[The evaluation of patients with ischemic cerebral lesions by CT, SPECT and qEEG in acute, subacute and chronic phases]. / Evalución mediante TC, SPECT y qEEG de pacientes con lesiones isquémicas cerebrales durante las fases aguda, subaguda y crónica.

INTRODUCTION: SPECT, EEG AND CT scan offer information with several pathophysiologic meanings. Their results vary with time and according to the vascular affected territory. OBJECTIVE: We wanted to study how the sensibility varies and the relationship with the clinic of SPECT, qEEG and CT scan in the acute, subacute and chronic stages and according to the vascular affected territory. We also wanted to analyze the several pathophysiologic aspects of the cerebral ischemia. METHODS: Thirty-six patients with symptoms of hemispheric stroke were evaluated with CT scan, qEEG, SPECT99mTc-HMPAO during the acute (0-5 days), subacute (0-15 days) and chronic (16 days to 1 year) stages. RESULTS: The decrease of ipsilateral CBF depend on the time (p = 0.0061), being not very frequent during the two first weeks. The qEEG was the most sensitive study in the first phase, its sensibility did not depend on the vascular affected territory and was dependent on the time (p = 0.0011), diminishing in the chronic phase. The slow activity was habitually ipsilateral. The CT scan was the less sensitive study. CONCLUSION: After 24 hours and until the second week, there is habitually an increase of the ipsilateral rCBF. The luxury perfusion could explain the fogging effect in the CT scan. The slow activity of the qEEG represents the alteration of the oxygen metabolism. The interpretation of the variation of the CBF and the qEEG allow us to define oligemia of the ischemia and between reactive hyperemia and the increase of CBF due to the necrotic tissue.

Evaluación mediante TC, SPECT y qEEG de pacientes con lesiones isquémicas cerebrales durante las fases aguda, subaguda y crónica / Evaluation of patients with ischemic cerebral lesions using TC, SPECT and qEEG during acute, subacute and chronic phases

Introducci›n. La SPECT, el EEG y la TC brindan informaci›n con diferente significado fisiopatol›gico; sus resultados var­an en el tiempo y segœn el territorio vascular afectado. Objetivos. En este estudio nos propusimos estudiar c›mo var­a la sensibilidad y la relaci›n con la cl­nica de la SPECT, el qEEG y la TC en las etapas aguda, subaguda y cr›nica, segœn el territorio vascular afectado. Asimismo, analizamos diferentes aspectos fisiopatol›gicos de las ECV isqu micas. M todos. Se realiz› un estudio con TC, qEEG, SPECT 99mTc-HMPAO durante las etapas aguda (0-5 d­as), subaguda (6-15 d­as) y cr›nica (de 16 d­as a 1 a o) a 36 pacientes. Resultados. La disminuci›n ipsilaeral del FSC dependi› del tiempo (p=0,0061), siendo poco frecuente durante las dos primeras semanas. El qEEG fue el estudio mÿs sensible en la primera fase, su sensibilidad no estuvo relacionada con el territorio vascular afectado y si dependi› del tiempo (p=0,0011), disminuyendo en la fase cr›nica. La actividad lenta habitualmente fu ipsilateral. La TC de crÿneo result› ser el estudio menos sensible. Conclusiones. Despu s de las 24 horas y hasta la segunda semana, habitualmente se produce un aumento del FSCr ipsilateral. la perfusi›n de lujo podr­a explicar el efecto fogging en la TC simple de crÿneo. La actividad lenta del qEEG representa la alteraci›n del metabolismo de ox­geno. La interpretaci›n de la variaci›n del FSC y del qEEG permite delimitar entre la perfusi›n de miseria de tipo oligo mica de la isqu mica, as­ como la hiperemia reactiva del aumento del FSC debido a la necrosis tisular

Evaluaci›n mediante TC, SPECT y qEEG de pacientes con lesiones isqu micas cerebrales durante las fases aguda, subaguda y cr›nica / Evaluation of patients with ischemic cerebral lesions using TC, SPECT and qEEG during acute, subacute and chronic phases

Introducci›n. La SPECT, el EEG y la TC brindan informaci›n con diferente significado fisiopatol›gico; sus resultados var­an en el tiempo y segœn el territorio vascular afectado. Objetivos. En este estudio nos propusimos estudiar c›mo var­a la sensibilidad y la relaci›n con la cl­nica de la SPECT, el qEEG y la TC en las etapas aguda, subaguda y cr›nica, segœn el territorio vascular afectado. Asimismo, analizamos diferentes aspectos fisiopatol›gicos de las ECV isqu micas. M todos. Se realiz› un estudio con TC, qEEG, SPECT 99mTc-HMPAO durante las etapas aguda (0-5 d­as), subaguda (6-15 d­as) y cr›nica (de 16 d­as a 1 a o) a 36 pacientes. Resultados. La disminuci›n ipsilaeral del FSC dependi› del tiempo (p=0,0061), siendo poco frecuente durante las dos primeras semanas. El qEEG fue el estudio mÿs sensible en la primera fase, su sensibilidad no estuvo relacionada con el territorio vascular afectado y si dependi› del tiempo (p=0,0011), disminuyendo en la fase cr›nica. La actividad lenta habitualmente fu ipsilateral. La TC de crÿneo result› ser el estudio menos sensible. Conclusiones. Despu s de las 24 horas y hasta la segunda semana, habitualmente se produce un aumento del FSCr ipsilateral. la perfusi›n de lujo podr­a explicar el efecto fogging en la TC simple de crÿneo. La actividad lenta del qEEG representa la alteraci›n del metabolismo de ox­geno. La interpretaci›n de la variaci›n del FSC y del qEEG permite delimitar entre la perfusi›n de miseria de tipo oligo mica de la isqu mica, as­ como la hiperemia reactiva del aumento del FSC debido a la necrosis tisular

Local rules for protein folding on a triangular lattice and generalized hydrophobicity in the HP model.

We consider the problem of determining the three-dimensional folding of a protein given its one-dimensional amino acid sequence. We use the HP model for protein folding proposed by Dill (1985), which models protein as a chain of amino acid residues that are either hydrophobic or polar, and hydrophobic interactions are the dominant initial driving force for the protein folding. Hart and Istrail (1996a) gave approximation algorithms for folding proteins on the cubic lattice under the HP model. In this paper, we examine the choice of a lattice by considering its algorithmic and geometric implications and argue that the triangular lattice is a more reasonable choice. We present a set of folding rules for a triangular lattice and analyze the approximation ratio they achieve. In addition, we introduce a generalization of the HP model to account for residues having different levels of hydrophobicity. After describing the biological foundation for this generalization, we show that in the new model we are able to achieve similar constant factor approximation guarantees on the triangular lattice as were achieved in the standard HP model. While the structures derived from our folding rules are probably still far from biological reality, we hope that having a set of folding rules with different properties will yield more interesting folds when combined.

Recognizing circular decomposable metrics.

Circular decomposable metrics (CDMs) have been used in phylogenetic studies. The fastest algorithm for recognizing a CDM runs in time O(n5), given an n x n table of pairwise distances. We give an O(n2) time algorithm for this problem.

Numerical taxonomy on data: experimental results.

We consider the problem of fitting an n x n distance matrix D by a tree metric T. This problem is NP-hard for most reasonable distance functions between D and T. Recently, an approximation algorithm was presented (Agarwala et al., 1996) which achieves a factor of 3 approximation to the L infinity best fitting tree. We call this method the Single Pivot (SP) heuristic. Within the biology community, the so-called Neighbor-Joining (NJ) heuristic (Saitou and Nei, 1987) has wide acceptance. In this paper, we introduced a new Double Pivot (DP) heuristic, which is an extension of the SP heuristic, and show that DP outperforms NJ on biological and random data.

A monoclonal antibody specifically modulates dihydropyridine-sensitive calcium current in BC3H1 myocytes.

The nonfusing muscle cell line BC3H1 expresses functional Na and Ca channels similar to those found in skeletal muscle (1). We have utilized a monoclonal antibody that cross-reacts with a 45-50-kDa glycoprotein in differentiated BC3H1 myocytes but not in cardiac or neuronal cells. This antibody, MAb1223, specifically modulates 45Ca influx and slowly activating, dihydropyridine-sensitive Ca currents when added to BC3H1 myocytes. Low-threshold, dihydropyridine-insensitive Ca currents and Na and K currents are unaffected. MAb1223 action is voltage dependent. At very negative holding potentials, similar to those of the resting cell, MAb1223 increases slow Ca current without significant changes in the voltage dependence of activation. At more positive potentials, at which channel opening probability is reduced by inactivation, exposure to MAb1223 reduces current. Agonist and antagonist dihydropyridines modulate the action of MAb1223 on the slow Ca channel in a manner that suggests that their respective binding sites may interact directly with the channel.

Differential effects of p-nitrophenyl-D-xylosides on mouse blastocysts and uterine epithelial cells.

A number of studies have implicated glycoconjugates in cell recognition events associated with implantation of mammalian blastocysts into the uterus. We have found that p-nitrophenyl-D-xylosides inhibit mouse embryo attachment and outgrowth on monolayers of uterine epithelial cells when cocultured in vitro. Inhibition of attachment and trophoblast formation by alpha- and beta-xylosides was observed in embryos cultured on tissue culture plastic in serum containing medium or on monolayers of epithelial cells. The biochemical basis for this inhibition has been investigated. Consistent with their accepted mode of action, beta- but not alpha-D-xylosides greatly stimulated glycosaminoglycan chain production by uterine epithelial cells and likewise reduced proteoglycan assembly. In contrast, both alpha- and beta-anomers selectively inhibited embryo attachment and outgrowth without stimulating glycosaminoglycan chain production by embryos. The inhibitory effect of the xylosides on embryos was reversible and did not require concentrations that reduced the rate of protein synthesis. Both alpha- and beta-D-xylosides inhibited the synthesis of proteoglycans including heparan sulfate as well as certain other glycoconjugates by embryos. Collectively, these data indicate that proper assembly of glycoconjugates, including proteoglycans, is required for implantation-related processes, although the inhibition of embryo outgrowth by xylosides may be by an as yet uncharacterized mechanism.

Heparin/heparan sulfate is involved in attachment and spreading of mouse embryos in vitro.

The involvement of embryonic cell surface proteoglycans in the attachment and outgrowth of cultured mouse embryos has been investigated. Several lines of evidence indicate that periimplantation stage blastocysts express heparin/heparan sulfate proteoglycans on their cell surfaces that can mediate embryo attachment and trophoblast outgrowth on a variety of matrices. First, in the presence of soluble heparin, the rate at which embryos attach and outgrow on laminin, fibronectin, or monolayers of uterine epithelial cells is reduced considerably. In the case of fibronectin, the rate of outgrowth in the presence of the heparin is slower than in the presence of the Arg-Gly-Asp-Ser-containing peptide that is recognized by a fibronectin receptor. Embryos also attach and exhibit a limited ability to outgrow on platelet factor IV, a heparin binding protein that does not possess the additional binding domains of laminin or fibronectin. Attachment on platelet factor IV is inhibited by heparin. Second, cell surface digestion of attachment-component embryos with heparinase, but not chondroitinase ABC, slows the rate of outgrowth on tissue culture plates in the presence of serum. Third, selective staining for sulfated molecules on the trophectoderm surface of periimplantation stage embryos indicates that such molecules are abundant and uniformly distributed on these cell surfaces. Last, heparin/heparan sulfate proteoglycans are detected as major cell surface components of embryos using vectorial labeling with lactoperoxidase and Na125I. Collectively, these data indicate that heparin/heparan sulfate-bearing molecules have a direct role in attachment and outgrowth of implantation stage blastocysts.

Developmental expression of a cell-surface protein involved in calcium uptake and skeleton formation in sea urchin embryos.

The developmental expression of a cell-surface protein involved in Ca2+ accumulation and skeleton formation in sea urchin embryos has been studied. In Strongylocentrotus purpuratus, this protein is present in the egg and in all cell types of the early embryo. After gastrulation, its synthesis and expression are restricted to the skeleton-forming primary mesenchyme cells. In Lytechinus pictus, the protein cannot be detected in eggs or in embryos until the mesenchyme blastula stage. Hybrid embryos demonstrate a pattern of expression indistinguishable from that of the species contributing the maternal genome, which suggests that early expression of the protein in S. purpuratus embryos is due to utilization of maternal transcripts from the egg. Later expression of this protein in primary mesenchyme cells is the result of cell-type-specific synthesis, likely encoded by embryonic transcripts. This cell-type-specific expression in primary mesenchyme cells correlates temporally with Ca2+ accumulation during skeleton formation in the embryo.

A monoclonal antibody inhibits calcium accumulation and skeleton formation in cultured embryonic cells of the sea urchin.

The assembly of the spicules (primitive skeleton) of the sea urchin embryo is being studied in primary mesenchyme cells cultured in vitro. A monoclonal antibody (1223) has been prepared that inhibits the deposition of CaCO3 into the spicules. This antibody reacts with a 130,000 Mr cell-surface protein that is concentrated on the surface of approximately 5% of the cells of dissociated gastrula stage embryos. When primary mesenchyme cells in the embryo or cells cultured in vitro are examined, the 1223 antigen is detected on the surface of the cells and on the extracellular material associated with the spicule. We conclude that the 1223 antibody recognizes a cell-surface protein that plays an essential role in spicule formation.

Reductive methylation as a probe of the heat-labile alpha-bungarotoxin-acetylcholine receptor membrane complex: evidence for surface interactions.

alpha-Bungarotoxin (alpha-Bgt) is a potent postsynaptic neurotoxin which blocks neurotransmission by binding very tightly to the acetylcholine-receptor (AcChR) protein. We have previously shown (P. Calvo-Fernandez, and M. Martinez-Carrion (1981) Arch. Biochem. Biophys. 208, 154-159) that alpha-Bgt free in its native solution conformation incorporates 12 methyl groups when reductively methylated using formaldehyde and sodium cyanoborohydride. We now show that when the alpha-Bgt molecule is bound to the AcChR contained in native membranes prepared from Torpedo californica electroplax, the number of accessible methylation sites is significantly reduced. This favors a model of alpha-Bgt-AcChR interaction involving significant numbers of lysyl moieties distributed over a reasonably large surface of the toxin molecule. In addition, this paper presents a novel procedure for the rapid and nondestructive dissociation of the toxin-AcChR membrane complex which takes advantage of the thermal instability of the complex.

Ligand-induced effects at regions of acetylcholine receptor accessible to membrane lipids.

The effectiveness of fluorescence quenching of pyrene-1-sulfonyl azide, a hydrophobic probe used to photo-label acetylcholine receptor (AcChR)-rich electroplax membranes [Sator, V., Gonzalez-Ros, J. M., Calvo-Fernandez, P., & Martinez-Carrion, M. (1979) Biochemistry 18, 1200], is used to study the accessibility of the covalently attached fluorophore to extramembranous quenchers as a function of occupancy of cholinergic receptor binding sites. In these membranes, binding of water-soluble cholinergic ligands to specific sites at the extracellular side affects the fluorophore located in a distant topographical area of the AcChR molecule. When a neurotransmitter analogue (carbamylcholine) is present, the susceptibility of the covalently attached fluorophore to quenching with externally added nitromethane decreases in comparison with that of the same membranes in the absence of carbamylcholine. This neurotransmitter agonist effect is, however, reversible as removal of carbamylcholine by dialysis restores the quenching effectiveness to that of resting nonliganded membranes. The presence of bound alpha-bungarotoxin produces an opposite effect to that of carbamylcholine and induces an increase in susceptibility to quenching agent. These results are interpreted in terms of long-range effects induced by occupancy of cholinergic sites which are detected by covalently bound fluorophore located at regions of the AcChR protein accessible through the lipid matrix of the Torpedo membrane. Such effects are presumably due to molecular rearrangements within the membrane-bound AcChR structure.

A differential scanning calorimetry study of acetylcholine receptor-rich membranes from Torpedo californica.

Various acetylcholine receptor-rich membrane preparations from Torpedo californica electroplax tissue were examined using the techniques of differential scanning calorimetry coupled with gel electrophoretic analysis of heat-denaturing material and functional assays following passage through discrete transitions. In unfractionated membranes, four irreversible calorimetric transitions were observed, one of which (Td = 59 degrees C) could be assigned to a complete loss of acetylcholine receptor function. A second lower temperature transition apparently corresponds to loss of certain peripheral membrane proteins including the Mr = 43,000 polypeptide and the acetylcholinesterase activity. Membrane preparations highly enriched in acetylcholine receptor polypeptides contained a major transition at 59 degrees C which could be shown to be sensitive to the presence of added ligands of the acetylcholine receptor, supporting its assignment to structural alterations of the receptor protein or its arrangement in the membrane.

Pattern of acetylcholine receptor activity during regeneration of free muscle transplants of rats.

Acetylcholine receptor (AChR) activities were followed in regenerating muscle grafts of rats. The effects of prior nerve crush or treatment with Marcaine on alpha-Bungarotoxin binding in graft muscle extracts were compared to nontreated controls. All treatment groups displayed a similar but characteristic pattern in which minor differences in the time course and absolute levels of AChR activities were observed. These results are interpreted in terms of possible alterations in AChR levels brought about by the changes in the degree of graft innervation.

Characterization of acetylcholine receptor isolated from Torpedo californica electroplax through the use of an easily removable detergent, beta-D-octylglucopyranoside.

Non-ionic detergents used for the solubilization and purification of acetylcholine receptor from Torpedo californica electroplax may remain tightly bound to this protein. The presence of detergent greatly hinders spectrophotometric and hydrodynamic studies of the receptor protein. beta-D-Octylglucopyranoside, however, is found to be effective in solubilizing the receptor from electroplax membranes with minimal interference in the characterization of the protein. The acetylcholine receptor purified from either octylglucopyranoside- or Triton X-100-solubilized extracts exhibits identical amino acid compositions, alpha-Bungarotoxin and (+)-tubocurarine binding parameters, and subunit distributions in SDS-polyacrylamide gels. The use of octylglucopyranoside allows for the assignment of a molar absorptivity for the purified receptor at 280 nm of approx. 530000 M-1 . cm-1. Additionally, successful reconstitution of octylglucopyranoside-extracted acetylcholine receptor into functional membrane vesicles has recently been achieved (Gonzales-Ros, J.M., Paraschos, A. and Martinez-Carrion, M. (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 1796--1799). Removal of octylglucopyranoside by dialysis does not alter the specific toxin and antagonist binding ability of the receptor or its solubility at low protein concentrations. Sedimentation profiles of the purified acetylcholine receptor in sucrose density gradients reveal several components. Sedimentation coefficients obtained for the slowest sedimenting species agree with previously reported molecular weight values. Additionally, the different sedimenting forms exhibit distinctive behavior in isoelectric focusing gels. Our results suggest that both the concentration and type of detergent greatly influence the physicochemical behavior of the receptor protein.