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Purification and characterization of deep sea shark Centrophorus squamosus liver oil 1-O-alkylglycerol ether lipids.

The 1-O-alkylglycerol composition of the liver oil of the deep sea shark Centrophorus squamosus, a species which provides edible flesh, has been determined. After various fractionations of the oil, the unsaponifiable fraction was characterized by means of gas chromatography/mass spectrometry, electron impact, and positive-ion chemical ionization. The oil is composed of 60% unsaponifiable matter, containing 45% squalene, 4.5% cholesterol, and 10% of linear saturated and monounsaturated glycerol ethers with 14-18 carbon atoms. After a first separation by chromatography on silicic acid, monounsaturated glycerol ethers have been separated from the saturated homologues, in particular from 1-O-octadecylglycerol (batyl alcohol) and 1-O-hexadecylglycerol (chimyl alcohol), via urea complexation. This newer application of the urea method, already used in the past to extract saturated from polyunsaturated fatty acids, allowed the purification of the main components of the complex unsaturated glycerol ether fraction, namely, 1-O-octadecen-9'ylglycerol (selachyl alcohol) and 1-O-hexadecen-9'ylglycerol.

Animals↗

Structural analysis, selection, and ontogeny of the shark new antigen receptor (IgNAR): identification of a new locus preferentially expressed in early development.

The new antigen receptor (IgNAR) family has been detected in all elasmobranch species so far studied and has several intriguing structural and functional features. IgNAR protein, found in both transmembrane and secretory forms, is a dimer of heavy chains with no associated light chains, with each chain of the dimer having a single free and flexible V region. Four rearrangement events (among 1V, 3D, and 1J germline genes) generate an expressed NAR V gene, resulting in long and diverse CDR3 regions that contain cysteine residues. IgNAR mutation frequency is very high and "selected" mutations are found only in genes encoding the secreted form, suggesting that the primary repertoire is entirely CDR3-based. Here we further analyzed the two IgNAR types, "type 1" having one cysteine in CDR3 and "type 2" with an even number (two or four) of CDR3 cysteines, and discovered that placement of the disulfide bridges in the IgNAR V domain differentially influences the selection of mutations in CDR1 and CDR2. Ontogenetic analyses showed that IgNAR sequences from young animals were infrequently mutated, consistent with the paradigm that the shark immune system must become mature before high levels of mutation accompanied with selection can occur. Nevertheless, also in agreement with the idea that the IgNAR repertoire is entirely CDR3-based, but unlike studies in most other vertebrates, N-region diversity is present in expressed IgNAR clones at birth. During the investigation of this early IgNAR repertoire we serendipitously detected a third type of IgNAR gene that is expressed in all neonatal tissues; later in life its expression is perpetuated only in the epigonal organ, a tissue recently shown to be a (the?) primary lymphoid tissue in elasmobranchs. This "type 3" IgNAR gene still undergoes three rearrangement events (two D regions are "germline-joined"), yet CDR3 sequences were exactly of the same length and very similar sequence, suggesting that "type 3" CDR3s are selected early in ontogeny, perhaps by a self-ligand.

Amino Acid Sequence↗

Haemoglobin function and respiratory status of the Port Jackson shark, Heterodontus portusjacksoni, in response to lowered salinity.

Haemoglobin function and respiratory status of sub-adult sharks, Heterodontus portusjacksoni was investigated for up to 1 week following transfer from 100% to either 75% or 50% seawater. Metabolic rates were unusually low and arterial-venous differences in blood O2 small. Haemodilution from osmotic inflow lowered haematocrit and reduced blood O2 content by up to 50%. There was no change in O2 consumption rate, blood O2 partial pressure, cardiac output, or the arterial-venous O2 content difference, and thus O2 delivery was maintained. Ventilation was acutely elevated but returned to normal within 24 h. The O2 delivery to the tissues was facilitated by decreased blood O2-affinity that could not be simply ascribed to changes in the osmolyte concentration. The Hb was unaffected by changes in intra-erythrocyte fluid urea or trimethylamine-N-oxide (TMAO) but was sensitive to changes in NaCl. The Bohr shifts in whole blood were low and there was little role for pH in modulating O2 transport. Venous Hb saturation remained close to 65%, at the steepest part of the in vivo O2 equilibrium curve, such that O2 unloading could be facilitated by small reductions in pressure without increasing cardiac or ventilatory work. H. portusjacksoni tolerated 50% seawater for at least 1 month, but there was little evidence of respiratory responses being adaptive which instead appeared to be consequential on changes in osmotic and ionic status.

Acclimatization↗

Dangerous nutrition? Calcium, vitamin D, and shark cartilage nutritional supplements and cancer-related hypercalcemia.

The use of nutritional supplements in the general population and in cancer patients has become very popular. These supplements are not perceived as medications and are presumed to be safe by cancer patients, who may however be at risk for hypercalcemia. We note that many of our patients who have developed symptomatic hypercalcemia were taking vitamin D, calcium, or shark cartilage supplements. We report eight cases of hypercalcemia in cancer patients seen at the Cleveland Clinic Foundation in whom these nutritional supplements may have contributed to the prevalence or severity of hypercalcemia.

Aged↗

Redescription of Cetorhinicola acanthocapax Beveridge & Campbell, 1988 (Cestoda: Trypanorhyncha) from the basking shark Cetorhinus maximus (Gunnerus).

Cetorhinicola acanthocapax Beveridge & Campbell, 1988 is redescribed based on adult specimens collected from a basking shark Cetorhinus maximus (Gunnerus) off the east coast of New Zealand. Both the mature and gravid segments are described for the first time, and the first scanning electron micrographs of the armature and scolex microtriches are provided. Novel features include the opening of the vagina anterior to the cirrus-sac and the extension of the vagina anterior to the cirrus-sac. Pectinate microtriches are present on the anterior quarter of the pars pedunculus scolecis scattered between filiform microtriches; the bothrial margins are covered with tridigitate microtriches, while the adherent surface of the bothria have pectinate microtriches. The additional morphological features described are consistent with a close association between Cetorhinicola Beveridge & Campbell, 1988 and the Eutetrarhynchoidea Guiart, 1927.

Animals↗

Different regional specializations of neurons in the ganglion cell layer and inner plexiform layer of the California horned shark, Heterodontus francisci.

We have described a population of neurons in the retinal of a shark, Heterodontus francisci, which is precisely aligned within the inner plexiform layer (IPL) and which differs from neurons in the ganglion cell layer (GCL) in soma size and topographical distribution. GCL neurons are relatively small and form a horizontally oriented visual streak; IPL neurons are significantly larger and form a circular specialization in the far temporal retina. Thus, it appears that there are two distinct retinal specializations in Heterodontus: one subserving frontal vision and one which provides a panoramic view of the lateral visual field.

Animals↗

An autoradiographic study of the retinofugal projections in the shark, Hemiscyllium plagiosum.

The retinofugal projections in the shark, Hemiscyllium plagiosum were studied after unilateral eye injection of tritiated proline. The results showed that each eye in this animal projects not only to the contralateral side of the brain but also to several ipsilateral visual centers including the nucleus suprachiasmaticus, ventrolateral nucleus of the optic tract and the optic tectum. These results suggest that some of the visual centers in this animal receive direct retinal inputs from both eyes as is the case in many other animals and most notably in mammals.

Animals↗

The nervus terminalis of the shark: the effect of efferent impulses on ganglion cell activity.

Tonic activity was recorded extracellularly in the nervus terminalis of the bonnethead shark. Cutting or cooling the nerve central to its ganglion eliminated these spikes, and was followed by an increase in multi-unit activity which appeared to arise from the ganglion. After cutting centrally, electrical stimulation of the distal stump of the nerve suppressed this multi-unit activity. The data suggest that efferent impulses may normally suppress ganglion cell activity.

Action Potentials↗

Comparison of a carboxypeptidase E-like enzyme in human, bovine, mouse, Xenopus, shark and Aplysia neural tissue.

Several diverse species contain an enzyme with many properties in common with those of bovine carboxypeptidase E (CPE), a neuropeptide processing carboxypeptidase B-like enzyme. This enzyme has been designated EC 3.4.17.10, and is also known as enkephalin convertase and carboxypeptidase H. All tissues that are known to contain bioactive peptides also contain CPE-like enzymatic activity. In Xenopus laevis, enzyme activity is highest in the brain and pituitary, lower in the skin, and undetectable in liver and gut. In Aplysia californica, enzyme activity is highest in the atrial gland, but is also present in moderate amounts in the various neural tissue. CPE extracted from human, bovine, mouse, Xenopus, shark, and Aplysia neural tissue is substantially purified using substrate affinity chromatography and concanavalin A sepharose columns. The partially purified enzyme from all species examined possess very similar enzymatic properties. These properties include a pH optimum of 5.6, a stimulation by cobalt chloride, and an inhibition by chelating agents (1,10-phenanthroline). Arginine-derived active site-directed inhibitors show similar inhibition constants (Ki's) towards enzyme from the various species, whereas lysine-derived inhibitors are substantially less potent towards the Aplysia carboxypeptidase than towards enzyme isolated from the other species. The similar properties of the carboxypeptidase isolated from the various species suggests that a CPE-like is involved in the biosynthesis of many peptide neurotransmitters and hormones in a wide range of organisms.

Animals↗

A study on heterogeneity in molecular species of shark cartilage chondroitin sulfate C. Fractionation of the polysaccharide on Sepharose CL-4B in the presence of high concentrations of ammonium sulfate.

Shark cartilage chondroitin sulfate C was fractionated by chromatography on Sepharose CL-4B-2.5 to 1.5M ammonium sulfate in 10mM hydrochloric acid at 4 degrees. Both unit-disaccharide composition and molecular-size distribution clearly affected the fractionation. Comparison of this fractionation with the fractionation on Sepharose 6B gel in 0.2M sodium chloride revealed that the former is distinctly superior to the latter. The fractionation on Sepharose CL-4B in the presence of ammonium sulfate also showed that the chondroitin sulfate C molecules having a larger molecular size contain generally more chondroitin 6-sulfate units (as major constituent) and less chondroitin disulfate units (D type, as minor constituent) than those having a smaller molecular size).

Ammonium Sulfate↗

Substitution of arginine for lysine 134 alters electrostatic parameters of the active site in shark Cu,Zn superoxide dismutase.

The complete amino acid sequence was determined for the Cu,Zn superoxide dismutase from the shark Prionace glauca. The active site region shows the substitution of an Arg for Lys at position 134, which is important for electrostatic facilitation of the diffusion of O2- to the catalytically active copper. This change may be related to observed alterations of electrostatic parameters of the enzyme (pK of the pH dependence of the enzyme activity, rate of inactivation by H2O2), although it preserves a high efficiency of dismutation at neutral pH.

Amino Acid Sequence↗

Plasma levels of glucose, alanine, lactate, and beta-hydroxybutyrate in the unfed spiny dogfish shark (Squalus acanthias) after surgery and following mammalian insulin infusion.

Serial blood samples were collected from the unfed spiny dogfish shark (Squalus acanthias) by use of a cannula chronically implanted in the dorsal aorta. Plasma glucose, alanine, lactate, and beta-hydroxybutyrate levels were estimated by standard enzymatic procedures. The metabolite levels were estimated immediately after the surgery, for an additional 3 to 5 postoperative days before experimental use, and for 7 to 9 days after a single infusion of mammalian insulin (50, 100, and 250 IU/kg body wt) or the control solution. Plasma glucose, alanine, and lactate levels declined for 1 or 2 days after the surgery, but beta-hydroxybutyrate levels were essentially unchanged. Plasma glucose levels remained relatively stable for the remainder of the pretreatment period, and in the control animals during the subsequent treatment period. In contrast, plasma alanine and beta-hydroxybutyrate levels increased during both the pretreatment period and the treatment period in the control animals. Plasma lactate levels increased in the control animals after Day 3 of the treatment period. Insulin infusions resulted in severe, prolonged depressions of plasma glucose and alanine levels. Plasma lactate levels also fell after insulin infusion, but the depressed levels were interrupted by precipitous, large increases, followed by sharp declines, that were evident for one to three sampling periods and whose onset varied between Days 3 and 5 in individual animals. The levels increased again in some animals between Days 6 and 9. Plasma beta-hydroxybutyrate levels were comparatively resistant to insulin. After an early decline, plasma beta-hydroxybutyrate rose to levels that were near the control levels by Days 2 through 4, but fell to near the initial levels by Day 5 and thereafter. The results suggest that the spiny dogfish, a species that consumes reliable and abundant amounts of protein and lipid, but little carbohydrate, and that may feed as seldom as every 2 weeks, uses ketone bodies (beta-hydroxybutyrate and presumably acetoacetate) as primary fuels. Glucose is apparently synthesized by gluconeogenesis, alanine and lactate are probable substrates, and ketone bodies are the likely sources of energy. The evidence supports the hypothesis that glucose is synthesized to maintain muscle glycogen reserves that are used for anaerobic glycolysis when the animal is engaged in predation, escape, or other circumstances that dictate vigorous swimming. If other tissues and organs require glucose as a fuel source, the amounts must be exceptionally small.

3-Hydroxybutyric Acid↗

Cell types in peripheral blood of the nurse shark: an approach to structure and function.

Ultrastructural and functional studies were carried out on nurse shark (Ginglymostoma cirratum) peripheral blood cells in order ot identify cells of definitive morphology and specific function. Along with erythrocytes and thrombocytes, four morphologically distinct leucocytes are recognized in peripheral blood: two types of granulocytes, the 'eosiniphil' and the 'granulocyte', and two mononuclear agranulocytic cells, one resembling mammalian macrophage and monocyte, the other resembling mammalian lymphocyte. Also present in peripheral circulation are blast-like cells and mitotic cells. In vitro phagocytosis was demonstrated by the monocyte-macrophage and the granulocyte while thrombocytes, eosinophils and lymphocytes showed no phagocytic activity in the system studied. It is stressed that care must be used in drawing functional analogies between blood cells of a mammal and an elasmobranch on the basis of morphological similarity alone.

Animals↗

Ultrastructure of the pre-implantation shark yolk sac placenta.

During ontogeny, the yolk sac of viviparous sharks differentiates into a yolk sac placenta which functions in gas exchange and hematrophic nutrient transport. The pre-implantation yolk sac functions in respiration and yolk absorption. In a 10.0 cm embryo, the yolk sac consists of six layers, viz. (1) somatic ectoderm; (2) somatic mesoderm; (3) extraembryonic coelom; (4) capillaries; (5) endoderm; and (6) yolk syncytium. The epithelial ectoderm is a simple cuboidal epithelium possessing the normal complement of cytoplasmic organelles. The endoplasmic cisternae are dilated and vesicular. The epithelium rests upon a basal lamina below which is a collagenous stroma that contains dense bodies of varying diameter. They have a dense marginal zone, a less dense core, and a dense center. The squamous mesoderm has many pinocytotic caveolae. The capillary endothelium is adjacent to the mesoderm and is delimited by a basal lamina. The endoderm contains yolk degradation vesicles whose contents range from pale to dense. The yolk syncytium contains many morphologically diverse yolk granules in all phases of degradation. Concentric membrane lamellae form around yolk bodies as the main yolk granules begin to be degraded. During degradation, yolk platelets exhibit a vesicular configuration.

Animals↗

Poisonings from flesh of the Greenland shark Somniosus microcephalus may be due to trimethylamine.

The flesh of the Greenland shark, Somniosus microcephalus, especially in fresh condition, is toxic to both dog and man. Analysis of the toxic fractions indicates the presence of large amounts of trimethylamine oxide (TMAO) but no other substances that could be directly responsible for the poisoning. The symptoms appear to be due to an acute trimethylamine (TMA) poisoning arising from intestinal reduction of TMAO to TMA. TMA stimulates contraction of the guinea-pig ileum (antagonized by atropine but not hexamethonium). Low concentrations of TMA increase contraction of the rat phrenic nerve-diaphragm preparation, whereas higher concentrations cause blockade.

Animals↗

The second component (C2n) of the nurse shark complement system: purification, physico-chemical characterization and functional comparison with guinea pig C4.

C2n was isolated and purified from nurse shark serum by initial cold low ionic strength precipitation, followed by chromatography on DE-52 cellulose, hydroxylapatite and filtration on BioGel A-0.5. The molecular weight, determined by gel filtration and PAGE, was 170K daltons and 180K daltons respectively. The molecule retained 100% activity after heating for 2 hours at 56 degrees C. Optimal temperature and time for C2n fixation was 30 degrees C and 20 minutes respectively. Experiments were carried out to determine whether C2n and C4gp could replace each other in the lysis of cellular intermediate EAnC1n (with which both components are compatible) when the remaining homologous and heterologous complement components were supplied. Results showed that lysis only occurred with the homologous components indicating an incompatibility between C2n and C2gp and C4gp and C3n. Results from interference studies indicated that C4gp and C2n interfere with each others binding and/or interaction with EAnC1n.

Animals↗

The existence of glutathione and cysteine disulfide-linked to erythrocyte carbonic anhydrase from tiger shark.

In this study it is shown that the higher molecular weight previously reported for tiger shark carbonic anhydrase (carbonate hydro-lyase, EC 4.2.1.1) compared to other carbonic anhydrases is decreased to a normal value around 30 000 after disulfide reduction of the enzyme. This difference in molecular weight is at least partly due to the existence of disulfide-linked glutathione and cysteine residues. Approx. 3 mol glutathione and a similar amount of cysteine are shown to be bound per mol enzyme. The presence of these factors also has effects on the enzyme activity.

Amino Acids↗

K+-induced swelling of the dogfish shark (Squalus acanthias) rectal gland cells is associated with changes of the cytoskeleton.

Dogfish shark (Squalus acanthias) rectal gland cells swell massively when incubated in elasmobranch media in which Na+ was equivalently replaced by K+; this swelling was abolished when the impermeant gluconate replaced Cl-, while the cell depolarization was comparable in both media. The K+-effect was associated with (a) an increase of the steady-state 42K (and 86Rb) efflux (particularly of the rate constant of the fast cellular efflux component) and a rearrangement of the respective cellular pools of K+; (b) an alteration of cell morphology and the pattern of the F-actin staining along the basolateral cell membrane as revealed with fluorescent analogs of phallacidin. These changes were independent of cell volume, being identical in KCl and K-gluconate media. The observations were specific for K+ (and Rb+): replacement of media Na+ by Li+ (which is not actively extruded by the cells), or the presence of ouabain, produced only minor swelling without affecting cell morphology and F-acting distribution. The results are consistent with the following view: as opposed to Na+ or Li+ media, the K+-induced changes of the cortical F-actin component of the cytoskeleton permit the observed massive cell swelling due to the osmotic contribution of intracellular impermeant anion(s).

Actins↗