetuses. To determine, whether or not there is an imbalance between oxygen supply and demand in one or the other group, we compared the time course of the changes of cerebral concentrations of both high-energy phosphates and glycolytic intermediates between immature and mature guinea pig fetuses during acute asphyxia caused by arrest of uterine blood flow. The fall in the cerebral concentrations of adenosine triphosphate and glucose, and the rise in those of adenosine monophosphate and lactate were slower in immature than in mature fetuses. There were no differences between the levels of cerebral adenosine diphosphate and creatine phosphate of the two groups. From these results we conclude that during acute asphyxia the imbalance between cerebral oxygen supply and demand is less marked in immature than in mature fetuses.