Human placental sialidase complex: characterization of the 60 kDa protein that cross-reacts with anti-saposin antibodies

Sialidase isolated from human placenta is associated with several proteins including acid beta-galactosidase, carboxypeptidase, N-acetyl-alpha-galactosaminidase, and others. These proteins are thought to form an aggregated complex during isolation of sialidase. One of the proteins of 60 kDa was recently identified by Potier et al. (Biochem. Biophys. Res. Comm. 173, 449-456, 1990) as a sialidase protein: this protein also cross-reacted with anti-prosaposin antibodies. We have isolated this protein and from the following evidence identified it as a heavy chain component of immunoglobulin G and not sialidase or a derivative of prosaposin. On gel filtration HPLC, sialidase activity and the 60 kDa protein were clearly separated from one another. The 60 kDa protein cross-reacted not only with antibodies raised against human saposins A, C, and D, but also with second antibody (goat anti-rabbit immunoglobulin G antibody) alone. This 60 kDa protein strongly cross-reacted with anti-human immunoglobulin G antibodies. The sequence of the initial 15 amino acids from the N-terminus of the 60 kDa protein was identical to the sequence of an immunoglobulin G heavy chain protein Tie (gamma 1).     

The peripheral lymph node homing receptor, LECAM-1, is involved in CD18-independent adhesion of human neutrophils to endothelium.

The binding of polymorphonuclear granulocytes (PMN) to activated vascular endothelium is a crucial step in the recruitment of PMN to an inflammatory site. Studies employing cytokine-activated endothelium in culture have shown that PMN binding involves the CD18 family of leukocyte integrins, but also CD18-independent adhesion mechanism(s) on PMN that have not been defined. We unify here two previously disparate approaches to study cell adhesion events between endothelial cells and leukocytes. We show that antibodies to human LECAM-1, the peripheral lymph node homing receptor that is also expressed on PMN, partially inhibit the adhesion of human PMN not only to HEV in frozen sections of lymph node tissue, but also to cytokine-activated human umbilical vein endothelium in vitro. Inhibition with anti-LECAM-1 antibodies and anti-CD18 antibodies is additive. Furthermore, the anti-LECAM-1 antibodies inhibit the adhesion of CD18-deficient PMN to cytokine activated human endothelial cells. These findings indicate that LECAM-1 and CD18-mediated binding mechanisms are independent, and act coordinately or sequentially to mediate PMN attachment to cytokine activated endothelium.     

Cellular localization of thiol-proteinase inhibitor in the epidermis of the newborn rat

Summary In cichlid, poecilid and centrarchid fishes luteinizing hormone releasing hormone (LHRH)-immunoreactive neurons are found in a cell group (nucleus olfactoretinalis) located at the transition between the ventral telencephalon and olfactory bulb. Processes of these neurons project to the contralateral retina, traveling along the border between the internal plexiform and internal nuclear layer, and probably terminating on amacrine or bipolar cells. Horseradish peroxidase (HRP) injected into the eye or optic nerve is transported retrogradely in the optic nerve to the contralateral nucleus olfactoretinalis where neuronal perikarya are labeled. Labeled processes leave this nucleus in a rostral direction and terminate in the olfactory bulb. The nucleus olfactoretinalis is present only in fishes, such as cichlids, poecilids and centrarchids, in which the olfactory bulbs border directly the telencephalic hemispheres. In cyprinid, silurid and notopterid fishes, in which the olfactory bulbs lie beneath the olfactory epithelium and are connected to the telencephalon via olfactory stalks, the nucleus olfactoretinalis or a comparable arrangement of LHRH-immunoreactive neurons is lacking. After retrograde transport of HRP in the optic nerve of these fishes no labeling of neurons in the telencephalon occurred. It is proposed that the nucleus olfactoretinalis anatomically and functionally interconnects and integrates parts of the olfactory and optic systems.     

Generality of the action of various maturation-promoting factors

It has been known in amphibians and starfishes that a cytoplasmic factor called maturation-promoting factor (MPF), produced in maturing oocytes under the influence of the maturation-inducing hormones, can induce germinal vesicle breakdown (GVBD) and the subsequent process of meiotic maturation. The present study revealed that injection of cytoplasm of maturing starfish oocytes (starfish MPF) into immature sea cucumber oocytes brought about maturation of the recipients. Amphibian MPF obtained from mature oocytes of Xenopus laevis or Bufo bufo was found to induce maturation of starfish oocytes following injection. Cytoplasm taken from cleaving starfish blastomeres induced maturation when injected into immature starfish oocytes. The maturation-inducing activity of cytoplasm of starfish blastomeres changed along with the mitotic cell cycle during 1- to 4-cell stages so far tested and reached a peak just before cleaving. Furthermore, an extract of mammalian cultured cells, CHO or V-79, synchronized in M phase, induced GVBD in starfish oocytes following injection, whereas S phase extract had little activity. These facts suggest that MPF generally brings about nuclear membrane breakdown in both meiosis and mitosis, and that the nature of MPF is very similar among vertebrates and invertebrates.     

Unsaturated diacylglycerol as a possible messenger for the activation of calcium-activated, phospholipid-dependent protein kinase system

A small quantity of unsaturated diacylglycerol (DG) sharply decreased the Ca²⁺ and phospholipid concentrations needed for full activation of a Ca²⁺-activated, phospholipid-dependent multifunctional protein kinase described earlier (Takai, Y., Kishimoto, A., Iwasa, Y., Kawahara, Y., Mori, T. and Nishizuka, Y. (1979)$\text{J.}\text{Biol.}\text{Chem.}\text{254}$. 3692–3695). In the presence of unsaturated DG and micromolar order of Ca²⁺, phosphatidylserine (PS) was most relevant with the capacity to activate the enzyme, whereas phosphatidylethanolamine and phosphatidylinositol (PI) were far less effective. Phosphatidylcholine was practically inactive. It is possible, therefore, that unsaturated DG, which may be derived from PI turnover provoked by various extracellular stimulators, acts as a messenger for activating the enzyme, and that Ca²⁺ and various phospholipids such as PI and PS seem to play a role cooperatively in this unique receptor mechanism.     

A role of membranes in the activation of a new multifunctional protein kinase system.

A new multifunctional protein kinase, which normally exists as an inactive form in the soluble fraction in mammalian tissues, attaches to membranes to exhibit full enzymatic activity. A low concentration of Ca2+ is absolutely necessary for this activation. This process is reversible. cAMP shows no effect. The active factors in membranes are phosphatidylinositol, phosphatidylserine, phosphatidic acid, diphosphatidylglycerol, and phosphatidylethanolamine in that order. Phosphatidylcholine and sphingomyelin are far less effective. Cytoplasmic as well as other membrane fractions from various tissues are active in supporting the enzymatic activity. A possible role of this Ca2+ and phospholipid-activated protein kinase system in transmembrane control is proposed.     

Ceramide metabolism in brain

Summary Ceramide is the fundamental structure and key intermediate of all sphingolipids. Biosynthesis and catabolism of brain ceramide, especially their relationship to the metabolism of more complex sphingolipids in brain, are reviewed. Human genetic diseases which involve altered ceramide metabolism are also discussed.     

Fatty acid α-hydroxylation and its relation to myelination

Summary -Hydroxylation is an enzymatic reaction by which long-chain fatty acids are converted to their -hydroxy derivatives. This reaction, in animals, can be detected only in developing brain and is the rate-determining step in the synthesis of hydroxycerebroside, which is an indispensable and abundant myelin lipid. In addition to a particulate fraction from brain, two cytoplasmic factors, one heat-stable and the other heat-labile, are required for -hydroxylation. During the past eight years we have been investigating -hydroxylation. Our progress is summarized and discussed here.