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Query: UMLS:C0009443 (cold)
 92,137 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In this work, we investigated the role of a glycosylphosphatidylinositol (GPI)-anchored protein, the alkaline phosphatase, on the solubilization of detergent-resistant liposomes. In vivo, GPI-anchored proteins are clustered into sphingolipid- and cholesterol-rich membrane domains and this peculiar composition provides cold-detergent-insolubility. To better understand the mechanisms involved in the clustering of these subdomain components, we built a model, namely sphingolipid- and cholesterol-rich liposomes. We show the cold-Triton X-100 resistance of liposomes before and after insertion of GPI-anchored enzyme. When the amount of incorporated enzyme varied, significant changes in membrane stability occurred. Low protein contents into liposomes increased detergent insolubility, whereas high amounts decreased it. Furthermore, significant differences in the detergent-resistance of each lipid were exhibited between liposomes and proteoliposomes. Thus, the enzyme insertion led to a dramatic decrease of cholesterol solubilization, in line with the existence of cholesterol/GPI interactions. Effect of temperature on detergent resistance was also investigated. Liposome solubilization increased with temperature up to a threshold value of 40/45 degrees C. This was also the temperature at which a phase transition of liposome membrane occurred, as evidenced by Laurdan fluorescence. Although the GPI-anchored enzyme insertion modified membrane stability, no change was observed on phase transition. Our work highlights the importance of GPI-anchored proteins in the structure of sphingolipid- and cholesterol-rich membrane domains, in the detergent-insolubility of these peculiar domains, as well as in interaction of GPI proteins with cholesterol.
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PMID:Role of GPI-anchored enzyme in liposome detergent-resistance. 1257 56

In order to obtain some informations on the nature and relative activity of the phosphatases present in various helminths, biochemical studies have been made in thirteen kinds of worm parasites including the adults and larvae (Fasciola hepatica, Eurytrema pancreaticum, Paramphistomum sp., Taenia solium, Taenia pisiformis, Dipylidium caninum, Diphyllobothrium mansoni, Cysticercus cellulosae, Cysticercus fasciolaris and Sparganum). A comparison based on the analysis of pH-activity curves was made among these helminths. The worm materials were mostly obtained alive from an abattoir and removed from the organs or tissues of the animal hosts naturally infected. Sparganum and Cysticercus cellulosae, however, are collected from the subcutaneous tissue of the patients by surgical removal. The worms thoroughly washed were weighed and transferred with 0.1 M Tris buffer to a chilled glass grinder (Capacity; 15 ml) and homogenized in the cold. The homogenate was centrifuged at 5000 RPM for 30 minutes. The supernatant was pipetted off for determination of the phosphatase activity. Incubation mixtures consisted of 1 ml substrate, 1 ml buffer and 0.5ml extract. The buffers used were Tris (Hydroxymethyl) aminomethane and citric acid monohydrate and the substrate was paranitrophenyl phosphate (1 gm/25 ml). These mixtures were incubated at the temperature of 37 degrees C for 30 minutes in water bath. The absorbance or transferance of mixture was determined colorimetrically by "Spectronic 20 "spectrophotometer at 410 nm against a distilled water blank. The amount of phenol liberated was then calculated from a standard curve using phenol solutions. Controls consisted of unincubated mixtures. The results were deducted from this experiment. The phosphatase activity occurred over all parasitic helminths used in this experiment. In trematodes, pH-activity curves have demonstrated two peaks of phosphatase activity in Fasciola hepatica and Paramphistomum species. However the acid phosphatase activity was predominantly found and the alkaline phosphatase activity was found distinctly to be low in all three species. In Eurytrema pancreaticum, the pH-activity curves displayed two peaks in acid phosphatase activity, one at pH 5.0 and the other pH 9.0. In cestodes, both alkaline and acid phosphatase activity displayed the pH optima 5.0 and 9.0 to 10.0 in the adult tapeworms. However, major activity in the adults is due to the alkaline phosphtases. In contrast to the adults, Cysticercus and sparganum showed the higher activity in acid phosphatases which predominates in the larvae. In all cases of nematodes, the pH optimum for acid phosphatase was 4.0 to 6.0. A preponderance of acid phosphatase activity was shown in the extract of intestine of Ascaris lumbricoides. The aspect that phosphatases are correlated with phosphorylated passage of substances through the cuticle of helminths and may also be involved in carbohydrate metabolism is discussed.
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PMID:[Studies On Phosphatase Activity In Some Parasitic Helminths] 1291 52

Atlantic cod is a marine fish that lives at low temperatures of 0-10 degrees C and contains a cold-adapted alkaline phosphatase (AP). Preparations of AP from either the lower part of the intestines or the pyloric caeca area were subjected to proteolytic digestion, mass spectrometry and amino acid sequencing by Edman degradation. The primary structure exhibits greatest similarity to human tissue non-specific AP (80%), and approximately 30% similarity to AP from Escherichia coli. The key residues required for catalysis are conserved in the cod AP, except for the third metal binding site, where cod AP has the same variable residues as mammalian APs (His153 and His328 by E. coli AP numbering). General comparison of the amino acid composition with mammalian APs showed that cod AP contains fewer Cys, Leu, Met and Ser, but proportionally more Asn, Asp, Ile, Lys, Trp and Tyr residues. Three N-linked glycosylation sites were found. The glycan structure was determined as complex biantennary in type with fucose and sialic acid attached, although a trace of complex tri-antennary structure was also observed. A three-dimensional model was obtained by homology modelling using the human placental AP scaffold. Cod AP has fewer charged and hydrophobic residues, but more polar residues at the intersubunit surface. The N-terminal helix arm that embraces the second subunit in dimeric APs may be more flexible due to a replaced Pro at its base. One disulfide bridge was found instead of the two present in most other APs. This may invoke greater movement in the structure that together with weaker subunit contacts leads to improved catalytic efficiency.
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PMID:Amino acid sequence of the cold-active alkaline phosphatase from Atlantic cod (Gadus morhua). 1294 38

1. Cytochemical studies of the intracellular distribution of alkaline phosphatase in rat liver have been made, using a fractionation procedure recently developed in this laboratory (8) and a similar but modified method not described previously. Aqueous media were used in both cases. 2. The alkaline phosphatase was found to consist of two forms, one of which is strongly activated by magnesium and one of which is not sensitive to this metal. 3. The form of the enzyme that is not activated by magnesium occurs mainly in the nuclear fraction, where it seems to be rather firmly bound. Some of this form of the enzyme is also found in the microsomes, but very little if any occurs in the soluble supernatant fraction. 4. The form of alkaline phosphatase which is activated by magnesium occurs mainly in the soluble supernatant fraction, but what is believed are significant amounts also occur in nuclei. A significant portion of this form of the enzyme can be extracted from the isolated nuclei with cold, isotonic saline solution. Some activity of this form of the enzyme is also found in the microsomal fraction. 5. Mitochondria appear to contain relatively little alkaline phosphatase of either kind. 6. The concept of a porous nuclear membrane has been invoked to explain some of the results obtained in this work. It is postulated that part at least of the form of the enzyme that is activated by magnesium is free to diffuse back and forth through pores in the nuclear membrane, whereas this is considered not to be possible for the form of the enzyme that is insensitive to magnesium as a result of the firm binding of the latter to nuclear substance.
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PMID:Intracellular distribution of alkaline phosphatase in rat liver cells. 1324 96

THE ALDEHYDES INTRODUCED IN THIS PAPER AND THE MORE APPROPRIATE CONCENTRATIONS FOR THEIR GENERAL USE AS FIXATIVES ARE: 4 to 6.5 per cent glutaraldehyde, 4 per cent glyoxal, 12.5 per cent hydroxyadipaldehyde, 10 per cent crotonaldehyde, 5 per cent pyruvic aldehyde, 10 per cent acetaldehyde, and 5 per cent methacrolein. These were prepared as cacodylate- or phosphate-buffered solutions (0.1 to 0.2 M, pH 6.5 to 7.6) that, with the exception of glutaraldehyde, contained sucrose (0.22 to 0.55 M). After fixation of from 0.5 hour to 24 hours, the blocks were stored in cold (4 degrees C) buffer (0.1 M) plus sucrose (0.22 M). This material was used for enzyme histochemistry, for electron microscopy (both with and without a second fixation with 1 or 2 per cent osmium tetroxide) after Epon embedding, and for the combination of the two techniques. After fixation in aldehyde, membranous differentiations of the cell were not apparent and the nuclear structure differed from that commonly observed with osmium tetroxide. A postfixation in osmium tetroxide, even after long periods of storage, developed an image that-notable in the case of glutaraldehyde-was largely indistinguishable from that of tissues fixed under optimal conditions with osmium tetroxide alone. Aliesterase, acetylcholinesterase, alkaline phosphatase, acid phosphatase, 5-nucleotidase, adenosine triphosphatase, and DPNH and TPNH diaphorase activities were demonstrable histochemically after most of the fixatives. Cytochrome oxidase, succinic dehydrogenase, and glucose-6-phosphatase were retained after hydroxyaldipaldehyde and, to a lesser extent, after glyoxal fixation. The final product of the activity of several of the above-mentioned enzymes was localized in relation to the fine structure. For this purpose the double fixation procedure was used, selecting in each case the appropriate aldehyde.
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PMID:Cytochemistry and electron microscopy. The preservation of cellular ultrastructure and enzymatic activity by aldehyde fixation. 1397 66

ATPase was localized in distinct regions of the mitotic apparatus of HeLa and Sarcoma 180 tissue culture cells. ATPase was demonstrated in the metaphase spindle of HeLa and Sarcoma 180 cells fixed in cold buffered 2 per cent formalin (pH 6.5 to 6.8) containing 2 x 10(-3)M CaCl(2). A high concentration of ATPase was frequently observed at the poles of the spindle. ATPase was also demonstrated in the interzonal region of both cell types during anaphase. The narrowing of the band of ATPase activity localized in the interzonal region during telophase indicates that ATPase activity is associated with the central spindle. In polar views of Sarcoma 180 cells fixed in cold, unbuffered, 2 per cent formalin, ATPase was frequently localized in granules in the region of the inner circumference of the ring of chromosomes formed at metaphase. ATPase in the mitotic apparatus of HeLa and Sarcoma 180 cells was shown not to be due to non-specific alkaline phosphatase. Mitotic apparatus ATPase in Sarcoma 180 cells was suppressed by an -SH inhibitor.
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PMID:CYTOCHEMICAL LOCALIZATION OF ADENOSINE TRIPHOSPHATASE IN THE MITOTIC APPARATUS OF HELA AND SARCOMA 180 TISSUE CULTURE CELLS. 1422 24

Control (normothermic) and cold-acclimated (environmental temperature gradually reduced from 20 to 5 degrees C for 4 weeks) groups of male rats and hamsters were compared to elucidate the nature of angiogenesis in oxidative and glycolytic muscles of these species during progressive cold exposure. Skeletal muscle capillarity and fibre cross-sectional area were measured in the tonic soleus (SOL) and phasic tibialis anterior (TA). Cryostat sections were stained for alkaline phosphatase (ALP) activity to identify all capillaries, and proliferating cell nuclear antigen (PCNA) to localise the site of cellular proliferation. Cold-induced angiogenesis, indicated by an increase in capillary to fibre ratio (C:F), occurred in SOL of rats (approximately 20 % increase, P < 0.05) but not hamsters (approximately 9.5 % increase, n.s.), and in TA of hamsters (approximately 22 % increase, P < 0.01) but not rats (approximately 1 % increase, n.s.). The change in C:F was highest in the glycolytic cortex region of TA where fibre size is larger than in the oxidative core. Capillary-specific cell proliferation (co-localised ALP and PCNA labelling) increased in parallel with C:F. The total PCNA label density within the interstitium was some 5-fold higher than that co-localised with capillaries, but where angiogenesis occurred the relative increase in capillary labelling was 2-fold greater than for other cells of the interstitium. These data suggest a significant role for endothelial cell proliferation in the angiogenic response, indicative of the sprouting form of angiogenesis. There was a tendency for fibre hypertrophy in both SOL and TA of rats, especially in the core region of TA (P < 0.01), such that capillary density (CD) and intramuscular diffusion distances (DD) were largely unchanged following cold exposure. In contrast, fibre size was maintained in hamsters, DD reduced and CD increased compared to control TA (P < 0.01). In conclusion, cold acclimation stimulated angiogenesis in muscle of hamsters more than in rats, possibly due to a higher metabolic rate in the smaller species. Angiogenesis was also seen in SOL of rat, where oxidative capacity and muscle activity is higher than the TA. Thus, a combination of oxidative capacity, muscle activity, and fibre size may determine the degree of angiogenesis in response to low environmental temperature.
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PMID:Cold exposure differentially stimulates angiogenesis in glycolytic and oxidative muscles of rats and hamsters. 1460 72

Utilization of hepatitis C seropositive kidney donors remains controversial. We examined the use of hepatitis C seropositive donors for renal transplantation. Data for creatinine, liver function tests, cold ischemia time, and graft and patient survival were analyzed from 20 hepatitis C seropositive recipients receiving cadaveric renal allografts from seropositive donors and were compared with 20 hepatitis C seropositive recipients receiving allografts from seronegative donors. Recipients receiving a kidney from a hepatitis C seropositive donor were on the waitlist for 9.9 +/- 1.8 months, compared with 17.8 +/- 3.3 months for those receiving a kidney from a seronegative donor (p < 0.05). There were no significant differences in graft or patient survival. Incidences of acute cellular rejection and acute tubular necrosis were similar. There were no significant differences in creatinine, alanine aminotransferase, alkaline phosphatase, or bilirubin values. While there was a significant difference in aspartate aminotransferase at 2 wk and 6 months, these differences were of questionable clinical importance. In conclusion, donor seropositivity for hepatitis C should not preclude renal transplantation into a hepatitis C seropositive recipient and utilization of these organs decreases waitlist time for hepatitis C seropositive recipients.
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PMID:Use of kidneys from hepatitis C seropositive donors shortens waitlist time but does not alter one-yr outcome. 1470 26

Trabecular bone is routinely analysed by histomorphological-histometrical and immunohistochemical techniques as means of assessing the differentiation status of bone deposition and growth. Currently few embedding resins exist for which both morphological and immunohistochemical analyses can be performed on mineralised tissue. Paraffin, the standard embedding medium for bone enzyme and immunohistochemistry, can only be used on demineralised tissue, but then trabecular structure may be badly preserved. Methyl methacrylate (MMA), the resin of choice for undecalcified bone histology can only be used for bone immunohistochemistry if the usual, highly exothermic polymerisation procedure is avoided which destroys tissue antigenicity. Consequently, most current practices involve cutting samples in half to be processed in separate resins when more than one type of analysis is required. Technovit 9100 New is a low temperature MMA embedding system that is purported to significantly improve tissue antigenicity preservation allowing polymerisation at -20 degrees C. In this study, Technovit 9100 New-embedded undecalcified trabecular bone samples (adult human, young bovine and ovine) yielded immunolabelling with several bone matrix markers and preserved morphological features in 7 microm sections when stained with Masson-Goldner, von Kossa, or toluidine blue. Bone samples from all resins used were immunolabelled with antibodies against osteocalcin, alkaline phosphatase, osteopontin, osteonectin, bone sialoprotein and procollagen type I amino-terminal propeptide. Technovit-embedded bone yielded more reliable immunolabelling of the matrix proteins when compared with heat or cold-cured LR White or standard embedded MMA samples. Technovit 9100 New provided better routine histology than LR White, and was comparable to MMA. Results demonstrated that Technovit 9100 New can be used as a low-temperature acrylic resin embedding method for routine undecalcified bone histology, as well as for immunohistochemistry.
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PMID:Immunohistochemistry of matrix markers in Technovit 9100 New-embedded undecalcified bone sections. 1472 3

Alkaline phosphatases (APs) are homodimeric metalloenzymes that catalyze the hydrolysis and transphosphorylation of phosphate monoesters. Each monomer contains a metal-binding triad that for optimal activity is usually occupied by two zinc ions and one magnesium ion. The recently determined crystal structure of cold-active shrimp alkaline phosphatase (SAP) was, however, fully occupied by zinc ions. This paper describes a metal-exchange experiment in which the zinc ion in one binding site (referred to as the M3 site) is replaced by magnesium. Crystal structures revealed a concomitant structural change: the metal exchange causes movement of a ligating histidine into a conformation in which it does not coordinate to the metal ion. The M3 site is relevant to catalysis: its occupation by magnesium is postulated to favour catalysis and it has been suggested to be a regulatory site for other APs. Further crystallographic studies show that ligand binding can induce a conformational change of an active-site arginine from a 'non-docked' (non-interacting) to a 'docked' conformation (interacting with the ligand). The first conformation has only been observed in SAP, while the latter is common in available AP structures. The observation that the arginine does not always bind the substrate may explain the increased catalytic efficiency that is generally observed for cold-active enzymes.
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PMID:Ligand-binding and metal-exchange crystallographic studies on shrimp alkaline phosphatase. 1533 25


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