Polyanionic Polysaccharide Research Articles

WIDESPREAD OCCURRENCE OF THE OCEANIC ULTRAPLANKTER, PRASINOCOCCUS CAPSULATUS (PRASINOPHYCEAE), THE DIAGNOSTIC “GOLGI‐DECAPORE COMPLEX” AND THE NEWLY DESCRIBED POLYSACCHARIDE “CAPSULAN”

A nonmotile green nanoalga was isolated from the waters over the Cayman Trench in March 1979 and has been maintained in culture as clone URI 266G (CCMP 1202). It was observed to form a copious polysaccharide capsule that presumably originated in the Golgi body and was secreted through a crown of 10 pores in the cell wall, the “decapore.” This multilaminate apical area, lying adjacent to the Golgi, underwent structural changes during morphogenesis. The polysaccharide precursors that coalesced to form the capsule apparently became stainable and visible as they exited the decapore when they cross‐linked with divalent ions in seawater. Cell wall precursors, or a cell wall lamina, surrounded the daughter cells both during synchronous binary fission and after cell separation, with the maternal capsule perhaps acting as a template. Similar prasinophyte isolates have been obtained from widespread areas of the North Atlantic and were divided into two subgroups on the basis of their pigment complement (Hooks et al. 1988). One subgroup, typified by clone Ω 48‐23 (CCMP 1203), was described by Guillard et al. (1991) as Pycnococcus provasolii Guillard within a new family, the Pycnococcaceae. The other subgroup, typified by clone URI 266G (CCMP 1202), contained two unique carotenoids, one of which was uriolide (Foss et al. 1986). Subsequently, Miyashita et al. (1993) described an alga from the western Pacific Ocean that is indistinguishable from URI 266G in both pigment composition and ultrastructure that they named Prasinococcus capsulatus Miyashita et Chihara and placed tentatively in the Pycnococcaceae. They described a curious asexual budding fission. Here we suggest an alternative form of cell division analogous to that observed in the other described Pycnococcaceae. We used theultrastructure of cells in exponential and stationary phases of growth to illustrate synchronous asexual binary fission, the “Golgi‐decapore complex,” and its apparent role in capsule formation. A unique sulfated and carboxylated polyanionic polysaccharide named capsulan is released from this complex.

The plasma membrane of Leishmania donovani promastigotes is the main target for CA(1-8)M(1-18), a synthetic cecropin A-melittin hybrid peptide.

Reports on the lethal activity of animal antibiotic peptides have largely focused on bacterial rather than eukaryotic targets. In these, involvement of internal organelles as well as mechanisms different from those of prokaryotic cells have been described. CA(1-8)M(1-18) is a synthetic cecropin A-melittin hybrid peptide with leishmanicidal activity. Using Leishmania donovani promastigotes as a model system we have studied the mechanism of action of CA(1-8)M(1-18), its two parental peptides and two analogues. At micromolar concentration CA(1-8)M(1-18) induces a fast permeability to H+/OH-, collapse of membrane potential and morphological damage to the plasma membrane. Effects on other organelles are related to the loss of internal homeostasis of the parasite rather than to a direct effect of the peptide. Despite the fast kinetics of the process, the parasite is able to deactivate in part the effect of the peptide, as shown by the higher activity of the d-enantiomer of CA(1-8)M(1-18). Electrostatic interaction between the peptide and the promastigote membrane, the first event in the lethal sequence, is inhibited by polyanionic polysaccharides, including its own lipophosphoglycan. Thus, in common with bacteria, the action of CA(1-8)M(1-18) on Leishmania promastigotes has the same plasma membrane as target, but is unique in that different peptides show patterns of activity that resemble those observed on eukaryotic cells.

Association of myeloperoxidase with heparin: oxidative inactivation of proteins on the surface of endothelial cells by the bound enzyme.

Chromatography of human myeloperoxidase (MPO) on a heparin-agarose column demonstrated a tight association of the protein with the resin. The electrophoretic mobility of mixtures of MPO and heparin in polyacrylamide gels under nondenaturing conditions was consistent with a strong interaction of the cationic enzyme with the polyanionic polysaccharide. Purified MPO prebound to bovine aorta endothelial cells (BAEC) and supplemented with hydrogen peroxide dose- and time-dependently abrogated the interaction of coagulation factor IX (FIX) with factor IX-binding protein (FIXBP) on the surface of BAEC reflecting oxidative modification of the binding protein. This inactivation of FIXBP required the presence of chloride implicating hypochlorite in the reaction. Hypochlorite and activated neutrophils exerted a similar effect. The oxidative modification of FIXBP was only partially dependent on the addition of hydrogen peroxide and was abolished by exogenous heparin which displaced MPO from the cell surface, emphasizing the functional differences between cell-bound and free enzyme.

The influence of physical structure and charge on neurite extension in a 3D hydrogel scaffold

Understanding neural cell differentiation and neurite extension in three-dimensional scaffolds is critical for neural tissue engineering. This study explores the structure-function relationship between a 3D hydrogel scaffold and neural cell process extension and examines the role of ambient charge on neurite extension in 3D scaffolds. A range of agarose hydrogel concentrations was used to generate varied gel physical structures and the corresponding neurite extension was examined. Agarose gel concentration and the corresponding pore radius are important physical properties that influence neural cell function. The average pore radii of the gels were determined while the gel was in the hydrated state and in two different dehydrated states. As the gel concentration was increased, the average pore radius decreased exponentially. Similarly, the length of neurites extended by E9 chick DRGs cultured in agarose gels depends on gel concentration. The polycationic polysaccharide chitosan and the polyanionic polysaccharide alginate were used to incorporate charge into the 3D hydrogel scaffold, and neural cell response to charge was studied. Chitosan and alginate were covalently bound to the agarose hydrogel backbone using the bi-functional coupling agent 1,1'carbonyldiimldazole. DRGs cultured in chitosan-coupled agarose gel exhibited a significant increase in neurite length compared to the unmodified agarose control. Conversely, the alginate-coupled agarose gels significantly inhibited neurite extension. This study demonstrates a strong, correlation between the ability of sensory ganglia to extend neurites in 3D gels and the hydrogel pore radius. In addition, our results demonstrate that charged biopolymers influence neurite extension in a polarity dependent manner.

AM218, a new polyanionic polysaccharide, induces radioprotection in mice when administered shortly before irradiation

We have shown that the polyanionic polysaccharide AM218 improves the survival rate of the potentially lethally irradiated mice. This radioprotective effect was highly dependent on the administration schedule, the most efficient protocol being that in which the drug was given shortly before irradiation. The haematopoietic implications in the pharmacological action of AM218 were confirmed by the improved recovery in the three peripheral blood lineages observed in the AM218-treated mice. However, because of a marked increase observed in the number of white blood cells during the period of highest mortality of the control irradiated mice, effects on the neutrophil lineage may account for the effects mediated by AM218 in the irradiated mice. Both in vitro and in vivo treatment with AM218 before irradiation improved the survival rate of CFU-GM progenitors, while no effects were observed on the CFU-S pool. This led us to postulate that the improved survival rate of the committed progenitors, at least the granulocyte-macrophage progenitors, accounts for the radioprotective capacity of AM218.

A detailed 1H and 13C NMR study of a repeating disaccharide of hyaluronan: the effect of sodium and calcium ions

Hyaluronan (HA), a polyanionic polysaccharide consisting of repeating glucuronate and N-acetylglucosamine residues, exists surrounded by ions in its physiological milieu. For example, the average concentration of sodium is 300 mM in bovine hyaline cartilage, or roughly twice that of typical extracellular fluid [1]. It has been shown that salts can modify strongly the properties of HA [2,3] and its constituent monomers [4,5]. For example, Van Damme et al. [2] reported that binding of HA to lysozyme is most efficient at pH 7.5 and 10-15 mM NaCI. Self-association of hyaluronate segments in aqueous solution requires a sodium concentration of 150 mM [3]. Several physical techniques (NMR [6-13], X-ray [14-17], CD [8,18], and IR [9,14]) have been employed to determine if salts interact with uronate residues non-specifically (via electrostatic interaction) [7] or through chelation by several ligands acting in concert. Most authors have postulated the existence of chelation sites. A review of studies conducted on HA in the solid and liquid states leads to the conclusion that the oxygens of the carboxyl groups on HA and of water molecules always take part in coordination of metal cations, while other oxygens around the sugar ring are sometimes involved as well. The type of

Uptake and transformation of metals and metalloids by microbial mats and their use in bioremediation

Constructed microbial mats, used for studies on the removal and transformation of metals and metalloids, are made by combining cyanobacteria inoculum with a sediment inoculum from a metal-contaminated site. These mats are a heterotrophic and autotrophic community dominated by cyanobacteria and held together by slimy secretions produced by various microbial groups. When contaminated water containing high concentrations of metals is passed over microbial mats immobilized on glass wool, there is rapid removal of the metals from the water. The mats are tolerant of high concentrations of toxic metals and metalloids, such as cadmium, lead, chromium, selenium and arsenic (up to 350 mg L−1). This tolerance may be due to a number of mechanisms at the molecular, cellular and community levels. Management of toxic metals by the mats is related to deposition of metal compounds outside the cell surfaces as well as chemical modification of the aqueous environment surrounding the mats. The location of metal deposition is determined by factors such as redox gradients, cell surface micro-environments and secretion of extra-cellular bioflocculents. Metal-binding flocculents (polyanionic polysaccharides) are produced in large quantities by the cyanobacterial component of the mat. Steep gradients of redox and oxygen exist from the surface through the laminated strata of microbes. These are produced by photosynthetic oxygen production at the surface and heterotrophic consumption in the deeper regions. Additionally, sulfur-reducing bacteria colonize the lower strata, removing and utilizing the reducing H2S, rather than water, for photosynthesis. Thus, depending on the chemical character of the microzone of the mat, the sequestered metals or metalloids can be oxidized, reduced and precipitated as sulfides or oxides. For example precipitates of red amorphous elemental selenium were identified in mats exposed to selenate (Se-VI) and insoluble precipitates of manganese, chromium, cadmium, cobalt, and lead were found in mats exposed to soluble salts of these metals. Constructed microbial mats offer several advantages for use in the bioremediation of metal-contaminated sites. These include low cost, durability, ability to function in both fresh and salt water, tolerance to high concentrations of metals and metalloids and the unique capacity of mats to form associations with new microbial species. Thus one or several desired microbial species might be integrated into mats in order to design the community for specific bioremediation applications.

Divalent cation-dependent interaction of sulfated polysaccharides with phosphatidylcholine and mixed phosphatidylcholine/phosphatidylglycerol liposomes

The Ca 2+-dependent interaction of various polyanionic polysaccharides (chondroitin sulfate, heparin, dextran sulfate, β-cyclodextrin sulfate, hyaluronic acid and carboxymethyldextran) with multilamellar dimyristoyl phosphatidylcholine (DMPC) liposomes was investigated by calorimetric and fluorescence spectroscopic measurements. It was found that an observed polysaccharide-induced phospholipid phase separation depends on the density of the sulfate groups along the polysaccharide chain independent of the presence of additional carboxyl groups. The phase separation resulting from the drastic dehydration of the covered membrane regions is monitored by the upward shift of the lipid phase transition and by the blue shift of the emission spectrum of a headgroup-dansylated phosphatidylethanolamine (DPE). This shift is only observable if the required polysaccharide chain length contains at least three glycosyl units. The Ca 2+-mediated interaction of dextran sulfate with various phosphatidylcholines, differing in their compressibility, showed the maximal difference between the phase transition temperatures of the lipid phase covered by the polysaccharide and the uneffected lipid domains for dielaidinoyl phosphatidylcholine (DEPC), the most compressible phospholipid investigated here. Mixed negatively charged DMPC/dimyristoyl phosphatidylglycerol (DMPG) liposomes were found to compete with the likewise negatively charged dextran sulfate for the binding of Ca 2+. At excess Ca 2+ concentrations, the binding of the polysaccharide was strengthened, compared to pure DMPC liposomes. The monovalent cation sodium, was able to inhibit the interaction between the membrane surface and dextran sulfate. Various divalent cations were found to mediate the interaction, depending on their ionic radii and electron configuration. Within the second group of the periodic system Ca 2+ is the most effective ion. However, within the horizontal forth period the ability to bind sulfated dextran to membrane surfaces decreases from Ca 2+ to Ni 2+, but then increases again if Cu 2+ or Zn 2+ was used as the mediating ion.

Immobilization of pectin, xyloglucan and other soluble plant polysaccharides on blotting membranes.

Soluble polysaccharides, of natural and synthetic origins, are immobilized by nylon, nitrocellulose, and polyvinylidene difluoride blotting membranes. Retention of an acid and a neutral polysaccharide, measured using radioactive pectin and xyloglucan-rich hemicellulose, exceeded 90% on some blotting membranes eluted in aqueous media. The two polysaccharides displayed different binding characteristics. Several polyanionic uronic acid polysaccharides, bound strongly to nylon 66 (Hybond N, Amersham) and to positively charge-modified or cationic nylon blotting membranes (Hybond N +, Amersham), but much less strongly to paper, nitrocellulose or polyvinylidene difluoride. Fragments of pectin, including Rhamnogalacturonan I and Rhamnogalacturonan II, and galacturonic acid oligosaccharides as small as the dimer also bind to nylon membranes, and can be detected using cationic dyes or by means of their radioactivity. Xyloglucan-rich hemicellulose in general binds more strongly than pectin to the same substrates. Of the substrates tested charge-modified nylon gave best retention of pectin, and paper gave best retention of xyloglucan during washes in water and solutions of salts, acids and bases. The influence of pH and solutions of mono, di- and trivalent salts on the retention of some pectic polysaccharides by nylon was investigated. A large proportion, in excess of 70% of pectin applied to charge-modified nylon, remained tightly bound at all salt concentrations up to 2 M. Soluble acid polysaccharides immobilized on blotting membranes could be detected by staining with cationic dyes, such as ruthenium red, alcian blue 8GX, and coriphosphine O, providing facile detection and a simple means of characterizing the cytochemical specificity of their staining reactions. Immobilized neutral polysaccharides, which do not react with cationic dyes, could usually be detected by the periodate-Schiff reaction, or, if labelled with radionuclides, by autoradiography or scintillation counting. Soluble polysaccharides, like proteins and nucleic acids, may therefore be immobilized on blotting membranes for investigation with cytochemical, immunocytochemical, and other molecular probing and detection procedures. Quantitative binding data showed marked differences in the affinity of different polysaccharides for blotting substrates. Detailed characterization of the binding behaviour is therefore a prerequisite for optimization and rational application of polysaccharide blotting.

Function of pulmonary M2 muscarinic receptors in antigen-challenged guinea pigs is restored by heparin and poly-L-glutamate.

The effect of heparin and poly-L-glutamate on the function of inhibitory M2 muscarinic autoreceptors on parasympathetic nerves in the lung was tested in antigen-challenged guinea pigs. After antigen challenge, M2 receptor function is decreased, thus increasing release of acetylcholine from the vagus and potentiating vagally induced bronchoconstriction. Guinea pigs were anesthetized, tracheostomized, vagotomized, paralyzed, and ventilated. Electrical stimulation of the vagi caused bronchoconstriction and bradycardia. In controls, pilocarpine attenuated vagally induced bronchoconstriction by stimulating neuronal M2 muscarinic receptors. Conversely, blocking these autoreceptors with gallamine potentiated vagally induced bronchoconstriction. In challenged animals the effects of both drugs were markedly reduced, confirming M2 receptor dysfunction. 20 min after heparin or poly-L-glutamate, the effects of both pilocarpine and gallamine on vagally induced bronchoconstriction were restored, demonstrating recovery of M2 receptor function. Neither heparin nor poly-L-glutamate affected vagally induced responses in control animals. Thus antigen-induced dysfunction of M2 receptors can be reversed by polyanionic polysaccharides (heparin) or polyanionic peptides (poly-L-glutamate). This suggests that a polycationic substance such as eosinophil major basic protein, cationic protein, or peroxidase may be responsible for antigen-induced pulmonary M2 receptor dysfunction.

Ultrastructural characterization of an extracellular matrix produced by Leishmania mexicana pifanoi

In a previous study it was shown that Leishmania braziliensis promastigotes secreted metabolic products to the culture media. These metabolites called “excreted factors” were chemically characterized as polysaccharides and glycoproteins that reacted with antileishmania sera. These observations together with the recent characterization of lipophosphoglycans and phosphoglycan lead us to investigate by ultrastructural procedues if leishmania is able to produce an extracellular matrix, in axenic culture conditions. In this study, Leishmania mexicana were cultured in minimum essential medium, supplemented with 10% fetal calf serum, at 22°C. Under these conditions, the parasites tend to associate forming small aggregates, called “rosettes” in which the flagellum is oriented inwards and the cell body outwards (Fig.1). In order to retain their association I supposed the existance of an adhesive substance that kept the parasites together. To test this possibility, rosettes were fixed with glutaraldehyde in cacodylate buffer 0.1 M, or with glutaraldehyde containing 1 mg of ruthenium red, that binds to glycoproteins or polyanionic polysaccharides of extracellular matrices.

Dextran sulfate blocks antibody binding to the principal neutralizing domain of human immunodeficiency virus type 1 without interfering with gp120-CD4 interactions

The mechanism of the antiviral activity of sulfated polysaccharides on human immunodeficiency virus type 1 (HIV-1) was investigated by determining the effect of dextran sulfate on the binding of CD4 and several anti-gp120 monoclonal antibodies to both recombinant and cell surface gp120. Dextran sulfate did not interfere with the binding of sCD4 to rgp120 on enzyme-linked immunosorbent assay (ELISA) plates or in solution and did not block sCD4 binding to HIV-1-infected cells expressing gp120 on the cell surface. Dextran sulfate had minimal effects on rgp120 binding to CD4+ cells at concentrations which effectively prevent HIV replication. In contrast, it potently inhibited the binding of both rgp120 and cell surface gp120 to several monoclonal antibodies directed against the principal neutralizing domain of gp120 (V3). In an ELISA format, dextran sulfate enhanced the binding of monoclonal antibodies against amino-terminal regions of gp120 and had no effect on antibodies directed to other regions of gp120, including the carboxy terminus. The inhibitory effects of polyanionic polysaccharides on viral binding, viral replication, and formation of syncytia therefore appear mediated by interactions with positively charged amino acids concentrated in the V3 region. This high local positive charge density, unique to the V3 loop, leads us to propose that this property is critical to the function of the V3 region in mediating envelope binding and subsequent fusion between viral and cell membranes. The specific interaction of dextran sulfate with this domain suggests that structurally related molecules on the cell surface, such as heparan sulfate, may be additional targets for HIV binding and infection.

Polyclonal activation of rat B cells. II. Dextran sulfate as a cofactor in mitogen-induced and antigen-induced differentiation of rat B lymphocytes.

Salmonella typhimurium mitogen (STM) is a polyclonal activator of rat B lymphocytes, triggering them to proliferate, but not differentiate, to antibody-secreting cells. When lymphokines in the form of a supernatant from Con A-stimulated splenocytes (CAS) are added to B cell cultures activated by STM, only a small number of cells are driven to differentiate. Only with the addition of a third signal provided by the polyanionic polysaccharide dextran sulfate (DXS) is significant rat B cell differentiation observed. In this study, we have shown that this requirement for DXS is not unique to the STM mitogen. LPS, Staphylococcus aureus Cowan I-fixed cells, and anti-Ig antibody all induced rat B cell proliferation with little differentiation, even in the presence of CAS. DXS was necessary to induce differentiation in all cultures costimulated with mitogen and CAS. The requirement for DXS for optimal B cell differentiation is also observed with other lymphokine preparations such as the supernatants from PMA-stimulated EL-4 cells and PHA-stimulated human T cells. Furthermore, this augmentative effect of DXS in rat B cell differentiation was not confined to polyclonal activation systems. Ag-specific IgG secretion was also increased when DXS was added to Ag and CAS costimulated cultures of B cells harvested from the draining lymph nodes of rats immunized with DNP-keyhole limpet hemocyanin. Within the polyclonal activation system, a method of staged additions of STM, DXS, and CAS to B cell cultures was used to investigate the role of DXS during B cell differentiation. Optimal differentiation occurred only when DXS was present in the B cell cultures in conjunction with CAS. The augmentation in differentiation seen with DXS did not appear to be due to the recruitment of an additional CAS-responsive B cell subset, because cycling, low density B cell blasts showed large increases in IgM secretion with subsequent exposure to DXS and CAS. These studies suggest tha DXS acts as a cofactor to various differentiation factors, augmenting polyclonal and Ag-specific rat B cell differentiation. The relevance of DXS to in vivo immune responses is discussed.

Sulphoevernan, a polyanionic polysaccharide, and the narcissus lectin potently inhibit human immunodeficiency virus infection by binding to viral envelope protein

Sulphoevernan is a sulphated alpha-1----3, 1----4 polyglucan (Mr 20,000) with a helical structure. This compound effectively inhibits both human immunodeficiency virus type 1 (HIV-1) and type 2 infection of cells in vitro at concentrations around 0.5 micrograms/ml. Moreover, the compound completely inhibits HIV-1-induced syncytium formation at a concentration of 1 microgram/ml. Competition experiments with 35S-labelled sulphoevernan revealed that the mannose-specific lectin from Narcissus pseudonarcissus prevented binding of sulphoevernan to HIV-1, whereas the antibody OKT4A did not reduce the amount of sulphoevernan bound to MT-2 cells. These data indicate that the non-cytotoxic polymer sulphoevernan binds to the virus rather than to the host cell. In vivo studies, using Rauscher leukaemia virus in NMRI mice, revealed that, at a daily dose of 20 mg/kg, the animals were protected against virus-induced increases in spleen weight. From these in vitro and in vivo data we conclude that sulphoevernan has potential in the treatment of acquired immunodeficiency syndrome.

Carrageenan stimulates reduction of nitroblue tetrazolium by human neutrophils without membrane depolarization, myeloperoxidase secretion, or increased oxygen consumption.

Carrageenan, a sulfated polyanionic polysaccharide, is commonly used to induce inflammation in experimental animals, and this model is used to screen for the effectiveness of antiinflammatory drugs. Carrageenan-induced inflammation has been attributed to a variety of autocoids including histamine, bradykinin, complement, superoxide, and prostaglandins. This study examines the effects of carrageenan on human PMN in a serum-free system. Carrageenan was found to stimulate the reduction of NBT by PMNs without stimulating membrane depolarization, oxygen consumption, H2O2 production, or myeloperoxidase secretion. Carrageenan stimulates a heat-labile, NBT-reducing system which is unassociated with the usual stimulus-response coupling seen with other PMN activators such as PMA, FMLP, and zymosan.

Influence of nutrient limitation and low pH on serogroup B Neisseria meningitidis capsular polysaccharide levels: correlation with virulence for mice.

Neisseria meningitidis strain M986, which possesses a polyanionic sialic acid capsular polysaccharide, was resistant to the bactericidal effects of normal rabbit serum, but sensitive when immune serum and complement were present. An isogenic strain PRM102, deficient in the ability to produce capsular polysaccharide, was sensitive to normal serum. Strain M986, when grown under conditions of low pH or nutrient limitation, synthesized increased levels of capsular polysaccharide. This was accompanied by an increase in cell surface hydrophilicity and virulence for mice. Cells grown in low-pH, iron-limited medium synthesized the highest concentration of polysaccharide and exhibited the highest cell surface hydrophilicity and virulence among the cases examined. The increase in capsular polysaccharide was partly explained by a decrease in the specific activity of a membrane-bound cytidine monophosphate-N acetylneuraminic acid hydrolase. The results suggest that conditions of nutrient limitation and low pH exert profound effects on the physicochemical nature of the meningococcal cell surface which, in turn, cumulate in enhanced virulence of this organism for mice.

Substratum requirements for bacterial gliding motility

Flexibacter FS-1 filaments are unable to glide on agarose gels or on charge-neutralized glass unless those substrata are supplemented with the charged, cold water-soluble fraction of agar, or a variety of polyanionic polysaccharides derived from plants, yeasts and bacteria. Two graft polymers of xanthan gum also promote gliding motility under these conditions, as do an extracellular product of the bacteria themselves. A number of other polymers and small molecular species are ineffective as supplements. These results are considered in the context of the soil habitat of this Flexibacter. Among a variety of other gliding bacteria tested, several strains of the order Cytophagales also were unable to glide on agarose.

The development of a cell-substrate attachment system in a euglenoid flagellate

The transition of the euglenoid Colacium libellae from the free-swimming flagellated to the nonflagellated stage attached by a stalk occurs in three distinct stages: (1) initial attachment, perhaps by the flagellum, (2) substrage contact by the anterior region of the cell concomitant with attachment disc formation, and (3) the extrusion of stalk material. The flagellum is resorbed within minutes of initial attachment. Polysaccharide and acid phosphatase derived from within intracellular cisternae initially accumulate in the reservoir, then transfer to the canal and finally become incorporated into the stalk, thus delineating the source and routte of transport for most stalk material. Indeed, the inner “shaft” and central “core” of the stalk contain a neutral polysaccharide that is continuous with similar material found in the reservoir and canal. The peripheral region of the stalk (the “cortex”), however, contains a polyanionic polysaccharide which may be extruded by a special secretory system composed of a ring of granules and canals located beneth the anterior pellicle of the organism.

Mitogenic response of C3H/HeJ mouse lymphocytes to polyanionic polysaccharides obtained from Bordetella pertussis endotoxin and from other bacterial species.

Lipopolysaccharide extracted from Bordetella pertussis was mitogenic for spleen cells of endotoxin-resistant C3H/HeJ mice. Although endotoxic lipid A was inactive, mitogenic activity of lipopolysaccharide was exhibited by purified preparations of polysaccharides I and II, which constitute the carbohydrate moiety of the macromolecule. These low-molecular-weight (2,800 and 3,600) polysaccharides, containing carboxyl groups, were not mitogenic for thymocytes and splenic T-cells of C3H/HeJ mice, but did show mitogenic activity for splenic B-cells of C3H/HeJ mice and for spleen cells of C57BL/6 athymic nude mice. The mitogenic activities of polysaccharides I and II were also compared with those of other polyanionic polysaccharides, and the results indicate that high molecular weight is not necessary, and negative charges are not sufficient, for mitogenicity.

Radiation induced diffusion controlled reactions: A probe for enhanced solute diffusion in polysaccharide matrices

Rate constants for the reaction of the hydrated electron, e - aq with certain solutes, which are diffusion controlled in normal aqueous solution, have been investigated using pulse radiolysis techniques in binary solvent systems of different viscosities. The same reactions were studied in various polysaccharide matrices, such as viscous polyanionic sols of kappa (к) carrageenan and sodium carboxymethylcellulose (CMC), and also polyanionic and neutral gels of к-carrageenan and agarose. The vastly enhanced viscosities do not reduce the reaction rates, and in certain instances, significant enhancement of rates is observed. A similar behaviour was encountered with the unimolecular decomposition of the cystamine (RSSR) −, anion radical, formed by reaction of the hydrated electron with cystamine, and also for the electron transfer reaction between the cystamine anion radical and oxygen to form the superoxide anion radical. The results can best be understood in terms of facilitated solute movement rather than Donnan equilibria. Facilitated solute movement is a phenomenon noted in certain types of polyanionic polysaccharide matrix, where enhanced diffusion of both ionic and neutral small molecular species is observed relative to that in a matrix of agarose.